Hypothesis

4,133 Matching Annotations

Jul 2020
niklasblog.com niklasblog.com

Obsidian: my new knowledge base or base for knowledge

1
1. pivic 09 Jul 2020
  
  in Public
  
  Nefarious corporations
  
  As somewhat detailed here.
  
  For far more information I strongly recommend reading Shoshana Zuboff's The Age of Surveillance Capitalism.
  
  shoshana zuboff surveillance capitalism book microsoft google facebook amazon
Visit annotations in context

Tags

shoshana zuboff

surveillance capitalism

google

facebook

book

amazon

microsoft

Annotators

pivic

URL

niklasblog.com/

SciScore for 10.1101/2020.07.07.20148106: (What is this?)

Please note, not all rigor criteria are appropriate for all manuscripts.

Table 1: Rigor

<table><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Institutional Review Board Statement</td><td style="min-width:100px;border-bottom:1px solid lightgray">Written informed consent obtained from all participants in this study and was approved by the following IRBs: 1 ) IRB# SUNY:269846 .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Randomization</td><td style="min-width:100px;border-bottom:1px solid lightgray">In this regard , it is interesting to note that a bruton tyrosine kinase ( BTK ) inhibitor , that targets Fc-receptor signaling in macrophages , is being tested in a randomized clinical trial 32 .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Blinding</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Power Analysis</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Sex as a biological variable</td><td style="min-width:100px;border-bottom:1px solid lightgray">Subdividing the subjects by sex did not reveal any statistical difference in IgG levels at any of the disease stages , although hospitalized females in the non-ICU setting had significantly lower antibody levels than ICU/deceased patients , whereas the difference in males was not significant ( Fig . 2f) .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Cell Line Authentication</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr></table>

Table 2: Resources

<table><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2">Antibodies</td></tr><tr><td style="min-width:100px;text=align:center">Sentences</td><td style="min-width:100px;text-align:center">Resources</td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">CoV-2 Spike protein or Nucleocapsid protein specific IgG antibodies at titers more than 1:100,000 were detectable in all PCR+ subjects (n=87) and were absent in the negative controls.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>CoV-2 Spike protein or Nucleocapsid protein specific IgG</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Other isotype antibodies (IgA, IgG1-4) were also detected.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>IgA, IgG1-4</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">CoV-2 infection2-6 To predict protection against CoV-2, it is critical to understand the quantity, quality and duration of the antibody responses during different stages of COVID-19 and in the convalescent period.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>CoV-2</div> <div>suggested: (Abcam Cat# ab272504, AB_2847845)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">In this assay, we immobilized biotinylated CoV-2 Spike protein receptor binding domain (RBD) or the Nucleoprotein (N) on streptavidin beads, to detect specific antibodies from patient plasma (Fig.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>CoV-2 Spike protein receptor binding domain (RBD)</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Different antibody isotypes were measured using anti-Ig (IgG, IgA, IgM) specific secondary antibodies conjugated to a fluorescent tag (Fig. 1a).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-Ig ( IgG</div> <div>suggested: None</div> </div>

      <div style="margin-bottom:8px">
        <div><b>IgA , IgM</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Using either anti-SRBD antibody or soluble ACE2-Fc, we show very high sensitivity in detecting Spike protein binding, down to picogram ranges (Fig. 1b).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-SRBD</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Furthermore, Nucleocapsid protein-specific IgG levels and S-RBD specific IgA positively correlated with S-RBD IgG antibodies (Supplementary Fig. 1b, c).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>S-RBD IgG</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Notably, IgG1 subclass antibody levels were comparable to total IgG levels whereas the other subtypes were relatively lower (Fig. 2b).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>IgG1</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">To evaluate membrane expression of Spike protein, cells were stained with recombinant soluble ACE2-Fc fusion protein followed by a secondary staining with an anti-Fc antibody (Fig 3a).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-Fc</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">ACE2 overexpression of ACE2-IRES-GFP or ACE2mKO2 was confirmed by staining with CoV-2 Spike-protein fused with mouse Fc (mFc) and antimFc secondary antibody (Supplementary Fig. 2a, b).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>antimFc</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Of note, there was a significant negative correlation between the number of days and the IgG or IgA to S-RBD, anti-nucleocapsid IgG or the NT50 values ( !" = -0.67) (Fig. 6d), suggesting a potential decline in antibody titers over time.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-nucleocapsid IgG</b></div>
        <div>suggested: (Imported from the IEDB Cat# 3E9, <a href="https://scicrunch.org/resources/Any/search?q=AB_2848062">AB_2848062</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Neutralization of the virus by antibodies (NAbs) is one of the goals to achieve protection against CoV-218.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>CoV-218</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">However, another study showed IgA antibodies, but not IgG, increased in severe patients28.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>IgA</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Although it will be important to follow the same individual subject convalescent plasma over time to better assess this finding, our data point towards a relatively short-lived antibody response to COVID-19.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>COVID-19</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">WT and ACE2 over-expressing HEK-293T were also stained with SARS-CoV-2 S1 protein, Mouse IgG2a Fc Tag (Acro Biosystems) followed with APC Goat anti-mouse IgG2a Fc Antibody (Invitrogen).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Mouse IgG2a</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Anti-S-RBD antibody and ACE2-Fc was tested both at 5 µg/mL starting concentration and in additional 5-fold serial dilutions.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Anti-S-RBD</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Pseudotype virus neutralization assay Three-fold serially diluted monoclonal antibodies including anti-SARS-CoV-2 Neutralizing human IgG1 Antibody from Acro Biosystems, NAb#3 (Fig 4D), Genscript clone ID 6D11F2, NAb#2 (Fig 4D) and Genscript clone ID 10G6H5, NAb#1 (Fig 4D)</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-SARS-CoV-2</b></div>
        <div>suggested: (Abcam Cat# ab272854, <a href="https://scicrunch.org/resources/Any/search?q=AB_2847844">AB_2847844</a>)</div>
      </div>

      <div style="margin-bottom:8px">
        <div><b>human IgG1</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Percent infection obtained was normalized samples derived from cells infected with CoV-2 or SARS pseudotyped virus in the absence of plasma, ACE2-Fc or monoclonal antibodies.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>ACE2-Fc</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">In addition, we also developed SARS Spike protein pseudotyped lentivirus, which similarly infected 293-ACE2 cells at almost 100% efficiency at higher virus supernatant volumes (Fig. 3f)</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>293-ACE2</b></div>
        <div>suggested: <a href="https://scicrunch.org/resources/Any/search?q=CVCL_DR94">CVCL_DR94</a></div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">HEK-293T cells (ATCC; mycoplasma-free low passage stock) were transfected with the expression plasmids using Lipofectamine 3000 (Invitrogen) according to the manufacturer’s protocol as previously described33.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>HEK-293T</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Generating human ACE2 over-expressing cells Wildtype ACE2 sequence was obtained from Ensembl Gene Browser (Transcript ID: ENST00000252519.8) and codon optimized with SnapGene by removing restriction enzyme recognition sites necessary for subsequent molecular cloning steps, preserving the amino acid sequence.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Ensembl Gene Browser</b></div>
        <div>suggested: None</div>
      </div>

      <div style="margin-bottom:8px">
        <div><b>SnapGene</b></div>
        <div>suggested: (SnapGene, <a href="https://scicrunch.org/resources/Any/search?q=SCR_015052">SCR_015052</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Flow cytometry data were analyzed using FlowJo (BD biosciences).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>FlowJo</b></div>
        <div>suggested: (FlowJo, <a href="https://scicrunch.org/resources/Any/search?q=SCR_008520">SCR_008520</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Statistical analyses were performed using GraphPad Prism 8.0 software (GraphPad Software)</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>GraphPad Prism</b></div>
        <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div>
      </div>

      <div style="margin-bottom:8px">
        <div><b>GraphPad</b></div>
        <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Author contributions M.D., L.K. and D.U. conceived, designed the experiments. M.D., L.K., L.P., M.Y. and R.H. carried out the experiments. B.T.L. designed the clinical research study on UConn Healthcare workers and M.K. recruited participants and executed clinical protocols. R.G. and O.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>UConn Healthcare</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr></table>

About SciScore

SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.

Visit annotations in context

Annotators

sciscore

URL

medrxiv.org/cgi/content/10.1101/2020.07.07.20148106

laulima.hawaii.edu laulima.hawaii.edu

someTitle

1
1. sr63 08 Jul 2020
  
  in Public
  
  EI tag c
  
  I wasn't fully grasping TEI and this site helped: https://tei-c.org/about/frequently-asked-questions/
Visit annotations in context

Annotators

sr63

URL

laulima.hawaii.edu/access/content/group/MAN.96998.202040/readings/A_Companion_to_Digital_Humanities_----__A_COMPANION_TO_DIGITAL_HUMANITIES_.pdf
twobithistory.org twobithistory.org

Whatever Happened to the Semantic Web?

1
1. theresia 04 Jul 2020
  
  in Public
  
  As a result, search engines came to ignore the <meta> tag in favor of using complex algorithms to analyze the actual content of a webpage.
  
  this is why we can't have nice things.
  
  human, spams, selfishness
  
  metadata x PageRank == CV x trial period == the dialogue x subtext == what they say x what they do
Visit annotations in context

Annotators

theresia

URL

twobithistory.org/2018/05/27/semantic-web.html
www.dndbeyond.com www.dndbeyond.com

The Village of Barovia

1
1. jromney 03 Jul 2020
  
  in Public
  
  E3. Mad Mary’s Townhouse A moaning sob floats through the still, gray streets, coloring your thoughts with sadness. The sounds flow from a dark, two-story townhouse. The house, which is about 40 feet square, is boarded up and barricaded from the inside. Mad Mary (CN female human commoner) sits in the center of the floor in an upstairs bedroom, clutching a malformed doll. She is lost in her sorrow and despondency. She barely recognizes the presence of anyone in the room. She says nothing in the presence of anger, but she will talk, albeit haltingly, to someone who talks with her gently. Mary hid her beloved daughter, Gertruda, in this house for the girl’s entire life. Gertruda, now a teenager, broke out of the house a week ago and has not been seen since. Her mother fears the worst — and is justified in doing so. See area K42 in chapter 4 for more information on Gertruda’s fate. The malformed doll has a strange leer and wears a sackcloth dress. It belonged to Mary in her youth and was passed down to Gertruda. Gadof Blinsky, the toymaker of Vallaki (see chapter 5, area N7), made the doll. Stitched into the hem of its dress is a frayed tag bearing the words “Is No Fun, Is No Blinsky!”
  
  Cut Mad Mary
Visit annotations in context

Annotators

jromney

URL

dndbeyond.com/sources/cos/the-village-of-barovia
via3.hypothes.is via3.hypothes.is

I Confess, Dec 1 1923

1
1. machadorobert 02 Jul 2020
  
  in Public
  
  behind
  
  big behind
  
  dog tag
Visit annotations in context

Tags

dog tag

Annotators

machadorobert

URL

via3.hypothes.is/proxy/static/https://www.research.thelab.blue/wp-content/uploads/2020/07/Thrills-Thirty-Years-Ago-TEST.pdf
www.medrxiv.org www.medrxiv.org

https://doi.org/10.1101/2020.05.31.20118554

1
1. sciscore 01 Jul 2020
  
  in Public
  
  SciScore for 10.1101/2020.05.31.20118554: (What is this?)
  Please note, not all rigor criteria are appropriate for all manuscripts.
  Table 1: Rigor
  <table><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Institutional Review Board Statement</td><td style="min-width:100px;border-bottom:1px solid lightgray">Let 's study immune responses , but let 's not create a dystopian society based on them . Materials and Methods Human specimens and data All experiments and analyses involving samples from human donors were conducted with the approval of the local ethics committee ( KEK-ZH-Nr . 2015-0561 , BASEC-Nr . 2018-01042 , and BASEC 2020-00802) , in accordance with the provisions of the Declaration of Helsinki and the Good Clinical Practice guidelines of the International Conference on Harmonisation .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Randomization</td><td style="min-width:100px;border-bottom:1px solid lightgray">To directly validate our method , we selected 210 high scoring samples and 122 random samples from known negatives and aimed to reproduce our results .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Blinding</td><td style="min-width:100px;border-bottom:1px solid lightgray">A blinded comparison with commercial test kits showed that our approach – combining three individual assays into one single score – was suitable for large-scale epidemiologic studies .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Power Analysis</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Sex as a biological variable</td><td style="min-width:100px;border-bottom:1px solid lightgray">Lastly , seropositivity can be found across all age groups and in both genders , with more male individuals affected in the USZ and BDS cohorts ( Fig . 3A , B , Table S1) .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Cell Line Authentication</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr></table>
  Table 2: Resources
  <table><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2">Antibodies</td></tr><tr><td style="min-width:100px;text=align:center">Sentences</td><td style="min-width:100px;text-align:center">Resources</td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Solution-equilibrium measurements revealed immunodominant antibodies with nanomolar affinity in COVID samples, whereas prepandemic plasma showed lower affinities despite similar titers for individual SARS2 antigens.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>SARS2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">However , those with strong binding properties to SARS2 RBD ( > 2.5 ) cluster at high values for SARS1 RBD , indicating that some anti-SARS2 RBD antibodies are likely cross-reactive to SARS1 RBD .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-SARS2 RBD</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Anti-his-tag antibody was included as a positive control .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>Anti-his-tag</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">In a comparative approach , we investigated IgG and IgA antibodies to S , RBD , and NC as well as responses to multiple control antigens , in four asymptomatic blood donors and 4 convalescent individuals recruited to the BDS for a plasmapheresis study .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>IgA</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Counter screening using commercial and custom-designed platforms We used the following commercial tests for the detection of anti-SARS-SARS2 antibodies in 56 plasma samples of 27 patients who were diagnosed by RT-PCR to be infected by SARS-SARS2 as well as 83-90 plasma samples which were collected before December 2019 and , hence , before the start of the COVID19 pandemics: The double-antigen sandwich electro-chemiluminescence immunoassay from Roche diagnostics ( Rotkreuz , Switzerland ) was performed with the E801 of the COBAS8000® system ( Roche diagnostics , Rotkreuz , Switzerland) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-SARS-SARS2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The test detects any antibody against the nucleocapsid antigen .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>antibody against the nucleocapsid antigen .</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Baseline and plateau values are fixed by the respective positive and negative controls in a plate-wise fashion and the signal is fitted following these equations: 1 = 1 − ( + + 1 − √ ( + )2 + 2 ( − ) + 1 ) , 2 where cbound , ca and c are concentration of the antigen-antibody , antigen , and blood concentration respectively .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>+ + 1 − √ ( + )2 + 2 ( − ) + 1 ) , 2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Assume that we have data for samples with known serostatus and antibody measurements , that is , we have ( , ) , = 1 , . . , , where is the vector of size ( in our case our antigen measurements ) and is a Boolean variable defining group membership ( in our case , whether the individual is seropositive or not) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>we have ( , ) , = 1 , . . ,</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The day after, membranes were washed four times with PBS-T and incubated for 1 hours with an anti-human secondary antibody, HRP-conjugated, diluted 1:10000 in 1% SureBlock.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-human secondary antibody, HRP-conjugated,</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">As a positive control, one membrane was incubated overnight with mouse anti-Histag antibody (ThermoFisher, dilution 1:10000 in 1% SureBlock) and subsequently with anti-mouse secondary antibody, HRP-conjugated (Jackson, dilution 1:10000 in 1% SureBlock).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-Histag</div> <div>suggested: (RevMAb Biosciences Cat# 54-1161-00, AB_2716428)</div> </div>
  
  <div style="margin-bottom:8px"> <div><b>anti-mouse</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Then, the plates were washed five times with PBS-T and the presence of IgGs was detected using an HRP-linked anti-human IgG antibody (Peroxidase AffiniPure Goat Anti-Human IgG, Fcγ Fragment Specific, Jackson, 109-035-098, at 1:4000 dilution in sample buffer).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Anti-Human IgG</b></div> <div>suggested: (Jackson ImmunoResearch Labs Cat# 109-035-098, <a href="https://scicrunch.org/resources/Any/search?q=AB_2337586">AB_2337586</a>)</div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">We confirmed these findings by using the samples of the asymptomatic and convalescent individuals as primary antibodies in Western Blot and detected bands for both S and the NC in the Expi293 cells overexpressing the viral proteins but not in the Expi293 control lysate ( Fig . 5B) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Expi293</b></div> <div>suggested: <a href="https://scicrunch.org/resources/Any/search?q=CVCL_D615">CVCL_D615</a></div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">To benchmark TRABI, we compared the results with an in-house high-throughput assay under development at the University of Oxford (optimizations ongoing at the time of data acquisition), the Roche Elecsys, the DiaSorin, the EuroImmun, and the Abbott systems (Fig. 1C), using 139 of 149 samples (10 were removed from the analysis because of insufficient sample volume to perform all tests).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Abbott systems</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">COVID and prepandemic samples were used to assess the performance of TRABI, commercial tests (Roche, DiaSorin, Abbott, Euroimmun) and an early version of an assay under development at the Target Discovery Institute (Oxford).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Abbott</b></div> <div>suggested: (Abbott, <a href="https://scicrunch.org/resources/Any/search?q=SCR_010477">SCR_010477</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Details of viral proteins used for this study For high-throughput serology , the following proteins were used: SARS-CoV-2 S ( pHL-Sec; aa . 11208 , C-terminal 8His-Twin-Strep ) and RBD ( pOPINTTGNeo; aa . 330-532 , C-terminal 6His ) produced at the SGC in Oxford and the nucleocapsid protein from AcroBiosystems ( AA Met 1 - Ala 419 , C-terminal his-tag , NUN-C5227) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>AcroBiosystems</b></div> <div>suggested: (ACRObiosystems, <a href="https://scicrunch.org/resources/Any/search?q=SCR_012550">SCR_012550</a>)</div> </div> </td></tr></table>
  
  Results from OddPub: We did not find a statement about open data. We also did not find a statement about open code. Researchers are encouraged to share open data when possible (see Nature blog).
  About SciScore
  
  SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.
Visit annotations in context

Annotators

sciscore

URL

medrxiv.org/cgi/content/10.1101/2020.05.31.20118554
www.biorxiv.org www.biorxiv.org

https://doi.org/10.1101/2020.06.12.148726

1
1. sciscore 01 Jul 2020
  
  in Public
  
  SciScore for 10.1101/2020.06.12.148726: (What is this?)
  Please note, not all rigor criteria are appropriate for all manuscripts.
  Table 1: Rigor
  <table><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Institutional Review Board Statement</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Randomization</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Blinding</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Power Analysis</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Sex as a biological variable</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Cell Line Authentication</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr></table>
  Table 2: Resources
  <table><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2">Antibodies</td></tr><tr><td style="min-width:100px;text=align:center">Sentences</td><td style="min-width:100px;text-align:center">Resources</td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">To independently confirm that similar number of virions was analyzed, the lower part of the same membrane was blotted with an anti-p30 MLV gag antibody (Fig. 2c).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-p30 MLV gag</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The M2 antibody used in this experiment binds the Flag tag located at both the N- and C-termini of a protein, but it binds N-terminal Flag tag more efficiently33.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>M2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">To differentiate these possibilities, we appended the Myc-tag to the N-terminus of the S-protein that is Flag tagged at its C-terminus and repeated the study, this time detecting the S1 domain with an anti-Myc antibody.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-Myc</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">For western blot analyses, 5-10 µl of purified PV, which is equivalent to 0.5-1.0 x 1010 vector genomes, was loaded per lane of the 4-12% Bis-Tris gel (Life Technologies), transferred to the PVDF membrane, and blotted with 1 µg/ml anti-Flag M2 antibody (Sigma-Aldrich, F1804) to detect the S-protein bands.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-Flag</div> <div>suggested: (Sigma-Aldrich Cat# F1804, AB_262044)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">1 µg/ml anti-p30 MLV gag antibody (Abcam, ab130757) and 1:10,000 dilution of goat-anti- mouse IgG-HRP polyclonal antibody (Jackson ImmunoResearch, 115-036-062) were used to detect MLV gag protein as an internal control.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>1 µg/ml anti-p30 MLV gag antibody (Abcam, ab130757)</div> <div>suggested: None</div> </div>
  
  <div style="margin-bottom:8px"> <div><b>anti-p30 MLV gag antibody (Abcam, ab130757)</b></div> <div>suggested: None</div> </div> <div style="margin-bottom:8px"> <div><b>mouse IgG-HRP</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">As with MLV PV, the S-protein bands were visualized using the anti-Flag M2 antibody, and the N-protein band was detected using pooled convalescent plasma at a 1:500 dilution and 10 ng/ml goat-anti-human IgG antibody conjugated with polymerized HRP (Fitzgerald, 61RI166AHRP40).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>IgG</b></div> <div>suggested: (Fitzgerald Industries International Cat# 61R-I166AHRP40, <a href="https://scicrunch.org/resources/Any/search?q=AB_10815602">AB_10815602</a>)</div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">We observed PVG614 infected hACE2-293T cells with approximately 9-fold higher efficiency than did PVD614 (Fig. 1c,d).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>hACE2-293T</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">MLV PVs were produced by transfecting HEK293T cells at ~60% confluency in T175 flasks using the calciumphosphate method with 70 µg of total DNA.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>HEK293T</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Mock- and hACE2HEK293T cells on 96-well plates were infected with the preincubation mixes and infection levels were assessed 24 h later by measuring luciferase activity using the LucPair Firefly Luciferase HS Assay Kit (GeneCopoeia).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>hACE2HEK293T</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Genotype frequency at residue 614 was calculated using R (R Foundation for Statistical Computing) with the Biostrings package.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Biostrings</b></div> <div>suggested: (Biostrings, <a href="https://scicrunch.org/resources/Any/search?q=SCR_016949">SCR_016949</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Logo plots of D614G variation were generated by WebLogo after sequence alignment.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>WebLogo</b></div> <div>suggested: (WEBLOGO, <a href="https://scicrunch.org/resources/Any/search?q=SCR_010236">SCR_010236</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">All appropriate data were analyzed with GraphPad Prism 7 (GraphPad Software Inc.).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>GraphPad Prism</b></div> <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div> </div> <div style="margin-bottom:8px"> <div><b>GraphPad</b></div> <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div> </div> </td></tr></table>
  
  Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
  Results from OddPub: Thank you for sharing your data.
  About SciScore
  
  SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.
Visit annotations in context

Annotators

sciscore

URL

biorxiv.org/cgi/content/10.1101/2020.06.12.148726
www.biorxiv.org www.biorxiv.org

ESI mutagenesis: A one-step method for introducing point mutations into bacterial artificial chromosome transgenes

1
1. EMBOpress 01 Jul 2020
  
  in Review Commons
  
  Note: This rebuttal was posted by the corresponding author to Review Commons. Content has not been altered except for formatting.
  
  Learn more at Review Commons
  
  Reply to the reviewers
  
  We thank the reviewers for their comments and outline below how we plan to address them.
  
  Reviewer #1 (Evidence, reproducibility and clarity (Required)): **Summary:** Provide a short summary of the findings and key conclusions (including methodology and model system(s) where appropriate). The authors here describe a method to modify bacterial artificial chromosomes (BAC) harbouring gene loci from eukaryotes. When wanting to modify a BAC an antibiotic selection cassette is often included alongside the desired mutation/modification to increase the number of successful recombinants in E.coli. Traditionally, this is removed in a second recombination process to leave only the desired modification. The novelty in the procedure described herein is to add a synthetic intron consensus sequence around the selection cassette, which eliminates the need for the subsequent removal of the antibiotic cassette from the BAC before transfection into mammalian cells, saving time and resources. The technique is clever in its simplicity and appears to function for a number of gene loci. The authors validated the correct functioning of the modified BACs for a number of genes using three main assays - transcript level, protein level and localisation. **Major comments:** *Are the key conclusions convincing?* The conclusion that the method described generates functional modified BACs is valid. *Should the authors qualify some of their claims as preliminary or speculative, or remove them altogether?* While the method is successfully employed in this study, its efficiency is not quantified in relation to the state-of-the-art as described in the introduction. One assumes it would be more efficient, but this has not been tested empirically in the paper. Does the inclusion of the synthetic intron sequence have an effect on the efficiency of modifying BACs compared to a more typical two-step positive/negative antibiotic selection cassette? *
  
  *
  
  This is a good point that we did not directly address. In general, the efficiency is similar to that of integrating any cassette with selectable marker, as has been published (Poser et al 2008), and therefore also higher than the two-step counterselection method, which requires such a cassette integration in the first step alone. We will include new data specifically addressing the efficiency of our new method (see specifics below)
  
  The functionality of this approach rests entirely on the ability of the target cell to correctly splice out the synthetic intron. The authors are aware of this potential problem as highlighted in the lines below, but do not make efforts to explicitly test splicing. On lines 224-225, the authors state "We cannot exclude that a small portion of synthetic introns within individual cells are misspliced". On lines 230-231 it is stated that "mis-spliced mRNAs are probably minimal and degraded by nonsense-mediated decay". On lines 215-217, the authors describe an "investigation of transgenic lines at the single-cell level" that suggests "the synthetic intron is correctly spliced out in all the cells of the population". How do the authors reach this conclusion? U2OS and HeLa cells are considered very "robust" and may not show detectable consequences when stressed with an increased level of nonsense-mediated decay. Further, many genes maintain a high level of expression that buffers them against small changes in transcription/splicing. The synthetic intron might have a bigger impact on more tightly regulated genes, so assessing the splicing rate would be essential if the authors wish to advocate their technique as generally applicable.
  
  *
  
  We will assay for splicing efficiency as outlined below.
  
  The ability of the synthetic intron to be removed from final transcripts depends on functioning splicing machinery. The authors might emphasise this issue, as spliceosome mutations are important fields of study and might not be compatible with this method.
  
  *
  
  We can add this in the text
  
  The authors used un-directed integration of each BAC under study. Therefore, it is hard to assess what effect the synthetic intron has, as the authors only ever assess the downstream levels of the correctly spliced, translated and localised protein. The authors themselves state that this can lead to clonal variations in expression of up to 2-fold and on line 250 that this variation "could compensate for synthetic intron effects", but make no effort to test this. Again, lines 267-268 highlight the potential dangers of potential effects of the synthetic introns, but do not test these. \Would additional experiments be essential to support the claims of the paper? Request additional experiments only where necessary for the paper as it is, and do not ask authors to open new lines of experimentation.* If not already performed, a large number of bacterial colonies should be screened for the correct modification and frequency of correct ones reported. This frequency - reported for at least three different modifications - would estimate what sort of efficiency this method provides. The modified region of each BAC should be sequenced and the results reported. The rate of exactly modified clones is important, in case of spontaneous or low fidelity integration of the antibiotic cassette. The percentage of transcripts that have the synthetic intron correctly spliced out should be measured for some of the BAC constructs used in the study. A direct head-to-head comparison of this newer method compared to other techniques, or even the authors' own previous two-step approach is necessary to assess the benefits of this method. Preferably, the experiment would be run in parallel with and without antibiotic selection applied, to show that it drastically improves chances of finding a correct clone. *
  
  We will generate 3 new mutations in BACs and analyze both the efficiency of integration by PCR and accuracy via sequencing. In practice, we have observed that the efficiency is similar to any other cassette integration, such as a GFP tag (Poser et al Nature Methods 2008) or a counterselection cassette (Bird et al Nature Methods 2012) (80-90%). Integrating a mutation via the second step of the counterselection method introduces a further 20% decrease in efficiencies on average.
  
  \Are the suggested experiments realistic in terms of time and resources? It would help if you could add an estimated cost and time investment for substantial experiments.* Repeating the transformation of the BAC and targeting cassette and assessing the recombination efficiency and sequencing should only require existing reagents and take less than a week or two to complete. Quantitative RT-PCR to assess the percentage of transcripts that have the synthetic intron spliced out would take a little more work. However, this should not be a considerable investment in time or resources for a standard microbiology laboratory and could be completed within a few weeks using modern techniques, such as that described in Londoño et al. 2016. Repeating all the experiments in parallel would be considerable work and would only be strictly necessary if the authors wish to emphasise the benefits of their method over the many others already in wide use. *
  
  *
  
  We will use quantitative PCR to estimate the fraction of transcripts that correctly splice out the artificial intron for two clonal cell lines characterized in the study: RNAi-resistant AurA-GFP (Fig 4), and GTSE1-14A (newly introduced; see below). While the exact method described in Londoño et al 2016 will not be applicable due to the larger size of the artificial intron, we believe we can adapt it to detect different splicing events.
  
  \Are the data and the methods presented in such a way that they can be reproduced?* Barring the omission of Table S1, which presumably includes exact information on the BACs modified and sequences used etc., there is sufficient other data and methods to allow the experiments to be repeated. Targeting the ESI procedure to the middle of exons is likely to have a bigger impact for smaller exons as the authors mention on lines 99-100. Making it clear which exon sizes for each gene were successfully targeted in this study would help give some idea of how significant a problem this might be. Perhaps Table S1 contains this information, but it was not provided. It would also help reviewers check the design strategies. *
  
  We apologize for inadvertently failing to upload Table S1 on bioRxiv. It has been uploaded now as part of this submission process. This table indeed contains BAC and target sequence information, including the size of the targeted exon (and the 2 “new” resulting exons). Targeted exons range in size from 138bp to 1537bp, and “new” exons are as small as 48bp.
  
  \Are the experiments adequately replicated and statistical analysis adequate?* The replication and statistically analysis of the data as presented appear adequate. Figure Legends should state the statistic used to generate error bars. *
  
  This will be updated
  
  \*Minor comments:** Specific experimental issues that are easily addressable. Are the promoters used in the vectors described universally functional? For example, is the PGK promoter functional in yeast? *
  
  *
  
  The PGK promoter contained in the cassettes is a mammalian promoter, which has also been reported to work in flies.
  
  \Are prior studies referenced appropriately?* The manuscript may benefit from the referencing of BAC modification techniques from a wider variety of groups, such as those using CRISPR-guided recombineering (Pyne et al. 2015). *
  
  We will add citations of more techniques
  
  \Are the text and figures clear and accurate?* The body text is very clear save minor typographical or grammatical errors. Regarding figures, some of the coloured text in Figure 1 is somewhat illegible when printed in grayscale. Line 278 - The acronyms LAP and NLAP are not defined/explained. Antibody section starting Line 282 may fit better next to Western Blot section. Figure 2C - The blot images would benefit from arrows to indicate expected sizes of proteins. Figure 3A - the graph may benefit from a dashed line at 100% to highlight that values are normalised to controls. Figure 4 - The differences between panels B & C are unclear. Figure 4E - The legend could provide a little more detail on cell cycle stage/status of the captured cells. *
  
  All of the above will be addressed accordingly
  
  \Do you have suggestions that would help the authors improve the presentation of their data and conclusions?* Lines 23-27 are somewhat unclear and feel out of context. Perhaps the authors could clarify this as a further advantage of using BACs instead of endogenous gene modifications. *
  
  Thanks for the input, we will clarify this.
  
  While not affecting the factual content of the paper, I would advocate that the authors format the method described in Figure S3 into a more detailed text based layout similar to that seen in a typical Nature Methods article. However, this may depend on the format required by any eventual publishing journal.
  
  *
  
  We prefer the graphical protocol, but will discuss whether to add a text protocol with the journal editor.
  
  That all of the work the paper was carried out in human cell lines and using human genes is a further caveat, but the authors admit this in the discussion and one would assume that most mammalian cells would respond similarly in their ability to splice out the synthetic intron. Reviewer #1 (Significance (Required)): \Describe the nature and significance of the advance (e.g. conceptual, technical, clinical) for the field.* This work is a formal description of a newer method that could be useful for many of those employing bacterial artificial chromosomes in numerous studies, such as gene regulation. *Place the work in the context of the existing literature (provide references, where appropriate).* This work builds on methodology previously published by the authors - a counter-selection two-step procedure (Bird et al. 2011). It sets out to formally describe a method merely mentioned as "BAC intronization" in a later paper by some of the authors (Zheng et al. 2014). Other alternative one-step procedures are also available, but present a different set of challenges (Lyozin et al. 2014). Some newer approaches, such as those using CRISPR-guided recombineering (Pyne et al. 2015) or systems that combine CRISPR and positive/negative selection cassettes (Wang et al. 2016) may be slightly more efficient, but are also more complex in their design. Bird et al. 2011 DOI: 10/dv776q Pyne et al. 2015 DOI: 10/f7jx92 Wang et al. 2016 DOI: 10/f89db5 Zheng et al. 2014 DOI: 10/f5pkr6 *State what audience might be interested in and influenced by the reported findings.* As a technology paper this work should have interest from a broad field of research. While the use of BACs could sometimes be considered more traditional in light of the explosion in CRISPR-based genome editing capabilities, it is definitely seeing a resurgence as the limitations of CRISPR in modifying large regions of genome become more apparent. Therefore, technologies that accelerate the modification of BACs could prove increasingly useful. As category of audience, all those involved in significant recombineering or gene/genome engineering would potentially benefit. *Define your field of expertise with a few keywords to help the authors contextualize your point of view. Indicate if there are any parts of the paper that you do not have sufficient expertise to evaluate.* Synthetic genomics, synthetic biology, cancer cell biology, gene and genome engineering REFEREES CROSS COMMENTING I would agree with reviewer two's assessment that we both view the paper in a similar light. Reviewer #2 (Evidence, reproducibility and clarity (Required)): This is a methods-focused paper that presents a strategy to efficiently introduce mutations into a bacterial artificial transgene using synthetic introns. BAC-based methods have been an effective strategy for introducing trans genes into human cells to achieve near-endogenous expression, including extensive work from these authors. However, generating mutations and changes within the internal coding sequence presents some challenges for how to target these mutations and select for the mutated form. Here, the authors describe a way to overcome this by introducing synthetic introns into an adjacent sequence. This allows them to introduce a selectable marker and conduct the molecular biology without creating complications downstream for the functionality of the protein. This method is carefully described and presented. The authors also provide clear validation by using this to create RNAi-resistant versions of multiple different mitotic factors as well as creating targeted mutants that alter the functional properties of a protein. This work clearly takes advantage of other ongoing studies from these labs (including mutants and cell lines that appear to also have been described elsewhere), but the ability to combine these in a single paper and clearly describe the method provides a helpful advance and validation. Based on the description and data presented, I think that things are clear and carefully validated. As such, I do not have technical comments or concerns and I would be comfortable with this paper appearing in an appropriate journal in its present form. Reviewer #2 (Significance (Required)): This is a solid methods paper, but for considering the nature of the impact and significance of this paper, there are several things to note: 1.The BAC-based method does appear to be a powerful and effective strategy. However, beyond the work of Mitocheck and the authors that are part of this paper, this has not seen widespread adoption. It is possible that this current method may increase its usage due to the value of the targeted mutations within the coding sequence, but at present it is not a broadly used strategy. *
  
  We agree that using BACs as transgenes has not seen widespread adoption as a tool on the broader cell biology community (although certainly beyond members of the Mitocheck consortium). This is likely because many erroneously think that it is a technique for specialist laboratories. We are trying to change this! For reasons outlined below, there is still an increasing desire for conditional analysis of mutated genes under physiological expression/regulation frequently not attainable via directed Cas9-based mutation. A major aim of this paper is thus to further simplify the methods for generating modified BAC transgenes.
  
  2.This BAC-based approach (and also RNAi) are becoming increasingly replaced by the use of CRISPR/Cas9 genome editing. The absence of Cas9-based strategies in this paper limits the potential impact and reach of this paper. The authors do mention the possibility of using a similar synthetic intron strategy for use with Cas9 in the Discussion, and appear to have conducted some experiments. If possible, it would substantially increase the value of this paper if this data and strategy were also included in the Results section (acknowledging that this may still be a work in progress).
  
  While some uses of BAC transgenes are in some cases better replaced by CRISPR/Cas9 techniques (i.e. GFP tagging), there are several occasions where using BACs are preferable: As stated in the text, RNAi-resistant BACs allow for conditional analysis of recessive mutations. Mutations in essential genes that are lethal will prevent growth and recovery of viable cells if integrated into the genome via Cas9. Additionally, deleterious mutations are prone to accumulate suppressive changes in chromosome integrity or gene expression during the procedure of selecting and expanding Cas9-modified cells for analysis, particularly in the genomically instable cancer cell lines frequently employed.
  
  We use both BACs and CRISPR/Cas9 in our lab according to our needs.
  
  We do have an ongoing project to apply this intronization technique to enable more efficient selection of CRISPR/Cas9 integrations. Preliminary results suggest that it works to allow selection of point mutations, but it is still being optimized, including a redesign of the cassette, and is not ready for publication.
  
  3.The method is solid and well-validated, but there are no new results or insights presented in this paper from the work that is described (this is fine, just commenting for considering the right journal fit).
  
  As “biological insights” gained as a result of this technique we had cited a couple studies that made use of the technique already (to functionally analyze a microcephaly-associated mutation in the centriolar protein CPAP at the single cell level in HeLa cells and neural progenitor cells (Zheng et al 2014, Gabirel et al 2016)). As a response to this critique to include “new biology” in this paper, we will add new unpublished data investigating a specific question: Is the cell-cycle-regulated disruption of the EB1-GTSE1 (microtubule plus-end tracking proteins) interaction in mitosis required for chromosome segregation fidelity? We have generated a GTSE1 mutant with 14 phosphosites mutated to alanine using this technique. We will present the effect on chromosome segregation.
  
  REFEREES CROSS COMMENTING It appears that both reviewers are largely on the same page regarding this paper.
  
  PeerReviewed
Visit annotations in context

Tags

PeerReviewed

Annotators

EMBOpress

URL

biorxiv.org/content/10.1101/844282v2

SciScore for 10.1101/2020.06.17.157982: (What is this?)

Please note, not all rigor criteria are appropriate for all manuscripts.

Table 1: Rigor

NIH rigor criteria are not applicable to paper type.

Table 2: Resources

      <div style="margin-bottom:8px">
        <div><b>c-Myc epitope tag ,</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">For library expression experiments , 45 OD units yeast were washed twice with PBS-BSA and labeled in 3mL 1:100 diluted anti-Myc-FITC antibody for 1hr at 4°C with gentle mixing .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-Myc-FITC</b></div>
        <div>suggested: (Sigma-Aldrich Cat# SAB4700448, <a href="https://scicrunch.org/resources/Any/search?q=AB_10896411">AB_10896411</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Antibody epitopes were mapped from crystal structures 6W41 ( Yuan et al. , 2020b) , 6WAQ ( Wrapp et al. , 2020b) , 2DD8 ( Prabakaran et al. , 2006) , 3BGF ( Pak et al. , 2009) , 2GHW ( Hwang et al. , 2006) , 7BZ5 ( Wu et al. , 2020) , and cryo-EM structures 6NB6 and 6NB7 ( Walls et al. , 2019) , and 6WPS ( Pinto et al. , 2020) .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>6NB7</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Briefly , 2.5e5 293T cells per well were seeded in 12-well plates in 1 mL D10 growth media ( DMEM with 10 % heat-inactivated FBS , 2 mM lglutamine , 100 U/mL penicillin , and 100 μg/mL streptomycin) .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>293T</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Media was removed from the 293TACE2 cells and replaced with fresh D10 containing 50 μL of pseudovirus supernatant in a final volume of 150 μL .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>293TACE2</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Plasmids were transfected into 150mL suspension expi293F or HEK293F cells at 37°C in a humidified 8% CO2 incubator rotating at 130 rpm and harvested 3 days later.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>HEK293F</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Methods Data and Code Availability We provide all data and code in the following ways: ● Raw data tables of our replicate functional scores at the level of single mutations ( Supplemental File 3 , and GitHub: https://github.com/jbloomlab/SARS-CoV-2-RBD_DMS/blob/master/results/single_mut_effects/single_mut_effects.csv ) ● Raw data tables of our replicate functional scores among sarbecovirus homologs ( Supplemental File 1 and GitHub: https://github.com/jbloomlab/SARS-CoV-2-RBD_DMS/blob/master/results/single_mut_effects/homolog_effects.csv ) ● Illumina sequencing counts for each barcode among FACS bins ( https://github.com/jbloomlab/SARS-CoV-2RBD_DMS/blob/master/results/counts/variant_counts.csv ) ● The complete variant:barcode lookup table ( https://github.com/jbloomlab/SARS-CoV-2RBD_DMS/blob/master/results/variants/codon_variant_table.csv ) ● The complete computational workflow to generate and analyze these data , including reproducible code within a programmatically constructed computational environment ( https://github.com/jbloomlab/SARS-CoV-2-RBD_DMS ) ● A Markdown summary of the organization of analysis steps , with links to key data files and Markdown summaries of each step in the analysis pipeline ( https://github.com/jbloomlab/SARS-CoV-2RBD_DMS/blob/master/results/summary/summary.md) , with specific Markdown summaries linked in the relevant Methods sections below ● All raw sequencing data are uploaded to the NCBI Short Read Archive ( BioProject PRJNA639956)</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>NCBI Short Read Archive</b></div>
        <div>suggested: None</div>
      </div>

      <div style="margin-bottom:8px">
        <div><b>BioProject</b></div>
        <div>suggested: (NCBI BioProject, <a href="https://scicrunch.org/resources/Any/search?q=SCR_004801">SCR_004801</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">For one bin in which the number of HiSeq reads was less than the number of cells sorted into a bin , we re-amplified PCR product from a newly purified plasmid aliquot , and obtained reads via a single lane of MiSeq 50bp single end sequencing .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>MiSeq</b></div>
        <div>suggested: (A5-miseq, <a href="https://scicrunch.org/resources/Any/search?q=SCR_012148">SCR_012148</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Data visualization The interactive heatmap of mutational effects shown at https://jbloomlab.github.io/SARS-CoV-2-RBD_DMS/ was made using the altair ( VanderPlas et al. , 2018 ) Python package.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Python</b></div>
        <div>suggested: (IPython, <a href="https://scicrunch.org/resources/Any/search?q=SCR_001658">SCR_001658</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Structural images were rendered in PyMol .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>PyMol</b></div>
        <div>suggested: (PyMOL, <a href="https://scicrunch.org/resources/Any/search?q=SCR_000305">SCR_000305</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">RBD nucleotide sequences were aligned via mafft with a gap opening penalty of 4.5 , and the maximum likelihood phylogeny was inferred in RAxML ( Stamatakis , 2014 ) under the GTR model with 4 gamma-distributed discrete categories of among-site rate variation .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>RAxML</b></div>
        <div>suggested: (RAxML, <a href="https://scicrunch.org/resources/Any/search?q=SCR_006086">SCR_006086</a>)</div>
      </div>
    </td></tr></table>

Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:

. To some degree, these caveats are universal of experimental studies, as even sophisticated animal models are imperfect proxies for true fitness

Results from OddPub: Thank you for sharing your code and data.

About SciScore

Visit annotations in context

Annotators

sciscore

URL

biorxiv.org/cgi/content/10.1101/2020.06.17.157982

www.biorxiv.org www.biorxiv.org

https://doi.org/10.1101/2020.06.14.151357

1
1. sciscore 01 Jul 2020
  
  in Public
  
  SciScore for 10.1101/2020.06.14.151357: (What is this?)
  Please note, not all rigor criteria are appropriate for all manuscripts.
  Table 1: Rigor
  <table><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Institutional Review Board Statement</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Randomization</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Blinding</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Power Analysis</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Sex as a biological variable</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Cell Line Authentication</td><td style="min-width:100px;border-bottom:1px solid lightgray">Cells were regularly passaged and tested for presence of mycoplasma contamination ( MycoAlert Plus Mycoplasma Detection Kit , Lonza)</td></tr></table>
  Table 2: Resources
  <table><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2">Antibodies</td></tr><tr><td style="min-width:100px;text=align:center">Sentences</td><td style="min-width:100px;text-align:center">Resources</td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Primary antibody incubations were performed overnight at 4°C using the following antibodies: rabbit anti-GAPDH 14C10 ( 0.1 μg/mL , Cell Signaling 2118S) , mouse anti-rhodopsin antibody clone 1D4 ( 1 μg/mL , Novus NBP1-47602 ) which recognizes the C9-tag added to the Spike proteins .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-GAPDH</div> <div>suggested: (Cell Signaling Technology Cat# 2118, AB_561053)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Following the primary antibody , the blots were incubated with IRDye 680RD donkey anti-rabbit ( 0.2 μg/mL , LI-COR 926-68073 ) or with IRDye 800CW donkey anti-mouse ( 0.2 μg/mL , LI-COR 926-32212 ) for 1 hour at room temperature.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-rabbit</div> <div>suggested: (LI-COR Biosciences Cat# 926-68073, AB_10954442)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Spike was immunoprecipitated using 2 µg C9 antibodies ( Novus NBP1-47602 ) per sample and incubated on a rotator at 4°C for at least 4 hours .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>C9</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Western blotting was performed as described above using mouse anti-rhodopsin antibody clone 1D4 ( 1 μg/mL , Novus NBP1-47602 ) which recognizes the C9-tag added to the Spike proteins .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-rhodopsin</div> <div>suggested: (Novus Cat# NBP1-47602, AB_10010560)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Following the primary antibody , the blots were incubated with IRDye 800CW donkey anti-mouse ( 0.2 μg/mL , LI-COR 926-32212 ) for 1 hour at room temperature.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-mouse</div> <div>suggested: (LI-COR Biosciences Cat# 926-32212, AB_621847)</div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2">Experimental Models: Cell Lines</td></tr><tr><td style="min-width:100px;text=align:center">Sentences</td><td style="min-width:100px;text-align:center">Resources</td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Cell culture A549 cells were obtained from ATCC , HEK293FT cells were obtained from Thermo Scientific , and Huh-7.5 and Caco-2 were a kind gift of T . Jordan and B . tenOever ( Mt . Sinai) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>A549</div> <div>suggested: None</div> </div>
  
  <div style="margin-bottom:8px"> <div><b>HEK293FT</b></div> <div>suggested: ATCC Cat# PTA-5077, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_6911">CVCL_6911</a></div> </div> <div style="margin-bottom:8px"> <div><b>Huh-7.5</b></div> <div>suggested: <a href="https://scicrunch.org/resources/Any/search?q=CVCL_7927">CVCL_7927</a></div> </div> <div style="margin-bottom:8px"> <div><b>Caco-2</b></div> <div>suggested: CLS Cat# 300137/p1665_CaCo-2, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0025">CVCL_0025</a></div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Briefly , for each virus , a T-225 flask of 80 % confluent HEK293T cells ( Thermo ) was transfected in OptiMEM ( Thermo ) using 25 µg of the transfer plasmid , 20 µg psPAX2 , 22 µg spike plasmid , and 175 µl of linear Polyethylenimine ( 1 mg/ml ) ( Polysciences) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>HEK293T</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">ACE2 lentiviral cloning and ACE2 stable cell line overexpression To generate pLenti-ACE2-Hygro , we amplified human ACE2 ( hACE2 ) from pcDNA3.1-ACE2 ( Addgene 1786 ) and cloned it into a lentiviral transfer pLEX vector carrying the hygromycin resistance gene using Gibson Assembly Master Mix ( NEB E2611L) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>ACE2</b></div> <div>suggested: <a href="https://scicrunch.org/resources/Any/search?q=CVCL_DR94">CVCL_DR94</a></div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Huh7.5-ACE2 and A549-ACE2 cell lines were generated by lentiviral transduction of ACE2 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Huh7.5-ACE2</b></div> <div>suggested: None</div> </div> <div style="margin-bottom:8px"> <div><b>A549-ACE2</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Z . D . is supported by an American Heart Association postdoctoral fellowship . N.E . S. is supported by New York University and New York Genome Center startup funds , National Institutes of Health ( NIH)/National</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>American Heart Association</b></div> <div>suggested: (American Heart Association, <a href="https://scicrunch.org/resources/Any/search?q=SCR_007210">SCR_007210</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Band intensity quantification was performed by first converting Odyssey multichannel TIFFs into 16-bit grayscale image ( Fiji ) and the then selecting lanes and bands in ImageLab 6.1 ( BioRad) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Fiji</b></div> <div>suggested: (Fiji, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002285">SCR_002285</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">For each peptide , we computed the difference in predicted affinity between the D614 and G614 variant using R/RStudio and visualized them using the pheatmap R package.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>pheatmap</b></div> <div>suggested: (pheatmap, <a href="https://scicrunch.org/resources/Any/search?q=SCR_016418">SCR_016418</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Statistical analysis Data analysis was performed using R/Rstudio 3.6.1 and GraphPad Prism 8 ( GraphPad Software Inc . )</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>GraphPad</b></div> <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div> </div> </td></tr></table>
  
  Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.
  Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
  Results from OddPub: Thank you for sharing your code.
  About SciScore
  
  SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.
Visit annotations in context

Annotators

sciscore

URL

biorxiv.org/cgi/content/10.1101/2020.06.14.151357
doi.org doi.org

https://doi.org/10.1101/2020.04.28.20083691

1
1. sciscore 01 Jul 2020
  
  in Public
  
  SciScore for 10.1101/2020.04.28.20083691: (What is this?)
  Please note, not all rigor criteria are appropriate for all manuscripts.
  Table 1: Rigor
  <table><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Institutional Review Board Statement</td><td style="min-width:100px;border-bottom:1px solid lightgray">All samples were collected under approval of the Institutional Review Board for Huma Subjects Research at Massachusetts General Hospital.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Randomization</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Blinding</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Power Analysis</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Sex as a biological variable</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Cell Line Authentication</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr></table>
  Table 2: Resources
  <table><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2">Antibodies</td></tr><tr><td style="min-width:100px;text=align:center">Sentences</td><td style="min-width:100px;text-align:center">Resources</td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Many serological enzyme-linked immunosorbent assays (ELISA) have been recently developed to detect anti-SARS-CoV-2 antibodies.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-SARS-CoV-2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">To address these limitations, we developed ultra-sensitive Single Molecule Array (Simoa) assays for anti-SARS-CoV-2 IgG, IgM, and IgA antibodies against four immunogenic viral proteins, providing us with detailed information about early stages of immune activation 17.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>IgA antibodies against four immunogenic viral proteins, providing us with detailed information about early stages of immune activation</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Developing an ultra-sensitive Simoa assay for anti-SARS-CoV-2 antibodies We developed a multiplexed ultra-sensitive Simoa assay for detection of IgG, IgM, and IgA against SARS-CoV-2 in human plasma.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>IgA against SARS-CoV-2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Finally, the beads are incubated with the enzyme streptavidin-βgalactosidase (SβG), which binds to the biotinylated anti-human immunoglobulin antibody, forming a complete enzyme-labeled immunocomplex.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-human immunoglobulin</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">We quantitatively validated viral target conjugation to the bead surface using anti-His tag antibodies as well as recombinant human anti-RBD antibody, as described in the Supplementary Information (Supplementary Figure 3).</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-His tag</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">For spike , S1 , and nucleocapsid , confirmation of antigen attachment to the beads was demonstrated by Simoa with His tags experiments using a biotinylated anti-His tag antibody ( ThermoFisher MA121315BTI ) on the HD-X Analyzer ( Quanterix) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>S1</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The anti-His-tag antibody was plated at concentrations of 0.1 pg/mL to 10,000 pg/mL using tenfold dilutions .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-His-tag</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">RBD conjugation to beads was confirmed by Simoa with an anti-RBD antibody ( clone CR3022 ) and a biotinylated anti human-IgG antibody ( Bethyl Laboratories A80-148B)</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>anti-RBD</div> <div>suggested: None</div> </div>
  
  <div style="margin-bottom:8px"> <div><b>anti human-IgG</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Biotinylation Detection antibodies for IgA , IgG and IgM were purchased from Thermo Fisher , Bethyl Laboratories , Abcam , Biolegend , and R&D systems ( see Immunoglobulin Simoa assay format ) and were biotinylated for use in Simoa assays as described previously by Cohen et al 23 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>IgA , IgG</b></div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Anti-human immunoglobulin antibodies were diluted in Homebrew Detector/Sample Diluent to final concentrations of: IgG (Bethyl Labratories A80-148B): 7.73ng/mL, IgM (Thermo Fisher MII0401): 216ng/mL, IgA (Abcam ab214003): 150ng/mL.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>IgM</b></div> <div>suggested: (Thermo Fisher Scientific Cat# MII0401, <a href="https://scicrunch.org/resources/Any/search?q=AB_11153935">AB_11153935</a>)</div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The expression construct was transiently transfected in HEK 293T cells using polyethylenimine.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>HEK 293T</b></div> <div>suggested: KCB Cat# KCB 200744YJ, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0063">CVCL_0063</a></div> </div> </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Logistic regression analysis was conducted in R version 3.6.2 for the multivariate analysis and Graphpad Prism 7 for the univariate analysis25.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Graphpad Prism</b></div> <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">All figures were plotted in Graphpad Prsim 7, Igor Pro7 and Adobe Illustrator version 2015. Acknowledgments: The authors would like to thank Liangxia Xie for the helpful discussion regarding the experimental design.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Graphpad</b></div> <div>suggested: (GraphPad, <a href="https://scicrunch.org/resources/Any/search?q=SCR_000306">SCR_000306</a>)</div> </div> <div style="margin-bottom:8px"> <div><b>Adobe Illustrator</b></div> <div>suggested: (Adobe Illustrator, <a href="https://scicrunch.org/resources/Any/search?q=SCR_010279">SCR_010279</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">In addition, support came from the Globa TravEpiNet (GTEN) system sponsored by the US Centers for Disease Control and Preventio (Grant No. U01CK000490: ETR, RCC) as well as a T32GM007753 grant from NIGMS, T32 AI007245 from NIAID, and an R01AI146779 from NIAID.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>NIAID</b></div> <div>suggested: (NIAID, <a href="https://scicrunch.org/resources/Any/search?q=SCR_016598">SCR_016598</a>)</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">He is an inventor of the Simoa technology, a founder o the company and also serves on its Board of Directors. Dr. Walt’s interests were reviewed an are managed by BWH and Partners HealthCare in accordance with their conflict of interes policies.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div><b>Partners HealthCare</b></div> <div>suggested: (Partners HealthCare Biobank, <a href="https://scicrunch.org/resources/Any/search?q=SCR_001316">SCR_001316</a>)</div> </div> </td></tr></table>
  
  Results from OddPub: Thank you for sharing your data.
  About SciScore
  
  SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore is not a substitute for expert review. SciScore checks for the presence and correctness of RRIDs (research resource identifiers) in the manuscript, and detects sentences that appear to be missing RRIDs. SciScore also checks to make sure that rigor criteria are addressed by authors. It does this by detecting sentences that discuss criteria such as blinding or power analysis. SciScore does not guarantee that the rigor criteria that it detects are appropriate for the particular study. Instead it assists authors, editors, and reviewers by drawing attention to sections of the manuscript that contain or should contain various rigor criteria and key resources. For details on the results shown here, including references cited, please follow this link.
Visit annotations in context

Annotators

sciscore

URL

doi.org/10.1101/2020.04.28.20083691

SciScore for 10.1101/2020.03.21.990770: (What is this?)

Please note, not all rigor criteria are appropriate for all manuscripts.

Table 1: Rigor

<table><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Institutional Review Board Statement</td><td style="min-width:100px;border-bottom:1px solid lightgray">This study received approval from the Research Ethics Committee of Shenzhen Third People 's Hospital , China ( approval number: 2020-084) .</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Randomization</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Blinding</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Power Analysis</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Sex as a biological variable</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr><tr"><td style="min-width:100px;margin-right:1em; border-right:1px solid lightgray; border-bottom:1px solid lightgray">Cell Line Authentication</td><td style="min-width:100px;border-bottom:1px solid lightgray">not detected.</td></tr></table>

Table 2: Resources

<table><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2">Antibodies</td></tr><tr><td style="min-width:100px;text=align:center">Sentences</td><td style="min-width:100px;text-align:center">Resources</td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Among a total of 69 antibodies from P#2 , the majority ( 59 % ) were scattered across various branches and the remaining ( 41 % ) were clonally expanded into three major clusters ( Figure 3A) .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>total of 69</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Control antibodies from P#1 demonstrated even lower competing power with ACE2 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>ACE2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">We selected a total of six antibodies with ACE2 competitive capacities of at least 70 % and analyzed them in a pairwise competition fashion using SPR .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>SPR</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The most potent antibody , P2C-1F11 , did not seem target the same epitope as the relatively moderate antibody P2C-1C10 .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>P2C-1F11</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Finally , despite successfully isolating and characterizing a large of number mAbs against SARS-CoV-2 , we cannot draw any firm correlation between antibody response and disease status at this time.</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>SARS-CoV-2</div> <div>suggested: None</div> </div> </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The third staining at 4 °C for 30min involved either: Streptavidin-APC ( eBioscience ) and/or Streptavidin-PE ( BD Biosciences ) to target the Strep tag of RBD , or antihis-APC and anti-his-PE antibodies ( Abcam ) to target the His tag of RBD .</td><td style="min-width:100px;border-bottom:1px solid lightgray"> <div style="margin-bottom:8px"> <div>antihis-APC</div> <div>suggested: None</div> </div>

      <div style="margin-bottom:8px">
        <div><b>anti-his-PE</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The IgG heavy and light chain variable genes were amplified by nested PCR and cloned into linear expression cassettes or expression vectors to produce full IgG1 antibodies as previously described 29,41 .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>full IgG1</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The PCR products were purified and cloned into the backbone of antibody expression vectors containing the constant regions of human IgG1 .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>human IgG1</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">SARS-CoV antibodies ( S230 and m396 ) previously isolated by others 42 were synthesized and sequences verified before expression in 293T cells and purification by protein A chromatography .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>SARS-CoV</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">HIV-1 antibody VRC01 was a broadly neutralizing antibody directly isolated from a patient targeting the CD4 binding site of envelope glycoprotein 40 .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>CD4 binding site of envelope glycoprotein 40</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The cells were then stained with PE labeled anti-human IgG Fc secondary antibody ( Biolegend ) at a 1:20 dilution in 50 μl staining buffer at room temperature for 30 minutes .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-human IgG</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">VRC01 is negative control antibody targeting HIV-1 envelope glycoprotein.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>HIV-1 envelope glycoprotein.</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The recombinant RBD was labeled with either a Strep or His tag and used alone or in combination to identify and isolate RBD-specific single B cells through staining with the Streptavidin-APC and/or Streptavidin-PE, or anti-His- APC and anti-His-PE antibodies.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>anti-His- APC</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Experimental Models: Cell Lines</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">SARS-CoV-2 , SARS-CoV and MERS-CoV pseudovirus were generated by cotransfection of human immunodeficiency virus backbones expressing firefly luciferase ( pNL43R-E-luciferase ) and pcDNA3.1 ( Invitrogen ) expression vectors encoding the respective S proteins into 293T cells ( ATCC ) 37,38,44,45</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>293T</b></div>
        <div>suggested: KCB Cat# KCB 200744YJ, <a href="https://scicrunch.org/resources/Any/search?q=CVCL_0063">CVCL_0063</a></div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Huh7 cells ( ATCC ) ( approximately 1.5 × 104 per well ) were added in duplicate to the virusantibody mixture.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Huh7</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The isolate was amplified in Vero cell lines to make working stocks of the virus ( 1 × 105 PFU/ml) .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Vero</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Serial dilutions of mAbs were mixed separately with 100 PFU of SARS-CoV-2 , incubated at 37 °C for 1 h , and added to the monolayer of Vero E6 cells in duplicates .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Vero E6</b></div>
        <div>suggested: <a href="https://scicrunch.org/resources/Any/search?q=CVCL_XD71">CVCL_XD71</a></div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The genes encoding the heavy and light chains of isolated antibodies were separately cloned into expression vectors containing IgG1 constant regions and the vectors were transiently transfected into HEK293T or 293F cells using polyethylenimine ( PEI ) ( Sigma) .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>293F</b></div>
        <div>suggested: <a href="https://scicrunch.org/resources/Any/search?q=CVCL_D615">CVCL_D615</a></div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">HEK 293T cells transfected with expression plasmid encoding the full length spike of SARS-CoV-2, SARS-CoV or MERS-CoV were incubated with 1:100 dilutions of plasma from the study subjects.</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>HEK 293T</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr><tr><td style="min-width:100px;text-align:center; padding-top:4px;" colspan="2"><b>Software and Algorithms</b></td></tr><tr><td style="min-width:100px;text=align:center"><i>Sentences</i></td><td style="min-width:100px;text-align:center"><i>Resources</i></td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Bioinformatic and biologic characterization indicates that these antibodies are derived from broad and diverse families of antibody heavy and light chains .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>Bioinformatic</b></div>
        <div>suggested: (QFAB Bioinformatics, <a href="https://scicrunch.org/resources/Any/search?q=SCR_012513">SCR_012513</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Finally , the cells were re-suspended and analyzed with FACS Calibur instrument ( BD Biosciences , USA ) and FlowJo 10 software ( FlowJo , USA)</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>FlowJo</b></div>
        <div>suggested: (FlowJo, <a href="https://scicrunch.org/resources/Any/search?q=SCR_008520">SCR_008520</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Half-maximal inhibitory concentrations ( IC50 ) of the evaluated mAbs were determined by luciferase activity 48h after exposure to virusantibody mixture using GraphPad Prism 6 ( GraphPad Software Inc . ) .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>GraphPad Prism</b></div>
        <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div>
      </div>

      <div style="margin-bottom:8px">
        <div><b>GraphPad</b></div>
        <div>suggested: (GraphPad Prism, <a href="https://scicrunch.org/resources/Any/search?q=SCR_002798">SCR_002798</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">The IgG heavy and light chain variable genes were aligned using Clustal W in the BioEdit sequence analysis package ( https://bioedit.software.informer.com/7.2/).</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>BioEdit</b></div>
        <div>suggested: (BioEdit, <a href="https://scicrunch.org/resources/Any/search?q=SCR_007361">SCR_007361</a>)</div>
      </div>
    </td></tr><tr><td style="min-width:100px;vertical-align:top;border-bottom:1px solid lightgray">Phylogenetic analyses were performed by the Maximum Likelihood method using MEGA X ( Molecular Evolutionary Genetics Analysis across computing platforms) .</td><td style="min-width:100px;border-bottom:1px solid lightgray">
      <div style="margin-bottom:8px">
        <div><b>MEGA X</b></div>
        <div>suggested: None</div>
      </div>
    </td></tr></table>

Results from OddPub: Thank you for sharing your data.

About SciScore

Visit annotations in context

Annotators

sciscore

URL

doi.org/10.1101/2020.03.21.990770

Jun 2020
web.archive.org web.archive.org

Bokardo » Blog Archive » Why We Can’t Compare Folksonomies to Search

1
1. theresia 28 Jun 2020
  
  in Public
  
  The more I use del.icio.us and observe other folksonomies, the more I realize that we don’t use them to find “stuff”. We use them to discover “personally-related stuff”, which is really hard to do with a search engine.
  
  searching vs tagging.
  
  clay says: search is for finding. tag is for keeping
  
  tags are contextual and personal
  
  searching are keyword-dependent but more or less more objective
Visit annotations in context

Annotators

theresia

URL

web.archive.org/web/20051220054408/http://bokardo.com/archives/why-we-cant-compare-folksonomies-to-search/
www.editions-ellipses.fr www.editions-ellipses.fr

Manuel de psychogénéalogie

1
1. alexandre_JEMS 24 Jun 2020
  
  in Public
  
  test Alex tag
  
  hello youpi oula olé roger test youyou
Visit annotations in context

Tags

youyou

youpi

olé

hello

oula

roger

test

Annotators

alexandre_JEMS

URL

editions-ellipses.fr/accueil/10461-manuel-de-psychogenealogie-9782340038721.html
clas3209.wordpress.com clas3209.wordpress.com

Zoroastrianism

1
1. zeromani 22 Jun 2020
  
  in Public
  
  Tags: Final Project, scythia, team steppes
  
  This piece is about the Parthians. You should tag it as Parthia, not Scythia.
Visit annotations in context

Annotators

zeromani

URL

clas3209.wordpress.com/2020/06/22/zoroastrianism/
www.biorxiv.org www.biorxiv.org

Structures of three MORN repeat proteins and a re-evaluation of the proposed lipid-binding properties of MORN repeats

2
1. EMBOpress 22 Jun 2020
  
  in Review Commons
  
  Note: This rebuttal was posted by the corresponding author to Review Commons. Content has not been altered except for formatting.
  
  Learn more at Review Commons
  
  Reply to the reviewers
  
  The response to reviewers consists of three parts:
  
  A summary of the main points from the two reviews, and the authors' response to these points.
  
  A detailed revision plan for the preprint, taking into account both the main points of the reviews, and other comments made by the reviewers.
  
  A point-by-point response to the reviewers.
  
  For figure citations, OV = old version, i.e. bioRxiv preprint 2019-826180v2, and NV = new version, i.e. revised and re-submitted version.
  
  1. Summary of main points by the reviewers, and authors’ responses:
  
  Both reviewers felt that the manuscript was overlong; Reviewer 1 recommended either shortening it or splitting it into two stories, while Reviewer 2 recommended cutting down the text.
  
  We have considerably shortened the manuscript in accordance with this request (see revision plan below). We had already considered splitting the manuscript into two parts during the drafting stage, and had rejected this possibility as the data are intertwined - the retroactive validation of the dimer interface by the mutagenesis constructs (OV Fig. S3 [NV Fig. S4]) being a good example.
  
  The revised manuscript features 7 main figures and 13 supplementals.
  
  Both reviewers felt too much text and figure space was allocated to negative data, specifically the investigation of potential lipid binding by the TbMORN1 protein, and that there should be more focus on the positive parts of the story.
  
  A key part of shortening the manuscript has been moving most of the negative data on lipid binding into the supplemental figures, and considerably shortening the associated text. This has allowed the main figures and associated text to focus more on the positive elements of the project, while still ensuring publication of all the data.
  
  The reviewers appear to be in slight disagreement concerning discussion of the data. Reviewer 1 has encouraged more speculation on the physiological role of PE binding, a potential lipid transfer function, a role for calcium ions, the relevance of the observed disulphide bond, and the role of zinc ions in apicomplexan proteins; Reviewer 2 has recommended avoiding excessive speculation or inference.
  
  Given that both reviewers have agreed that the original manuscript was overlong, we have implemented Reviewer 2's suggestion here and reduced the amount of speculation in the revised text.
  
  The reviewers agreed that the technical quality of the data was high and that the conclusions drawn were robust.
  
  We are glad that the reviewers were appreciative of the data quality. For this reason, we were reluctant to remove any of the data from the manuscript and would prefer instead to transfer it to the supplementals. We feel that the negative data still have considerable community value, given that they show that MORN repeats are not automatically lipid binding modules and can thus act as a caveat to other researchers.
  
  2. Detailed revision plan for the preprint:
  
  We have implemented the reviewers' suggestions and substantially shortened the manuscript, primarily by trimming the (phospho)lipid-binding section, which contains a large amount of negative data. The following main figures have been moved into the supplemental section:
  
  OV Fig. 2 ("TbMORN1 interacts with phospholipids but not liposomes") has become NV Fig. S2
  
  OV Fig. 4 ("TbMORN1(2-15) does not bind to liposomes in vitro") has become NV Fig. S6
  
  OV Fig. 8 ("Conservation and properties of residues in TbMORN1(7- 15)") has become NV Fig. S11
  
  This has left a total of 7 main figures and 13 supplementals.
  
  The text associated with the entirety of the lipid-binding part (OV lines 210- 530, OV Figs. 2-6 [NV Figs. 2-4, S2, S6], OV Supplemental Figs. 2-6 [NV Supplemental Figs. S3-S5, S7, S8]) has been condensed. The focus of this section is now on the positive parts of the data: the PE association (OV Fig. 3 [NV Fig. 2]) and the in vivo work (OV Figs. 5, 6 [NV Figs. 3, 4]).
  
  We have additionally limited the amount of inference and speculation in the manuscript.
  
  3. Point-by-point responses to the reviewers
  
  Reviewer #1 (Evidence, reproducibility and clarity):
  
  MORN (membrane occupation and recognition nexus) repeat proteins are found in prokaryotes and eukaryotes. They feature characteristic repeats in their primary sequence, have been assumed to play a role in lipid binding, but remain poorly characterized on the functional and structural level. This manuscript tries to address both these questions and is organized in major parts. In the first part the authors characterize a putative role of MORN repeat proteins in lipid binding and membrane association. In the second part, the authors use X-ray crystallography to establish the structure of MORN repeat proteins and to investigate the dimerization.
  
  As a cleverly chosen point of departure, they focus their study particularly on MORN1 from Trypanosoma brucei (TbMORN1), which is composed solely on MORN repeats. The structures of MORN repeats (from several species) in part two provide interesting insights into their mode of homotypic interactions and their role as dimerization or oligomerization devices. The lipid binding and membrane association of MORN proteins in the first part remains somewhat confusing and unclear, despite the use of a whole battery of techniques.
  
  We anticipate that the shortening and refocusing of the lipid binding data has addressed this issue.
  
  It is questionably, why the authors invest so many figures and words to inform the reader on negative results.
  
  We have chosen to publicise our negative data in full because, as noted in the manuscript, there is a widespread and erroneous assumption that MORN repeats are lipid binding modules. We feel that publishing these data will allow them to act as a caveat to other researchers working on MORN repeat proteins. We have, however, addressed the reviewer's request in that we have considerably shortened the text associated with these data and have moved the corresponding figures into the supplementals.
  
  The authors suggest that MORN proteins can bind to lipids via their hydrophobic acyl chainswhich is 'very hard to imagine under physiological conditions unless TbMORN1 is a lipid carrier and not a membrane-binding proteins. Unfortunately, a role as lipid carrier has not been rigorously tested.
  
  The reviewer is correct that we have not specifically tested for a function as a lipid carrier protein and although this was only speculation, it has been toned down accordingly.
  
  In this sense the first part remains somewhat immature and incoherent. Furthermore, they suggest based on the lack-of-evidence that MORN proteins do not bind membranes in vivo and in vitro.
  
  We are not clear where this suggestion was made. Our data indicate that TbMORN1 does not directly bind membranes in vivo or in vitro, and we therefore noted that putative lipid binding by other MORN repeat proteins should be viewed with caution. Specifically, we stated in the Discussion (OV lines 955-956) that "the presence of MORN repeats in a protein should not be taken as indicative of lipid binding or lipid membrane binding without experimental evidence". Again, our expectation is that the major changes planned for the data presentation in this section will make it more coherent.
  
  The main issue of this manuscript is, in my view, the way the data were presented.The manuscript is generally well-written, but much too long. The structural work is important and concise.
  
  We have considerably shortened the manuscript as per the reviewer's request, and especially the section on lipid binding.
  
  The first part, however, reports in five separate figures on a lack of membrane binding by a MORN protein and its ability to bind individual lipids. The physiologically relevance of this lipid binding is questionable as acknowledged by the authors.
  
  We have moved two of these figures (OV Figs. 2, 4) into the supplementals section [NV Figs. S2, S6], shortened the associated text, and limited the amount of speculation.
  
  Even though I find it important that the membrane/lipid binding ability of MORN proteins is rigorously tested, I would highly recommend to separate the current manuscript in two independent stories. Alternatively, I would recommend to reduce the first part into a single figure and to remove the most artifactual assays.
  
  We have implemented the second of these two suggestions for the manuscript. We had already considered splitting the manuscript during the drafting stage, but rejected this possibility as the data were too intertwined. Consequently, we have opted to considerably reduce the first part, and moved OV Figs. 2 and 4 into the supplementals [NV Figs. S2, S6]. We would prefer not to remove data altogether as they are likely to have community value even if they are negative and as noted, they are of good quality.
  
  In the current form, the first part and the second part of the manuscript remain somewhat detached from each other. The characterization of the lipid binding/membrane binding properties has a number of substantial weaknesses (e.g. use of quite different, nonphysiological buffers for membrane binding assays; use of deletion mutants for the binding assays, which do not show the full potential of oligomerization). This which makes it hard to read and confuses the reader. Even though I have no reason to doubt the conclusions by the authors, I do not think that all necessary caution has been invested to rule out other possibilities.
  
  We believe that the shortening and refocusing of the manuscript should address these issues. For consideration of the buffer and deletion mutant points, please see responses to Major Points below.
  
  In summary, even though the technical quality of the individual performed assays is high, there are some conceptual issues that make it hard to make a strong case based on a collection of individual, clear datasets. Even though I find the structures of the MORN proteins important, timely, and interesting, I would not recommend this study for publication in its current form. The manuscript would be more fun to read if both of the parts would be shortened substantially and more focused.
  
  We have implemented this suggestion: the manuscript has been considerably shortened (from 20,489/135,073 to 18,555/103,988 characters/words, focused on reducing the negative lipid-binding results).
  
  While I agree that most evidence provided on lipid/membrane binding of TbMORN1 argue against a direct role of MORN proteins in membrane binding, I feel that the experimental approach is not coherent enough. See a few major points of criticism below.
  
  Major Points:
  
  The authors decide to characterize the membrane binding of a MORN repeat protein using a deletion variant that lacks the N-terminal repeat. However, in Figure 1B they show that the N-terminal repeat is important for the formation of higher-order oligomers. While I fully understand that the presence of the most N-terminal repeat does hamper the structural work, I find it problematic to remove it for the lipid/membrane-binding assays. The formation of higher oligomeric species beyond the dimer, may be important for membrane binding/recruitment (avidity effects).
  
  As we explained in the manuscript, the reason for not using the full-length protein for in vitro work was because it was polydisperse, and that the yields were extremely low. See OV lines 178-179 ("The yields of TbMORN1(1-15) were always very low, making this construct not generally suitable for in vitro assays".) and OV lines 411-414 ("...TbMORN1(1-15), which was polydisperse in vitro and formed large oligomers (Fig. 1B). The membrane-binding activity of these polydisperse oligomers was not possible to test in vitro, as the purification yields of TbMORN1(1-15) were always low."). Consequently, we used the longest construct that was suitable in terms of chemical and oligomeric homogeneity. Using the full-length protein would have had inherent problems with aggregation, and consequently would have compromised the data and derived results. In order to make this clear in the manuscript we edited the sentence mentioned above as follows:
  
  “It was not possible to test the membrane-binding activity of these polydisperse oligomers in vitro however, as the purification yields of TbMORN1(1-15) were always low. As an alternative, the possible membrane association of TbMORN1(1-15) was examined in vivo."
  
  2) (Related to point 1) I do not understand the choice of the buffers used for some of the assays. The use of pH 8.5 and NaCl concentrations of 200 mM are non-physiological.
  
  These were the buffer conditions required to retain the protein in a monodisperse state, suitable for in vitro assays.
  
  For CD spectroscopy, a high ionic strength was obtained by the use of 200 mM NaF. If a high ionic strength is required to prevent the formation of higher oligomers of MORN, it raises the question if the formation of higher oligomers (under physiological conditions) may also contribute to their function.
  
  The oligomers of TbMORN1 may indeed be the most functionally relevant form of TbMORN1 but we do not currently have a means of testing this in vitro, as acknowledged in the text (OV lines 411-414, quoted above). The aim of CD spectroscopy was to assess fold integrity and stability of different constructs; we used buffers as recommended for the CD spectroscopy experiments by Kelly et al, 2005 (doi:10.1016/j.bbapap.2005.06.005) (Table 1 and section 4.2). Furthermore, the CD spectra of TbMORN(1-15) and TbMORN(2-15) (OV Fig. S1E [NV Fig. S1E]) are basically superimposable, suggesting identical secondary structure content at the concentration used for these experiments.
  
  It is unclear, in which buffer the fluorescence anisotropy measurements were performed.
  
  We have provided details on the buffer conditions for the fluorescence anisotropy experiments in the Materials and Methods section, NV page 23, lines 962-963.
  
  The sucrose-loaded vesicles were hydrated in a 20 mM HEPES pH 7.4, 0.3 M Sucrose. The composition of the buffer after the addition of MORN proteins is not clear.
  
  The Materials and Methods are now unambiguous on this point. Please see NV lines 1036- 1046: "6 μM Rhodamine B dihexadecanoyl phosphoethanolamine (Rh-DHPE) was added to all lipid mixtures to facilitate the visualisation of the SLVs. The lipid mixtures were dried under a nitrogen stream, and the lipid films hydrated in 20 mM HEPES pH 7.4; 0.3 M sucrose. The lipid mixtures were subjected to 4 cycles of freezing in liquid nitrogen followed by thawing in a sonicating water bath at RT. The vesicles were pelleted by centrifugation (250,000 × g, 30 min, RT) and resuspended in 20 mM HEPES pH 7.4, 100 mM KCl to a total lipid concentration of 1 mM. SLVs were incubated with 1.5 μM purified TbMORN1(2-15) in gel filtration buffer (20 mM Tris-HCl pH 8.5, 200 mM NaCl, 2% glycerol, 1 mM DTT) at a 1:1 ratio (30 min, RT)." The liposomes were at physiological pH and close to physiological ionic strength.
  
  Despite the use of an impressive array of techniques, this first part of the manuscript remains somewhat immature and incoherent. Due to the use of constructs that have not the full ability to oligomerize (point 1) and due to the inconsistent use of experimental conditions, it is hard to draw firm conclusions from this first part.
  
  Any biochemical study is conducted within the constraints of the choice of construct and the choice of buffer conditions, and the data are valid within those parameters. This applies as much to positive data as to negative data, so we are not clear why the reviewer is placing such emphasis on this point. In the case of the LiMA data, which are the most unbiased and comprehensive dataset in the manuscript, these experiments were well-controlled and there were also domains present that were recruited to membranes under the buffer conditions, allowing us to rule out that the assay conditions were completely unsuitable. Validating negative results should be done as carefully and with as many orthogonal approaches as the validation of positive results. The reviewer acknowledges below that "the data point in the direction that MORN proteins (or at least TbMORN1) does not directly bind to membranes". This is the conclusion that we wanted to communicate.
  
  For example: In Figure 2E TbMORN(2-15) does show some concentration-dependent binding, which -however- is interpreted as background binding. What are the results using this assay (or better: a liposome floatation assay) when using full-length TbMORN(1-15) in a more physiological buffer?
  
  As noted already, it is not possible to use the TbMORN1(1-15) construct for in vitro assays owing to the extremely low yields and polydisperse nature of the protein. The excess fulllength protein was associated with the cytosolic fraction and not the membrane fraction in vivo (OV Fig. 6B [NV Fig. 4B]).
  
  The statement that MORN proteins bind to lipids, but not to liposomes/membranes is -in my view- not sufficiently addressed to make a strong case.
  
  At no point do we suggest that MORN repeat proteins in general bind to lipids and not to liposomes/membranes. On the contrary, and as detailed in the manuscript, we set out to assay the lipid binding activity of TbMORN1, found that it appears to bind to lipids but not to liposomes/membranes, and have therefore cautioned that lipid or liposome/membrane binding of other MORN repeat proteins must be tested experimentally before claims of function are made.
  
  3) The physiological relevance of lipid binding to MORN proteins remains obscure (as also acknowledged by the authors). Does the binding of PE lipids to the MORN protein have a physiological role? Does the binding of fluorescent PI(4,5)P2 point to a physiological role of MORN proteins?
  
  These are interesting questions that we would like to address in future work.
  
  4) In light of recent data from the Chris Stefan lab (PMID: 31402097) a co-incidence detection of PI(4,5)P2, PS, and cholesterol seems possible. Can the authors address this possibility?
  
  Again, the involvement of cholesterol, PS, and PI(4,5)P2 would be interesting questions for subsequent work but are beyond the scope of the present study. We did partially address this issue in our use of PI(4,5)P2, POPC and cholesterol containing liposomes in liposome cosedimentation assays, which showed no binding (OV Fig. S3A [NV Fig. S4A]).
  
  Furthermore, the role of Ca2+ signaling / Ca2+ ions has not been addressed. In light of the important role of Ca2+ for the recognition of PI(4,5)P2 (PMID: 28177616), this point should be addressed.
  
  We carried out liposome pelleting assays in the presence of Ca2+ and Mg2+, and saw no binding by TbMORN1(2-15) in either condition (see data below). These data were not included in the MS because of the insufficient number of technical replicates available.
  
  5) For characterizing the binding of lipids to MORN proteins, the authors use nonphysiological fluorescent and short-chain lipid analogues at concentrations, which are unlikely to occur for endogenous PIPs in the cytosol of cells. Why choosing such an artificial system? Why introducing this system at length, if other -less artifact-prone- assays are available? I would recommend to not feature this assay as prominently as it was in the current study.
  
  Our aim was to stick to using the same fluorophore throughout all the experiments. The choice of short-chain lipids was constrained by what was commercially available with the BODIPY TMR fluorophore. We have implemented the reviewer's suggestion in the manuscript, and the text associated with the fluorescence anisotropy assays has been considerably shortened. We are aware that the chosen concentration of the fluorescent lipids was out of physiological range, but the requirements of the fluorescence anisotropy itself necessitated a compromise. The possible shortcomings of the fluorescence anisotropy assays are, we believe, more than amply compensated by the LiMA data.
  
  6) How would PE find its way to the lipid binding region in MORN? Would it diffuse to the MORN protein via the aqueous phase or would the MORN protein pickup PE form membranes up collision? The authors should address this point, by separating the lipiddepleted MORN protein from donor-vesicles containing PE by a dialysis membrane. If PE would not find its way to the lipid binding site of MORN, this would imply that MORN protein can extract lipids only upon colliding with the membrane. What is the stoichiometry of PE to MORN?
  
  These are all interesting questions that we would like to pursue in subsequent work, but we feel that they are beyond the scope of the present study. Until we have conditions suitable for obtaining high yields and monodisperse populations of the full-length protein, which probably also necessitates developing conditions for controlled oligomerisation, it would be premature to start this. As to how it picks up PE: it is well known that specific lipid binding/chaperoning proteins can deliver their lipid cargo to other proteins. Additionally, proteins that bind lipids use hydrophobic domains to both interact with and sequester fatty acids and/or lipids from membranes. The literature is populated with lots of such examples. https://www.sciencedirect.com/science/article/pii/S0092867416310765.
  
  Despite my critique raised above, I agree with the authors that the data point in the direction that MORN proteins (or at least TbMORN1) does not directly bind to membranes. Their data, however, would still be consistent with a role as lipid transfer protein and a recruitment of MORN proteins to the membrane by other proteins. Have the authors performed any additional experiments in this direction? Also, the potential role of palmitoylation is only mentioned in the discussion (page 22), while palmitoylation would provide a simple means for membrane recruitment.
  
  We are glad that the reviewer concurs with our main conclusion. We agree, as noted in the discussion, that a role as a lipid transfer protein might still be possible, and this is something that we would like to pursue in follow-up work. We have not yet performed any additional experiments in this direction. Concerning palmitoylation, the predictions using the CSS-Palm software were always weak and ambiguous, and in addition the best candidate cysteine residue was Cys351, which is in our structure engaged in the disulphide bond observed in the C2 crystal form. We feel that this is something to keep in mind, but is not yet a strong enough hypothesis to pursue intensively.
  
  Minor Points:
  
  Figure 1B: The authors should provide information on the void volume of the column.
  
  Implemented in the figure legend (7.2 ml).
  
  Page 17, line 696-701: The authors point out that the C2 crystal form is stabilized by two disulfide bridges. The authors should comment on the physiological relevance of these disulfide bridges.
  
  Given the reducing environment of the cytosol, it is an open question as to whether these disulphide bridges exist in vivo. We would prefer not to speculate on this point, as we do not feel it would be productive.
  
  Page 18, line 734-740: The authors should provide data on the potential role of Zn2+ on MORN function in a physiological context. The section describing that the dimer is stabilized by Zn2+ ions (pages 18 and 19) lacks a discussion if Zn2+ are functionally relevant. There is only a beautiful sequence analysis and a discussion of the conservation of the Zn2+ coordinating residues. Can the authors perform Zn2+ titrations and SEC-MALS experiments (or alternatives such as SAXS) to show that Zn2+ indeed affects the oligomeric state of only the PfMORN, but not the other MORN proteins that form alternative dimers?
  
  The known requirement for zinc ions in Plasmodium growth was already noted (OV lines 992- 993, Marvin et al., 2012), and is, we believe, sufficient to address the issue of physiological relevance at this stage. The zinc ions are predicted to affect the architecture of the apicomplexan (Plasmodium, Toxoplasma) MORN1 protein dimers, not their oligomeric state. For PfMORN1, SEC-MALS and SAXS were carried out in 20 mM Tris-HCl pH 7.5, 100 mM NaCl with no zinc present. When EDTA was added, no change in behaviour of the protein was seen by SEC-MALS. When “TPEN”, a strong zinc chelator, was added, the protein precipitated in SEC-MALS experiments.
  
  Reviewer #1 (Significance):
  
  A putative role of MORN proteins in membrane and lipid binding is addressed. The view the MORN proteins bind directly to membranes is challenged. Structures of dimeric MORN proteins provide important insight into the modes of dimerization.
  
  There is a recent structure of MORN proteins (which is referenced by the authors), but I feel that additional structural work is important and justified. The work on membrane vs. lipid binding is important, but not sufficiently addressed in the current manuscript.
  
  We are glad that the reviewer finds the structural work important and justified, although we disagree with the reviewer’s assessment of the lipid binding. As noted in the previous paragraph, our data challenge the assumption that MORN repeat proteins directly bind membranes, and we feel that this alone is a significant conceptual advance.
  
  I would recommend to separate the study in two parts. The audience is likely to confused (or bored) by the lengthy discussion on whether or not MORN proteins bind lipids and or membrane or not.
  
  We would prefer to implement the reviewer's other suggestion, namely that the manuscript is considerably shortened and less focus given to the negative data on lipid binding.
  
  I am not an expert in structural biology, but have a fair understanding of structural biology. I have worked on lipid binding proteins and have a very good understanding of lipid/membrane-binding assays.
  
  Reviewer #2 (Evidence, reproducibility and clarity):
  
  Summary
  
  The manuscript describes an extensive and detailed investigation into the structure and function(s) of MORN domains. It has to be acknowledged that, despite the considerable amount of work reported, the conclusions are rather limited. From a technical viewpoint, the experiments have been appropriately executed and, generally, I concur with the conclusions drawn. However, the manuscript is over-long: in general, I would recommend concentrating on positive conclusions which can be drawn from the data and avoid excessive speculation or inference (some examples given below).
  
  We are glad that the reviewer is satisfied with the technical quality of the work and (in general) the validity of the conclusions. We acknowledge that the original submission was fairly long, and have considerably shortened the revised manuscript and focused more on the positive conclusions in order to implement this suggestion.
  
  Major Comments
  
  There are three general- perhaps rather obvious- points to make. First, there is no particular reason to think that conservation of structure necessarily indicates conservation of a particular function. There seems to be an implicit assumption that MORN domains are associated with a specific, well-defined biological function. Given their diversity, are there particular reasons to think that this is the case?
  
  The reviewer is exactly right that there is an implicit assumption that MORN domains are associated with a specific, well-defined function: specifically, lipid binding. It is this assumption, which has been widely circulated in the almost complete absence of experimental evidence, that we are challenging. We agree that MORN repeats are likely to be capable of multiple functions, and protein-protein interactions are now better supported than protein-lipid interactions.
  
  Second, a strategy which examines the properties of just the recombinant MORN domains in vitro, removed from the context of the whole protein (eg junctophilin) or- importantly- its interacting partners in vivo, has obvious limitations. Frequently a reductionist approach is successful; however, in this case, MORN domains appear to be less tractable to that kind of approach. For all the in vitro binding and structural experiments presented, there is always a concern that the absence of other parts of the relevant MORN-containing protein or its partners could explain failure or inconsistency of in vitro biological activity measurements.
  
  Again, the reviewer is right that there is an inherent contextual limitation to any in vitro work that utilises a single protein, but this is a concern that - by definition - could be raised about any in vitro study utilising a single protein. It should be noted that we have also carried out in vivo experiments using TbMORN1 (OV Figs. 5, 6 [NV Figs. 3, 4]).
  
  Third, the possibility that MORN domains might mediate interactions with other proteins seems to be given little consideration, in spite of the Li et al (2019) paper. An experimental strategy which looked for binding partners (eg by pulldown assay) might have provided more insight.
  
  These data are already in the literature. A previous study by the same team (Morriswood et al., 2013) used proximity-dependent biotin identification to identify candidate binding partners and near neighbours of TbMORN1.
  
  In order to stress this point we added the following sentence in the discussion section, NV pages 18-19, lines 774-778.
  
  “The concluding data presented here suggest that TbMORN1 utilises this oligomerisation capacity to build mesh-like assemblies, which can reach considerable size in vitro (Fig. 7G). These mesh-like assemblies may reflect the endogenous organisation of the protein in vivo, where a number of binding partners have already been identified (Morriswood et al., 2013)”.
  
  Minor Comments
  
  In the abstract and elsewhere the authors refer to a possible function of MORN domains as 'dimerisation and oligomerisation devices' (line 53). What is the evidence that dimer formation is important for function in vivo?
  
  This is an interesting and important question and one that we would like to address in future work. We did attempt to generate trypanosome cell lines that inducibly expressed monomeric TbMORN1 (the double mutant, where the point mutations were simultaneously introduced in the dimerisation interface in repeats 13 and 14), but no expression of the ectopic protein was ever observed (9 separate clones obtained in 3 independent transfections). This might indicate the importance of the dimeric state in vivo, perhaps hinting that dimerisation is important for protection from degradation. In general, proteins assuming higher oligomeric states in homo- or heteromeric assemblies benefit from increased robustness in the cellular environment and optimised activity by the following means:
  
  Increased stability by decreasing the surface area/volume ratio
  
  Simple construction of larger complexes
  
  Allosteric regulation
  
  Co-localisation of distinct biological functions
  
  Substrate channelling
  
  Protection from aggregation or degradation
  
  Which or which combination of the factors is relevant for TbMORN1 being a functional dimer in vivo is difficult to say at this point.
  
  Did the authors attempt to co-crystallize TbMORN1(7-15) with PI(4,5)P2?
  
  No. For crystallisation, we used lysine methylated samples, and by doing this we neutralised positively-charged potential binding sites which would have interacted with the negatively charged lipid headgroup. We did not observe any bound lipids in the electron density maps obtained from the crystals.
  
  Fig 2C: did the authors also estimate binding stoichiometry as well as the equilibrium binding constants for these data? This should be determined by fitting a single binding site model to the data. Other methods (eg ITC) can probably determine this with more accuracy. The value of stoichiometry is sometimes forgotten in such binding measurements- is one ligand bound per monomer or dimer, for example?
  
  We discussed estimation of the binding stoichiometry in the fluorescence anisotropy assays at some length, but the conclusion was that the required experiments would contain too many approximations to provide high-confidence data. We did use ITC and also MST, but did not observe any binding with these assays.
  
  Lines 674-678 I found it hard to work out whether these constructs harbour the natural C-terminal sequence without truncation or addition of an affinity tag. I think the answer is 'yes' but it was difficult working this out from the details in M&M.
  
  TbMORN1(7-15) crystallisation was with a C-terminal Strep tag; TgMORN1(7-15) and PfMORN1(7-15) had their affinity tags removed by protease treatment prior to crystallisation. We have clarified this point in the M&M, page 29, lines 1189-1192: “Crystallisation of TbMORN1(7-15) (with a C-terminal Strep tag), TgMORN1(7-15) and PfMORN1(7-15) (both with affinity tags removed) was performed at 22 °C using a sitting-drop vapour diffusion technique and micro-dispensing liquid handling robots (Phoenix RE (Art Robbins Instruments) and Mosquito (TTP labtech).”
  
  Lines 688-694 The PISA interface analysis is useful here in distinguishing crystal contacts from those which persist in solution. The discussion of the results is unclear, however, on this critical point: were the dimer interfaces the only contacts which were significant in the various crystal forms?
  
  Yes, correct. PISA showed that the described dimerisation contacts were the only significant ones in the various crystal forms. Other crystals contacts had typically low P-values and poor ΔG and small “radar” surface in the complexive PISA analysis.
  
  In the case of both TbMORN1 crystal forms and in the case of the TgMORN1 P43212 crystal form we have a dimer in the asymmetric unit, while in the case of the PfMORN1 and TgMORN1 P6222 form we have one molecule in the asymmetric unit, and the dimer is created by the crystallographic twofold axis. In the latter cases the quaternary structure resulting from the symmetry operations was the top-scoring one considering either P-values and/or the number of stabilising interactions buried surface area.
  
  Lines 754-763 This paragraph seems rather speculative and is a good example where the text could be cut down.
  
  If the line citation is correct, then we disagree with this assessment and would prefer not to implement it. The paragraph in question concerns a detailed and very precise discussion of the side chain interactions that stabilise the V-shaped forms of TgMORN1 and PfMORN1.
  
  Line 765-788 This section is also rather overdone: such observations are only useful if they are subsequently tested by recording dimer conformation for a representative selection of MORN dimers from different species.
  
  Again, we disagree with the reviewer's assessment of this analysis. The analysis has considerable predictive power and already has some experimental validation via the SAXS observation that PfMORN1 is capable of forming extended dimers in solution (OV Fig. 10C [NV Fig. 7C]).
  
  Lines 800-801 I don't think this statement is strictly correct. The SAXS data show that PfMORN1(7-15) adopts an extended conformation, with no evidence of the 'V' shaped structure. Related to that point, from what I could glean from the SAXS Methods section, all solution conditions for these experiments were conducted without Zn2+? If some dimer interfaces require Zn2+, should it not be included?
  
  We have clarified this statement. The SAXS experiments were conducted without zinc, and, as we have stressed, the V-shaped form of TgMORN1 and PfMORN1 was only ever observed in the crystals. For PfMORN1, SEC-MALS and SAXS were carried out in 20 mM Tris-HCl pH 7.5, 100 mM NaCl with no zinc present. When EDTA was added, no change in behaviour of the protein was seen by SEC-MALS. When “TPEN”, a strong zinc chelator, was added, the protein precipitated in SEC-MALS experiments.
  
  Reviewer #2 (Significance):
  
  There is certainly value in establishing that MORN domains do not, in vitro, appear to bind to lipid vesicles, and to define their lipid binding capability (although it is rather complex). The crystal structures and SAXS data extend the rather limited structural data on MORN domains. Despite the effort involved, conclusions about likely functions of MORN domains in vivo are rather limited.
  
  We are glad that the reviewer acknowledges the value in challenging the assumption that MORN repeats are lipid binding devices, and that the structural data are important for expanding the knowledge base on this class of repeat motif proteins. In vivo functional work is being actively pursued at present.
  
  My expertise lies in X-ray crystallography and protein biochemistry.
  
  PeerReviewed
2. EMBOpress 16 Jun 2020
  
  in Review Commons
  
  Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.
  
  Learn more at Review Commons
  
  Referee #2
  
  Evidence, reproducibility and clarity
  
  Summary
  
  The manuscript describes an extensive and detailed investigation into the structure and function(s) of MORN domains. It has to be acknowledged that, despite the considerable amount of work reported, the conclusions are rather limited. From a technical viewpoint, the experiments have been appropriately executed and, generally, I concur with the conclusions drawn. However, the manuscript is over-long: in general, I would recommend concentrating on positive conclusions which can be drawn from the data and avoid excessive speculation or inference (some examples given below).
  
  Major Comments
  
  There are three general- perhaps rather obvious- points to make. First, there is no particular reason to think that conservation of structure necessarily indicates conservation of a particular function. There seems to be an implicit assumption that MORN domains are associated with a specific, well-defined biological function. Given their diversity, are there particular reasons to think that this is the case? Second, a strategy which examines the properties of just the recombinant MORN domains in vitro, removed from the context of the whole protein (eg junctophilin) or- importantly- its interacting partners in vivo, has obvious limitations. Frequently a reductionist approach is successful; however, in this case, MORN domains appear to be less tractable to that kind of approach. For all the in vitro binding and structural experiments presented, there is always a concern that the absence of other parts of the relevant MORN-containing protein or its partners could explain failure or inconsistency of in vitro biological activity measurements. Third, the possibility that MORN domains might mediate interactions with other proteins seems to be given little consideration, in spite of the Li et al (2019) paper. An experimental strategy which looked for binding partners (eg by pulldown assay) might have provided more insight.
  
  Minor Comments
  
  In the abstract and elsewhere the authors refer to a possible function of MORN domains as 'dimerisation and oligomerisation devices' (line 53). What is the evidence that dimer formation is important for function in vivo?
  
  Did the authors attempt to co-crystallize TbMORN1(7-15) with PI(4,5)P2?
  
  Fig 2C: did the authors also estimate binding stoichiometry as well as the equilibrium binding constants for these data? This should be determined by fitting a single binding site model to the data. Other methods (eg ITC) can probably determine this with more accuracy. The value of stoichiometry is sometimes forgotten in such binding measurements- is one ligand bound per monomer or dimer, for example?
  
  Lines 674-678 I found it hard to work out whether these constructs harbour the natural C-terminal sequence without truncation or addition of an affinity tag. I think the answer is 'yes' but it was difficult working this out from the details in M&M.
  
  Lines 688-694 The PISA interface analysis is useful here in distinguishing crystal contacts from those which persist in solution. The discussion of the results is unclear, however, on this critical point: were the dimer interfaces the only contacts which were significant in the various crystal forms?
  
  Lines 754-763 This paragraph seems rather speculative and is a good example where the text could be cut down.
  
  Line 765-788 This section is also rather overdone: such observations are only useful if they are subsequently tested by recording dimer conformation for a representative selection of MORN dimers from different species.
  
  Lines 800-801 I don't think this statement is strictly correct. The SAXS data show that PfMORN1(7-15) adopts an extended conformation, with no evidence of the 'V' shaped structure. Related to that point, from what I could glean from the SAXS Methods section, all solution conditions for these experiments were conducted without Zn2+? If some dimer interfaces require Zn2+, should it not be included?
  
  Significance
  
  There is certainly value in establishing that MORN domains do not, in vitro, appear to bind to lipid vesicles, and to define their lipid binding capability (although it is rather complex). The crystal structures and SAXS data extend the rather limited structural data on MORN domains. Despite the effort involved, conclusions about likely functions of MORN domains in vivo are rather limited. My expertise lies in X-ray crystallography and protein biochemistry.
  
  PeerReviewed
Visit annotations in context

Tags

PeerReviewed

Annotators

EMBOpress

URL

biorxiv.org/content/10.1101/826180v2
nixos.org nixos.org

Nixpkgs manual

1
1. toraritte 22 Jun 2020
  
  in Public
  
  fetchgit Used with Git. Expects url to a Git repo, rev, and sha256. rev in this case can be full the git commit id (SHA1 hash) or a tag name like refs/tags/v1.0.
  
  Not only is there no fetchgit (the right one is fetchGit), but there is also no sha256 argument.
  
  Backtracking: Got to IRC log https://logs.nix.samueldr.com/nixos/2018-08-14 (save on archive.org), search for Unsupported argument 'sha256' to 'fetchGit' (or part of it), and an answer will point to:<br> https://github.com/NixOS/nix/blob/master/src/libexpr/primops/fetchGit.cc#L198-L215
  
  We are again back to trying things out on hearsay.
  
  In the home-manager NixOS wiki it also shows a ref argument to fetchGit but it is not documented anywhere. Yay. Anyway, it works without it too.
Visit annotations in context

Annotators

toraritte

URL

nixos.org/nixpkgs/manual/
www.youtube.com www.youtube.com

How Can We Win Kimberly Jones Video Full Length David Jones Media Clean Edit #BLM 2020 What Can I Do

1
1. Carolwb 21 Jun 2020
  
  in Public
  
  On Saturday May 30th filmmaker and photographer David Jones of David Jones Media felt compelled to go out and serve the community in some way. He decided to use his art to try and explain the events that were currently impacting our lives. On day two, Sunday the 31st, he activated his dear friend author Kimberly Jones to tag along and conduct interviews. During a moment of downtime he captured these powerful words from her and felt the world couldn’t wait for the full length documentary, they needed to hear them now. Show less Show more
  
  video of Kimberly Jones who begins clearly and thoughtfully, "As long as we are focusing on the what we are not focusing on the why."
Visit annotations in context

Annotators

Carolwb

URL

youtube.com/watch
twitter.com twitter.com

Twitter

2
1. kgunanidhi1505 20 Jun 2020
  
  in Public
  
  The FBI said it has stopped using the "Black Identity Extremist" tag
  
  It is good to know that the FBI stopped using the "Black Identity Extremist" tag. I believe that the federal agencies like FBI should revisit their policies and procedures and ensure they stop using all these tags related to race, religion, origin, etc. to make the people and community safe and comfortable.
  
  I checked the root page's blue checkmark to ensure that is a proper and appropriate source of information. I also used the part of statements from this tweet to search google news and confirmed that this is a valid news.
2. tfuller656 19 Jun 2020
  
  in Public
  
  The FBI said it has stopped using the "Black Identity Extremist" tag and acknowledged that white supremacist violence is the biggest terrorist threat this country faces.
  
  I didn't know that the FBI was using "Black Identity Extremist" as a term, and I am glad they are stopping. Also, Ms. Pressley's face in the picture, says it all: "Its about time."
  
  I did check the website link out to check if it was fake by using Caulfield's "Wikipedia Method", and it was not. The link provided is for the online magazine "The Root."
Visit annotations in context

Annotators

kgunanidhi1505

tfuller656

URL

twitter.com/chribreuer
chem.libretexts.org chem.libretexts.org

Gas Chromatography

1
1. salman1993 20 Jun 2020
  
  in Public
  
  Applications Gas chromatography is a physical separation method in where volatile mixtures are separated. It can be used in many different fields such as pharmaceuticals, cosmetics and even environmental toxins. Since the samples have to be volatile, human breathe, blood, saliva and other secretions containing large amounts of organic volatiles can be easily analyzed using GC. Knowing the amount of which compound is in a given sample gives a huge advantage in studying the effects of human health and of the environment as well. Air samples can be analyzed using GC. Most of the time, air quality control units use GC coupled with FID in order to determine the components of a given air sample. Although other detectors are useful as well, FID is the most appropriate because of its sensitivity and resolution and also because it can detect very small molecules as well. GC/MS is also another useful method which can determine the components of a given mixture using the retention times and the abundance of the samples. This method be applied to many pharmaceutical applications such as identifying the amount of chemicals in drugs. Moreover, cosmetic manufacturers also use this method to effectively measure how much of each chemical is used for their products. Equations “Height equivalent to a theoretical plate” (HETP) use to calculate the flow rate by usingthe total number of theoretical plates (N) and column length (L). Some application, HETP concepts is used in industrial practice to convert number of theoretical plates to packing height. HETP can be calculate with the Van Deemter equation, which is given by HETP=A+Bυ+Cv(1)(1)HETP=A+Bυ+Cv HETP= A + \dfrac{B}{υ} + Cv \tag{1} Where A and B and C are constants and v is the linear velocity (carrier flow rate). A is the "Eddy-Diffusion" term and causes the broadening of the solute band. B is the "Longitudinal diffusion" term whereby the concentration of the analyte, in which diffuses out from the center to the edges.This causes the broadering of the analyte band. C is the "Resistance to Mass Transfer " term and causes the band of the analyte broader. HETP=LN(2)(2)HETP=LN HETP= \dfrac{L}{N} \tag{2} L is the length of the column, where N is the number of theoretical plates, tR is the retention time, and ω is the width of the elution peak at its base. N=16(tRω)2(3)(3)N=16(tRω)2 N= 16 \left (\dfrac{tR}{ω} \right)^2 \tag{3} In which, the more plates give a better resolution and more efficiency. Resolution can be determined by R=2[(tR)B–(tR)AWA+WB](4)(4)R=2[(tR)B–(tR)AWA+WB]R= 2\left[ \dfrac{(tR)B – (tR)A}{ WA +WB}\right] \tag{4} A relationship between the plates and resolution is giving by, R=(N)1/2/4)(α−1α)(1+K′BK′B)(5)(5)R=(N)1/2/4)(α−1α)(1+K′BK′B) R= (N)1/2 /4) ( \alpha -\dfrac{1}{\alpha}) ( 1+ \dfrac{K’B}{ K’B}) \tag{5} Where the selectivity, a, and k' is the capacity factors take places of the two solutes. The selectivity and capacity factors can be control by improving separation, such as changing mobile/ stationary phase composition, column temperature and use a special chemical effect. References Skoog, D. A.; Holler, F. J.; Crouch, S. R. Principles of Instrumental Analysis. Sixth Edition, Thomson Brooks/Cole, USA, 2007. Krugers, J. Instrumentation in Gas Chromatography. Centrex Publishing Company-Eindhoven, Netherlands, 1968. Hubschmann, H. Handbook of GC/MS: Fundamentals and Applications. Wiley-VCH Verlag, Germany, 2001. Scott, R. P. W. Chromatographic Detectors: Design, Function, and Operation. Marcel Dekker, Inc., USA, 1996. J.N. Driscoll. REview of Photoionization Detection in Gas Chromatography: The first Decade. Journal of CHromatographic Science , Vol 23. November 1985. 488-492. Boer, H. , "Vapour phase Chromatography", ed. Desty, D. H., 169 (Butterworths Sci. Pub., London, 1957). Dimbat, M. , Porter, P. E. , and Stross, F. H. , Anal. Chem., 28, 290 (1956). | Article | ISI | ChemPort | Contributors Kyaw Thet (UC Davis), Nancy Woo (UC Davis) /*<![CDATA[*/ $(function() { if(!window['autoDefinitionList']){ window['autoDefinitionList'] = true; $('dl').find('dt').on('click', function() { $(this).next().toggle('350'); }); } });/*]]>*/ /*<![CDATA[*/ var front = "auto"; if(front=="auto"){ front = "Gas Chromatography"; if(front.includes(":")){ front = front.split(":")[0]; if(front.includes(".")){ front = front.split("."); front = front.map((int)=>int.includes("0")?parseInt(int,10):int).join("."); } front+="."; } else { front = ""; } } front = front.replace(/_/g," "); MathJaxConfig = { TeX: { equationNumbers: { autoNumber: "all", formatNumber: function (n) { if(false){ return front + (Number(n)+false); } else{return front + n; } } }, macros: { PageIndex: ["{"+front+" #1}",1], test: ["{"+front+" #1}",1] }, Macros: { PageIndex: ["{"+front+" #1}",1], test: ["{"+front+" #1}",1] }, SVG: { linebreaks: { automatic: true } } } }; MathJax.Hub.Config(MathJaxConfig); MathJax.Hub.Register.StartupHook("End", ()=>{if(activateBeeLine)activateBeeLine()}); /*]]>*/ /*<![CDATA[*/window.addEventListener('load', function(){$('iframe').iFrameResize({warningTimeout:0, scrolling: 'omit'});})/*]]>*/ Back to top Chromatography High Performance Liquid Chromatography Recommended articles There are no recommended articles. 3.1: Principles of Gas ChromatographyNowadays, gas chromatography is a mature technique, widely used worldwide for the analysis of almost every type of organic compound, even those that a...10.23: ChromatographyChromatography is an efficient way for chemists to separate and analyze mixtures. Read on to find out how this critical process works.2.4: Gas Chromatography (GC)Gas chromatography (GC) is a powerful instrumental technique used to separate and analyze mixtures. A gas chromatograph is a standard piece of equipme...2.4D: Quantitating with GCPeak integrations are useful because it is possible to correlate the area under a peak to the quantity of material present in a sample. Note it is the...2.4A: Overview of GCGas chromato
  
  application
Visit annotations in context

Annotators

salman1993

URL

chem.libretexts.org/Bookshelves/Analytical_Chemistry/Supplemental_Modules_(Analytical_Chemistry)/Instrumental_Analysis/Chromatography/Gas_Chromatography
en.wikipedia.org en.wikipedia.org

The Root (magazine) - Wikipedia

1
1. tfuller656 19 Jun 2020
  
  in Public
  
  The Root (magazine) From Wikipedia, the free encyclopedia
  
  I used this Wikipedia page to check the legitamacy of website cited by the Twitter post "The FBI said it has stopped using the "Black Identity Extremist" tag and acknowledged that white supremacist violence is the biggest terrorist threat this country faces."
  
  The conclusion was that it a real website called "The Root."
Visit annotations in context

Annotators

tfuller656

URL

en.wikipedia.org/wiki/The_Root_(magazine)
www.nature.com www.nature.com

Parallel, tag-directed assembly of locally derived short sequence reads

3
1. edampf 19 Jun 2020
  
  in BehSci
  
  Hiatt, J., Patwardhan, R., Turner, E. et al. Parallel, tag-directed assembly of locally derived short sequence reads. Nat Methods 7, 119–122 (2010). https://doi.org/10.1038/nmeth.1416
  
  is:article lang:en DNA sequencing metagenomics next-generation sequencing library construction
2. edampf 19 Jun 2020
  
  in BehSci
  
  We demonstrate subassembly, an in vitro library construction method that extends the utility of short-read sequencing platforms to applications requiring long, accurate reads. A long DNA fragment library is converted to a population of nested sublibraries, and a tag sequence directs grouping of short reads derived from the same long fragment, enabling localized assembly of long fragment sequences. Subassembly may facilitate accurate de novo genome assembly and metagenome sequencing.
  
  ann:summary
3. edampf 19 Jun 2020
  
  in BehSci
  
  Parallel, tag-directed assembly of locally derived short sequence reads
  
  ann:title
Visit annotations in context

Tags

next-generation sequencing

DNA sequencing

ann:title

metagenomics

is:article

lang:en

ann:summary

library construction

Annotators

edampf

URL

nature.com/articles/nmeth.1416
www.educative.io www.educative.io

Prepare for a Coding Interview - Interactive Courses - Educative

1
1. dylansong 17 Jun 2020
  
  in Public
  
  The CSS above will ONLY select the h1 and h2 within the div. The other h1 and h2 within the p tag will be left unstyled.
  
  父级选择器用空格
  
  css html
Visit annotations in context

Tags

css

html

Annotators

dylansong

URL

educative.io/courses/the-complete-advanced-guide-to-css/JYlM1noxJ9o
inst-fs-iad-prod.inscloudgate.net inst-fs-iad-prod.inscloudgate.net

25 Years of Ed Tech

1
1. jhh1899 12 Jun 2020
  
  in Public
  
  I find key commands quite helpful when annotating: I use tabs to click between this block and the tag block. I use CTRL+Enter to submit my annotation. Makes life much much much better! :)
Visit annotations in context

Annotators

jhh1899

URL

inst-fs-iad-prod.inscloudgate.net/files/c36e1a8e-7065-49aa-99ab-40a2a1d30f0b/Weller_2020-25_Years_of_Ed_Tech-VRH6l.pdf
support.rentman.io support.rentman.io

UPDATE - Filters and presets

1
1. Lode 10 Jun 2020
  
  in Public
  
  Tags are labels that you can use to group your projects, equipment items, contacts, crew members, vehicles, invoices and tasks. First, you add a tag to your item(s) in a module. Afterwards, you can filter the shown items in a module by selecting your tag in the top right at label Tags expand_more.
  
  Style (no italics) + remove image
Visit annotations in context

Annotators

Lode

URL

support.rentman.io/hc/en-us/articles/360014345720-Filters-and-presets
www.sciencedirect.com www.sciencedirect.com

Metabolic engineering and classic selection of the yeast Candida famata (Candida flareri) for construction of strains with enhanced riboflavin production

1
1. voros.suleimaan 09 Jun 2020
  
  in Public
  
  Currently, the mutant of the flavinogenic yeast Candida famata dep8 isolated by classic mutagenesis
  
  No __notes were added.
  
  note riboflavin tag zib
Visit annotations in context

Tags

zib

tag

riboflavin

note

Annotators

voros.suleimaan

URL

sciencedirect.com/science/article/abs/pii/S1096717610000960
blog.jonudell.net blog.jonudell.net

hypothesis – Jon Udell

1
1. diegodlh 08 Jun 2020
  
  in Public
  
  A Firefox/Hypothesis extension has been in the works for quite a while,
  
  I published an unofficial Firefox extension here. Just download and install the XPI file. Go here for a discussion about it. So far only thing I found that doesn't work is annotating local PDFs, because Firefox is more restrictive about this than Chrome.
Visit annotations in context

Annotators

diegodlh

URL

blog.jonudell.net/tag/hypothesis/
shakespeare.mit.edu shakespeare.mit.edu

SCENE I. An apartment in the DUKE'S palace.

1
1. voros.suleimaan 08 Jun 2020
  
  in Public
  
  properties to 'unfold'
  
  boo tag mag zip
Visit annotations in context

Tags

zip

tag

mag

boo

Annotators

voros.suleimaan

URL

shakespeare.mit.edu/measure/measure.1.1.html
moz.com moz.com

Technical SEO – The Beginner's Guide to SEO

1
1. HollywoodHappenings 05 Jun 2020
  
  in Public
  
  How search engines understand websites Imagine being a search engine crawler scanning down a 10,000-word article about how to bake a cake. How do you identify the author, recipe, ingredients, or steps required to bake a cake? This is where schema markup comes in. It allows you to spoon-feed search engines more specific classifications for what type of information is on your page.Schema is a way to label or organize your content so that search engines have a better understanding of what certain elements on your web pages are. This code provides structure to your data, which is why schema is often referred to as “structured data.” The process of structuring your data is often referred to as “markup” because you are marking up your content with organizational code.JSON-LD is Google’s preferred schema markup (announced in May ‘16), which Bing also supports. To view a full list of the thousands of available schema markups, visit Schema.org or view the Google Developers Introduction to Structured Data for additional information on how to implement structured data. After you implement the structured data that best suits your web pages, you can test your markup with Google’s Structured Data Testing Tool.In addition to helping bots like Google understand what a particular piece of content is about, schema markup can also enable special features to accompany your pages in the SERPs. These special features are referred to as "rich snippets," and you’ve probably seen them in action. They’re things like:Top Stories carouselsReview starsSitelinks search boxesRecipesRemember, using structured data can help enable a rich snippet to be present, but does not guarantee it. Other types of rich snippets will likely be added in the future as the use of schema markup increases.Some last words of advice for schema success:You can use multiple types of schema markup on a page. However, if you mark up one element, like a product for example, and there are other products listed on the page, you must also mark up those products.Don’t mark up content that is not visible to visitors and follow Google’s Quality Guidelines. For example, if you add review structured markup to a page, make sure those reviews are actually visible on that page.If you have duplicate pages, Google asks that you mark up each duplicate page with your structured markup, not just the canonical version.Provide original and updated (if applicable) content on your structured data pages.Structured markup should be an accurate reflection of your page.Try to use the most specific type of schema markup for your content.Marked-up reviews should not be written by the business. They should be genuine unpaid business reviews from actual customers.Tell search engines about your preferred pages with canonicalizationWhen Google crawls the same content on different web pages, it sometimes doesn’t know which page to index in search results. This is why the rel="canonical" tag was invented: to help search engines better index the preferred version of content and not all its duplicates.The rel="canonical" tag allows you to tell search engines where the original, master version of a piece of content is located. You’re essentially saying, "Hey search engine! Don’t index this; index this source page instead." So, if you want to republish a piece of content, whether exactly or slightly modified, but don’t want to risk creating duplicate content, the canonical tag is here to save the day.
  
  How do websites communicate with search engines is there a special code that
Visit annotations in context

Annotators

HollywoodHappenings

URL

moz.com/beginners-guide-to-seo/technical-seo
docs.gitlab.com docs.gitlab.com

GitLab Markdown | GitLab

1
1. TylerRick 04 Jun 2020
  
  in Public
  
  You can add a table of contents to a Markdown file, wiki page, or issue/merge request description, by adding the tag [[_TOC_]] on its own line. It will appear as an unordered list that links to the various headers.
  
  table of contents GitLab-Flavored Markdown
Visit annotations in context

Tags

GitLab-Flavored Markdown

table of contents

Annotators

TylerRick

URL

docs.gitlab.com/ee/user/markdown.html
May 2020
github.com github.com

csae8092/posts

1
1. Ciceronianus 28 May 2020
  
  in Public
  
  <div class="templates:surround?with=templates/page.html&at=content"> <h1>Table of Contents</h1> <div data-template="app:toc"/> </div>
  
  This seems to combine two different ways of HTML templating:
  
  class
  
  data-tag
  
  Should it be done in this way?
  
  eXist-db
Visit annotations in context

Tags

eXist-db

Annotators

Ciceronianus

URL

github.com/csae8092/posts/blob/master/digital-edition-web-app/part-3-table-of-content.md
cdli-gh.github.io cdli-gh.github.io

translation_pipeline

1
1. epageperron 27 May 2020
  
  in Public
  
  -ta most likely to be verbs
  
  -ta are most likely ablatives. That's why we use those particles to tag morphology. Checkout the morphology charts.
  
  For Ur III sumerian, what is most telling about the nature of the word is its placement in the text. Suffixes are rarely present as compared to other genres.
Visit annotations in context

Annotators

epageperron

URL

cdli-gh.github.io/blog/gsoc20/translation_pipeline/index
gitlab.com gitlab.com

Index · Container registry · Packages · User · Help

1
1. TylerRick 26 May 2020
  
  in Public
  
  Using the Git SHA in your image tag makes this less necessary since each job will be unique and you shouldn't ever have a stale image. However, it's still possible to have a stale image if you re-build a given commit after a dependency has changed.
  
  dependencies stale cache
Visit annotations in context

Tags

dependencies

stale cache

Annotators

TylerRick

URL

gitlab.com/help/user/packages/container_registry/index
hypothes.is hypothes.is

Hypothesis

1
1. duc 25 May 2020
  
  in Public
  
  GitHub
  
  why don't just just show the script tag to embed Hypothesis in any website. No need to "head to Github" to spend more time, hassles etc.
Visit annotations in context

Annotators

duc

URL

hypothes.is/docs/help
sogo-cas.uni-osnabrueck.de sogo-cas.uni-osnabrueck.de

Exposé_Schulze.pdf

1
1. NeleHenke 24 May 2020
  
  in Public
  
  „Ein Bild: der höchste Alpengipfel, ausgehauen zu einem Gesicht unter wuchtendem Stahlhelm, das still und ernst über die Lande schaut, den deutschen Rhein hiunter aufs freie Meer. –Einst wird kommen der Tag....“
  
  Ich finde es sehr schön, wenn die Passagen aus dem Buch in Absätzen und somit hervorgehoben zitiert werden! Aber hier zitierst Du jetzt anders als in Deiner vorherigen Buchstelle (S.3). Hier benutzt Du Anführungszeichen und beendest mit einfachen Doppelpunkten ohne die eckigen Klammern. Eventuell nochmal darauf achten, dass dies alles einheitlich gemacht wird.
Visit annotations in context

Annotators

NeleHenke

URL

sogo-cas.uni-osnabrueck.de/SOGo/so/nehenke/Mail/0/folderINBOX/1123/3/Exposé_Schulze.pdf
www.oercamp.de www.oercamp.de

Screencasts und kommentierte Screenshots zur Tool-Einführung für Lernende

1
1. neleh 23 May 2020
  
  in Public
  
  am Tag nach dem Webinar hier.
  
  Falls Du schon vorab nähere Informationen suchst, empfehle ich Dir diesen Blogbeitrag im eBildungslabor
Visit annotations in context

Annotators

neleh

URL

oercamp.de/webinare/weboep/screen/
featuredcontent.psychonomic.org featuredcontent.psychonomic.org

Boosting COVID-19 related behavioral science by feeding and consulting an eclectic knowledge base

1
1. edampf 22 May 2020
  
  in BehSci
  
  Herzog, S. (2020, May 21). Boosting COVID-19 related behavioral science by feeding and consulting an eclectic knowledge base. Psychonomic Society Featured Content. https://featuredcontent.psychonomic.org/boosting-covid-19-related-behavioral-science-by-feeding-and-consulting-an-eclectic-knowledge-base/
  
  is:blog lang:en COVID-19 behavioral science boosting eclectic knowledge base database information sharing SciBeh research annotation tag filter search gitlab
Visit annotations in context

Tags

gitlab

database

research

SciBeh

annotation

tag

lang:en

knowledge base

boosting

COVID-19

behavioral science

search

is:blog

filter

eclectic

information sharing

Annotators

edampf

URL

featuredcontent.psychonomic.org/boosting-covid-19-related-behavioral-science-by-feeding-and-consulting-an-eclectic-knowledge-base/
stackoverflow.com stackoverflow.com

Do Git tags only apply to the current branch?

1
1. kip2 21 May 2020
  
  in Public
  
  git describe [--tags] describes the current branch in terms of the commits since the most recent [possibly lightweight] tag in this branch's history. Thus, the tag referenced by git describe may NOT reflect the most recently created tag overall.
  
  git tags releases
Visit annotations in context

Tags

releases

tags

git

Annotators

kip2

URL

stackoverflow.com/questions/14613540/do-git-tags-only-apply-to-the-current-branch
edtechfactotum.com edtechfactotum.com

Hypothesis annotation notification – a feature request

1
1. diegodlh 20 May 2020
  
  in Public
  
  You can get an RSS or Atom feed for annotations made at a specific url, a specific Hypothesis tag or group, or for a specific Hypothesis user.
  
  Domain-level query is also available with RSS and Atom feeds. This tool here makes the process of getting these feed urls a bit easier.
  
  rss notifications
Visit annotations in context

Tags

rss

notifications

Annotators

diegodlh

URL

edtechfactotum.com/hypothesis-annotation-notification-a-feature-request/
stats.hackmag.com stats.hackmag.com

Опасный тренд. Как утилита Trend Micro для борьбы с руткитами позволила устанавливать руткиты

1
1. Vega 20 May 2020
  
  in Public
  
  Tag` в экземпляре `MicroScan`
  
  Выделить или хотя бы убрать `
Visit annotations in context

Annotators

Vega

URL

stats.hackmag.com/pdf/298462.edit
support.rentman.io support.rentman.io

UPDATE - Using tags

2
1. Lode 20 May 2020
  
  in Public
  
  Filter by tag
  
  Change image, add tags button and change some text.
2. Lode 20 May 2020
  
  in Public
  
  Afterwards, you can filter the shown items in a module by clicking on the label tags expand_more button and selecting your tag(s).
  
  Change sentence and add tags button
Visit annotations in context

Annotators

Lode

URL

support.rentman.io/hc/en-us/articles/360013931999-Organize-with-tags-
support.google.com support.google.com

IP Anonymization (or IP masking) in Analytics - Analytics Help

1
1. TylerRick 19 May 2020
  
  in Public
  
  The Analytics JavaScript Tag When a JavaScript-enabled web browser loads a page with the Analytics tag (ga.js or analytics.js), it does two things asynchronously: load and process the Analytics function queue and request the Analytics JavaScript. The function queue is a JavaScript array where the different Analytics configuration and collection functions are pushed. These functions, which are set by the site owner when implementing Analytics can include functions like specifying the Analytics account number and actually sending page view data to the Analytics Collection Network for processing. When the Analytics JavaScript runs a function from the function queue that triggers data to be sent to the Analytics Collection Network (this function is typically ga('send', 'pageview') in the analytics.js JavaScript library and _trackPageview in the ga.js library), it sends the data as URL parameters attached to an HTTP request for http://www.google-analytics.com/_utm.gif (for ga.js) and http://www.google-analytics.com/collect (for analytics.js). If the anonymization function has been called prior to the page tracking function, an additional parameter is added to the pixel request. The IP anonymization parameter looks like this: &aip=1 The Analytics Collection Network The Analytics Collection Network is the set of servers that provide two main services: the serving of ga.js and analytics.js (the Analytics JavaScript) and the collection of data sent via requests for _utm.gif and /collect. When a request for ga.js, analytics.js, _utm.gif, or /collect arrives, it includes additional information in the HTTP request header (i.e. the type of browser being used) and the TCP/IP header (i.e. the IP address of the requester). As soon as a request for _utm.gif arrives, it is held in memory for anonymization. If the &aip=1 parameter is found in the request URL (as it would have been placed by the Analytics JavaScript after processing the anonymization function in ga.js or analytics.js ), then the last octet of the user IP address is set to zero while still in memory. For example, an IP address of 12.214.31.144 would be changed to 12.214.31.0. (If the IP address is an IPv6 address, the last 80 of the 128 bits are set to zero.) Only after this anonymization process is the request written to disk for processing. If the IP anonymization method is used, then at no time is the full IP address written to disk as all anonymization happens in memory nearly instantaneously after the request has been received.
  
  detailed explanation
Visit annotations in context

Tags

detailed explanation

Annotators

TylerRick

URL

support.google.com/analytics/answer/2763052
www.analyticsmania.com www.analyticsmania.com

Setup GDPR Cookie Consent Notification with Google Tag Manager - Analytics Mania

1
1. TylerRick 19 May 2020
  
  in Public
  
  To be fully compliant with GDPR, you would also need to enable Show Reject All Button setting.
  
  consent: must be as easy to withdraw/reject as it is to give/accept GDPR
Visit annotations in context

Tags

consent: must be as easy to withdraw/reject as it is to give/accept

GDPR

Annotators

TylerRick

URL

analyticsmania.com/post/gdpr-cookie-consent-notification-with-google-tag-manager/
h.readthedocs.io h.readthedocs.io

Hypothesis API documentation

1
1. diegodlh 15 May 2020
  
  in Public
  
  contain
  
  Isn't there a way for an exact tag match? I mean, a way in which "web" would only match a "web" tag, and not a "web annotation" tag too, for example.
Visit annotations in context

Annotators

diegodlh

URL

h.readthedocs.io/en/latest/api-reference/
www.projekt-gutenberg.org www.projekt-gutenberg.org

Novellen

1
1. andreas.gencyilmaz 15 May 2020
  
  in Public
  
  Und Tag für Tag nahm die Pest zu, die Sommersonne brannte auf die Stadt herab, es fiel kein Regentropfen, es rührte sich kein Wind,
  
  In den ersten Wochen während des Fernunterrichts war das Wetter ähnlich: Kein Regen für eine sehr lange Zeit, die Sonne schien; es war sehr warm (insbesondere für März/April).
Visit annotations in context

Annotators

andreas.gencyilmaz

URL

projekt-gutenberg.org/jacobsen/novelle1/chap005.html
support.rentman.io support.rentman.io

UPDATE - Set up QR codes and barcodes

1
1. parkiepants 13 May 2020
  
  in Public
  
  (Optional) Click save 'Save to item' to connect this tag to its equipment
  
  new line, CSS
Visit annotations in context

Annotators

parkiepants

URL

support.rentman.io/hc/en-us/articles/360013101260-UPDATE-Set-up-QR-codes-and-barcodes
developers.google.com developers.google.com

Loading Third-Party JavaScript | Web Fundamentals | Google Developers

1
1. TylerRick 12 May 2020
  
  in Public
  
  A "tag" is a snippet of code that allows digital marketing teams to collect data, set cookies or integrate third-party content like social media widgets into a site.
  
  This is a bad re-purposing of the word "tag", which already has specific meanings in computing.
  
  Why do we need a new word for this? Why not just call it a "script" or "code snippet"?
  
  unfortunate definition using an alternate dumbed-down/user-friendly name for something language: misuse of word words words to avoid using
Visit annotations in context

Tags

unfortunate

definition

language: misuse of word

words

using an alternate dumbed-down/user-friendly name for something

words to avoid using

Annotators

TylerRick

URL

developers.google.com/web/fundamentals/performance/optimizing-content-efficiency/loading-third-party-javascript
Local file Local file

Untitled

1
1. daweilai 12 May 2020
  
  in Public
  
  习经济学要目标高，因为偏向理论研究，所以本科就是基础阶段，硕士、博士才是追求。2、经济学的理论是有趣的，但是这些理论的产生过程是非常乏味的，在四年本科学习中，我们要花费大量的时间与表格、数据、数字模型打交道。你要做好这个思想准备。3、如果想深入学习这个专业，我个人的看法是必须本科双专业，也就是经济
  
  this is new annotation from daweilai
  
  这是tag 这是tag2
Tags

这是tag

这是tag2

Annotators

daweilai
support.rentman.io support.rentman.io

UPDATE - Using Tags

4
1. parkiepants 11 May 2020
  
  in Public
  
  Filter by tag
  
  Deleted screenshots, texbox, and explanation (one and all)
2. parkiepants 11 May 2020
  
  in Public
  
  Add a tag
  
  Deleted screenshots and text box about what you can do in widgets
3. parkiepants 11 May 2020
  
  in Public
  
  Don't forget to press enter before you click Save. Otherwise, your tag will not be added.
  
  "press enter", CSS Save
4. parkiepants 11 May 2020
  
  in Public
  
  Choose an existing tag to apply, search for an existing tag to apply, or type in a new tag to apply and press enter.
  
  new
Visit annotations in context

Annotators

parkiepants

URL

support.rentman.io/hc/en-us/articles/360013760599-UPDATE-Using-Tags
lxieyang.github.io lxieyang.github.io

Michael Xieyang Liu

1
1. ahorvath 06 May 2020
  
  in Public
  
  My current research interests include human-computer interaction
  
  test
  
  tag
Visit annotations in context

Tags

tag

Annotators

ahorvath

URL

lxieyang.github.io/
darkblueheaven.com darkblueheaven.com

Spatial Software

1
1. jborichevskiy 03 May 2020
  
  in Public
  
  Within minutes of testing out the app with me, my friends discovered we could play tag for fun, or have little footraces around the map. Behaviors I didn't design into the app at all.
  
  Cool example of emergent behavior in software
Visit annotations in context

Annotators

jborichevskiy

URL

darkblueheaven.com/spatialsoftware/
academic.oup.com academic.oup.com

Editorial: We need to talk about authorship

1
1. robeirne 01 May 2020
  
  in Gigascience Annotations
  
  For example, if two groups collaborate on a project, it can make sense that there is a primary contact for each group.
  
  How would this be modelled in the underlying JATS XML data?
  
  Although many journals specify only one corresponding author, in other journals there is no limit to the number of contributors who may be designated to receive correspondence for the article. Accordingly, more than one <contrib> element may have its @corresp attribute set to “yes”.
  
  From: https://jats.nlm.nih.gov/articleauthoring/tag-library/1.1d2/attribute/corresp.html
Visit annotations in context

Annotators

robeirne

URL

academic.oup.com/gigascience/article/7/12/giy122/5114264
Apr 2020
safetyholic.com safetyholic.com

Online Safety Tips: Things Not to Do Online · Tips & Tricks · Safetyholic. Internet Security & Privacy Online

1
1. TylerRick 30 Apr 2020
  
  in Public
  
  Don’t share your children’s photos with their real names Adults are able to un-tag themselves from images that they don’t want to be identified in, but children don’t have that option. A lot of parents these days are referring to their children as a hashtag or a nickname which protects their identity without taking the fun out of sharing family photos. This also has an added bonus of giving the child a clean slate when they are older and building their own web presence, instead of being able to Google their name and seeing hundreds of baby photos that their relatives posted in the past.
  
  protecting your kids online
Visit annotations in context

Tags

protecting your kids online

Annotators

TylerRick

URL

safetyholic.com/online-safety-tips-things-not-to-do-online/
www.iubenda.com www.iubenda.com

Prior Blocking of Cookies: Manual Tagging (with Practical Examples)

1
1. TylerRick 29 Apr 2020
  
  in Public
  
  Google Tag Manager allows you to avoid tagging scripts as described below, although this is limited to a certain category of scripts – scripts that are not positional/do not define a position. It, therefore, does not handle embed scripts such as those related to advertising banners, youtube video widgets, facebook like buttons etc.
  
  cookie blocking Google Tag Manager caveat limitations
Visit annotations in context

Tags

limitations

caveat

cookie blocking

Google Tag Manager

Annotators

TylerRick

URL

iubenda.com/en/help/1229-manual-tagging-blocking-cookies
annotate-hypothesis.herokuapp.com annotate-hypothesis.herokuapp.com

Text Annotation task

1
1. mturk.annotate 28 Apr 2020
  
  in Public
  
  won the CWA World Tag Team Championship
  
  More men than women in pro wrestling.
Visit annotations in context

Annotators

mturk.annotate

URL

annotate-hypothesis.herokuapp.com/study/22/313632/
www.samanthacasolaro.com www.samanthacasolaro.com

Untitled document

1
1. sc_geier 27 Apr 2020
  
  in Public
  
  Click on each price tag to learn about the costs of fashion.
  
  I like the illustrations, but as an interactive graphic, they feel a bit arbitrary and repetitive. What distinguishes one illustration from another, in terms of how they introduce their sections? Make unique illustrations for each section, e.g., a tree for Deforestation.
Visit annotations in context

Annotators

sc_geier

URL

samanthacasolaro.com/final_project/
stanbol.apache.org stanbol.apache.org

Apache Stanbol - Welcome to Apache Stanbol!

1
1. raj_reddy 25 Apr 2020
  
  in Public
  
  direct usage from web applications (e.g. for tag extraction/suggestion; or text completion in search fields), 'smart' content workflows or email routing based on extracted entities, topics, etc.
  
  autocomplete apache stanbol semantics
Visit annotations in context

Tags

apache stanbol

autocomplete

semantics

Annotators

raj_reddy

URL

stanbol.apache.org/
fortelabs.com fortelabs.com

Tagging is Broken

1
1. BillyPilgrim 25 Apr 2020
  
  in Public
  
  Thus the most problematic behavior is implicitly encouraged and enabled. Grrrrrrr. It could be a misspelling. It could be a slip of the finger. It could be a different capitalization, punctuation, or tense, whatever. No warning or indication is given, and a divergent tag is created, for you to hopefully notice and fix later, hopefully before you rely on it.
  
  This is a problem in the note taking app Roam. All it takes is a hashtag or two [[brackets]] around a word to create a new page/note.
  
  This leads to many dead pages from typos or just copypasted content with the hashtag symbol in them.
Visit annotations in context

Annotators

BillyPilgrim

URL

fortelabs.com/blog/tagging-is-broken/
www.nrn.com www.nrn.com

How restaurants are stepping up to help their communities during the coronavirus pandemic, from providing toilet paper to meal kits

1
1. brianne 21 Apr 2020
  
  in Public
  
  The community meals include menu options like prime rib, salmon, vegan stew, loaves of bread, Arcana’s homemade ice cream and various side dishes, costs $20 per person, with a sliding scale option for those who cannot afford the price tag.
  
  This menu appears to include traditionally high-cost items like prime rib and treats like ice cream. Interpretations: this restaurant is committed to providing all members of its community with healthy, hearty, and economical food options. Interpretations: $20/meal is still a hefty price tag when many fast food restaurants are able to provide a family of four a whole meal for $20.
Visit annotations in context

Annotators

brianne

URL

nrn.com/people/how-restaurants-are-stepping-help-their-communities-during-coronavirus-pandemic-providing
www.aare.edu.au www.aare.edu.au

Naomi Milthorpe – EduResearch Matters

1
1. laurapasquini 21 Apr 2020
  
  in Public
  
  ing was that the teacher-education students in our study did not see their set online tasks as being valuable to their
  
  FOllow up later
Visit annotations in context

Annotators

laurapasquini

URL

aare.edu.au/blog/
www.drugrehab.com www.drugrehab.com

Drug Use in Middle School

1
1. keelea 21 Apr 2020
  
  in Public
  
  Unfortunately, in their quest for peer acceptance, many middle schoolers believe that drinking or using drugs will make them more popular.
  
  While fitting in is very important to kids in middle school many kids are looking to develope the tag of "being popular". Many of these students believe that in order to make themselves more popular and socially accepted by their peers then they need to develope the habits of using drugs. This is a negative stance because while some may see themselves as being more popular what they don't see is the effect these substances have on their developement and their bodies.
Visit annotations in context

Annotators

keelea

URL

drugrehab.com/teens/middle-school/
www.biorxiv.org www.biorxiv.org

Glycolytic Protein Phosphofructokinase Relocalizes into SCompartments with Liquid-like Properties

3
1. EMBOpress 20 Apr 2020
  
  in Review Commons
  
  Note: This rebuttal was posted by the corresponding author to Review Commons. Content has not been altered except for formatting.
  
  Learn more at Review Commons
  
  Reply to the reviewers
  
  Reviewer #1:
  
  **Summary**
  
  Jang et al., address the important question of spatially localized or compartmentalized metabolic enzymes with a focus on the glycolytic enzyme PFK1. Using a good strategy of inserting a fluorescent tag at the endogenous PFK1 locus with tissue-specific inducible expression in C. elegans, combined with strong quantitative longitudinal imaging and innovative bioengineered microfluidic-hydrogels to control oxygen availability as well as optogenetic approaches, they show PFK1 condensates, which are not stress granules and not seen in normoxia, assemble with hypoxia. PFK1 condensates are dynamic, reversible, localized at the synapse in neurons, and recruit aldolase, another glycolytic enzyme. Although glycolytic proteins were previously shown to compartmentalize near the plasma membrane, and PFK1 was previously shown to assemble into filaments in vitro and be punctate at the plasma membrane in mammalian cells, evidence for cellular localized PFK1 condensates in animals is highly significant. The work includes strong biophysical characterization of PFK1 phase-separated condensates, but no clear indication of the composition of condensates. More significantly, the findings lack functional significance related to PFK1 activity or glycolytic flux with hypoxia vs normoxia. Despite previous work by this group showing that disrupting subcellular localization of glycolytic enzymes impairs neuronal activity in response with hypoxia, the reader is left with questions on the importance of localized and PFK1 condensates and their make-up .
  
  **Major comments:**
  
  Key conclusions are convincing, and most experimental approaches, biophysical characterization including thermodynamic principles, and data analysis are exemplary and well described. However, as indicated above, the work is limited to a descriptive analysis of cellular localization of PFK1 condensates and their biophysical properties without insights on functional significance relative to enzyme activity - or at least glycolytic flux or metabolic reprogramming with hypoxia. At best, only correlations can be drawn from hypoxia-induced localized PFK1 condensates and the authors' previous report (Jang et al., 2016) on hypoxia-regulated neuronal activity. Some insight or at least prediction in the discussion on the differences in spatially localized PFK1 in muscle vs neurons with regard to metabolic or energy distinctions should be included.
  
  We have added additional discussions on the differences of the spatially localized PFK-1.1 in muscles versus neurons, explaining that in both tissues the cellular enrichment appears to be at sites predicted to have high ATP consumption (lines 128-133; 482-484).
  
  Despite the strong biophysical analysis of condensates, several important features are not determined. First is at best a rudimentary analysis of the composition of condensates and also how PFK1 is assembled into these structures. For the former, is the core of the condensate predominantly PFK1 with perhaps aldolase only recruited to the periphery or is aldolase an integral component of the structure. Hence, is it a PFK1 condensate or a glycolytic condensate? For the latter question, is there a particular orientation for PFK1 in condensates, i.e a collection of filaments as previously reported, which might provide insight on assembly? Finally, and less critical but also important is the criterion for spherical, which is not well defined, and at least some idea or speculation on determinants for a spherical morphology - compared with filaments that have been reported for other non-glycolytic metabolic enzymes.
  
  We have now co-expressed PFK-1.1 and ALDO-1 and examined their dynamic formation during hypoxic conditions. We observe PFK-1.1 and ALDO-1 form condensates simultaneously, with gradual enrichment of both molecules. We now include this new data in Figure 7E and Video 8; lines 422-441, 964-989). We also include genetic data demonstrating the ALDO-1 requires pfk-1.1 to form condensates, and that PFK-1.1 requires aldo-1 as well. Therefore, the enzymes are interdependent on each other to form condensates (Figures 7G, 7H, S7B, and S7C).
  
  The spheroid geometry reflects liquid-like properties, which arises from surface tension of molecules loosely held together via multi-valent interactions. Filamentous arrangements reflect crystalline-like structures resulting from more stable interactions between molecules into solid-like states. While we did not perform high resolution studies, like Cryo-EM, to resolve this question, the spheroid geometry of PFK-1.1 condensates, along with its fluid-like properties, suggest the condensates are liquid-like compartment distinct to filamentous structures. We now add this discussion in lines 467-470.
  
  The work is an important advance in our understanding on the self-assembly of metabolic enzymes by showing hypoxia-induced PFK1 condensates in vivo, their spatially-restricted subcellular localization in muscle cells and neurons, and their biophysical properties, the latter being distinct from those of stress granules. Taken together, these findings are more extensive than many previous reports on the assembly of metabolic enzymes into filaments or condensates, but fall short for new insights on functional significance.
  
  We focus this study on the biophysical characterization of the condensates, and how that results in compartmentalized enrichment of glycolytic proteins. Examination of the functional significance of the phase separation to the enzymatic reactions in vivo is not currently possible because we lack probes we can use in vivo to measure the metabolites resulting from the reaction. We have now added discussion acknowledging this and framing its significance in the context of what has been published in the field (lines 484-492). For example, a recent manuscript in ChemRxiv demonstrated, in vitro, that the enzymatic activity of glycolytic proteins, hexokinase and glucose-6 phosphate dehydrogenase, promote these enzymes condensing into liquid droplets. The authors further found that the condensation accelerated the glycolytic reactions (Ura et al., 2020). This raises the question whether glycolytic proteins compartmentalize, and form condensates, in vivo, which we address in this manuscript. We capture this point in (lines 444-464) where we explain that, while it has long been hypothesized that glycolytic proteins like PFK-1 could be compartmentalized, this remained controversial due to lack of dynamic in vivo imaging. In our study, and through a systematic examination of endogenous PFK-1.1 via the use of a hybrid microfluidic-hydrogel device, we conclusively determine that PFK-1.1 indeed displays distinct patterns of subcellular localization in specific tissues in vivo.
  
  Reviewer #2 (Evidence, reproducibility and clarity (Required)):
  
  This paper reports on the condensation of the glycolytic enzyme PFK-1 in response to hypoxic conditions in neurons of C. elegans. The authors employ a microfluidic-hydrogel device to dynamically monitor the relocalisation of PFK-1 from a mostly diffuse state to clusters in response to hypoxia and show that PFK-1 can undergo multiple rounds of PFK-1 clustering and dissolution. The authors work through the key features of a liquid-like compartment (sphericity, fusion, fast internal rearrangements) and give evidence that PFK-1 may have all three. Finally, the authors tag PFK-1 with the light-inducible multimerization domain Cry2 and find that even without light PFK-1 will constitutively form clusters that sequestrate endogenous PFK-1 as well as other glycolytic proteins. The strength of this work is that it is characterizing what appears to likely be phase separation in the context of a whole animal experiencing a stress that it could encounter in the natural world. A limitation of the work is that it is unclear what the functional implications are of condensates of PFK-1 at the molecular or cell scale.
  
  **Major comments:**
  
  -All experiments were performed using fluorescently tagged PFK-1 expressed from endogenous promoter or from the native genetic locus which is important for excluding overexpression artifacts. However, there is still risk that the GFP tag is driving the assembly process. In order to exclude tag-specific effects that may cause aggregation of the tetrameric PFK-1, ideally a control would be done in which PFK-1 is visualized through immunofluorescence experiments of WT cells. Alternatively, a short tag (e.g HA, His) as epitope for is an alternative .
  
  We used fluorescent tags to observe the dynamic relocalization in vivo. While in the study we have not performed immunofluorescence, we established the validity of the labeling method by: 1) using monomeric versions of GFP; 2) using different fluorophores to show the same condensation phenomenon; 3) performing CRISPR for single copy insertions; 4) Demonstrating that different glycolytic proteins form condensates; 5) demonstrating the GFP-tagged versions of the protein are capable of rescuing the loss-of-function alleles and 6) Now adding new data demonstrating the observed localization specifically depend on the presence of other glycolytic proteins. This last result supports that GFP tag is not driving the assembly process of glycolytic condensate and that the glycolytic condensate formation requires the presence of specific molecules in the pathway. I add that we routinely use fluorophore markers to over a dozen distinct proteins that label subcellular compartments, and we have never observed the dynamic relocalization reported here, with the exception of other glycolytic proteins that interact with PFK, suggesting this is a property specific to glycolytic proteins, and, based on the genetic studies, dependent on the glycolytic reaction. We add and discuss these findings in Figures 7G, 7H, S7B, and S7C; lines 422-441, 964-989.
  
  -For the Cry2-section, the complementation of the pfk-1 mutant supports functionality of the synaptic clustering phenotype. Are there other features of function that can be evaluated or could you look at how Cry-2 vs wt worms recover from different durations of stress or frequencies. Could you see if the Cry-2-fusion will rescue function to a partial-loss-of-function allele or a tetramerization deficient allele? A detailed analysis of the effects of constitutive presence of PFK-1-Cry2 clusters would be necessary to bolster claims that this is fully functional construct. Can enzyme activity be somehow monitored?
  
  We did not observe any difference between wild-type worms and CRY2-expressing worms with regards to their development, survival, locomotive behavior or synaptic phenotype. While we can not discard the possibility that this is not a full rescue, with available tools, we can not distinguish the recue with PFK-1-Cry2 from that of just PFK-1.
  
  -The analysis of the sphericity of clusters (4A) is limited due to the diffraction limit of light which limits an analysis of a compartment of this size. While this is a limitation of the live organism, this should be more clearly acknowledged.
  
  We have included in the Methods section our criteria for quantifying condensates and avoiding diffraction limit artifacts. Briefly, “Considering the resolution limit of a spinning disc confocal (approximately 300nm), any structure with a diameter less than 500nm and an area smaller than 0.2 µm2 was excluded from the analyses”. To better clarify this point, we also now add a description of the criteria used in the main text (lines 242-243).
  
  In addition, we observed that PFK-1.1 condensates are not perfect spheres, but constrained spheroids (which can not be explained by diffraction-limited point spread functions). We can explain the observed spheroid shapes based on liquid-like properties of the condensates, and the constrains of the diameter of the neurite. To better highlight this finding, we have now moved Figure S4E into the main figure (Figure 4B’).
  
  -Fusion experiments (4C) do not fully exclude that clusters overlap instead of merging. It would be beneficial to show the foci for several subsequent frames. One would expect that upon fusion, the condensate size would increase, but video 3 suggests the opposite. It would be useful to quantify condensate size before and after fusion for several separate fusion events. -an alternative possible experiment would be the tagging of PFK-1 with a photoconvertible fluorophore (e.g. Dendra2) and subsequent analysis of fusion events
  
  To better show the fusion events in Figure 4C, we now include all xy, yz, and zx plane views of before and after fusion events of Figure 4C (Figure S5B). We also added a quantification of four independent fusion events in which we compare the sum of the areas of the two puncta before fusion and the size of the area of the single punctum after fusion (Figure S5C). These data support that we are observing fusions events.
  
  -4D). It is unclear if foci are indeed undergoing fission or if two clusters next to each other are moving apart.
  
  For Figure 4D, in all the frames we had recorded, a single structure maintains a continuous signal until fission occurs and splits into two structures. To better present this event, we now include an unabridged version of figure of 4D in the supplement that shows all the frames captured (Figure S5D).
  
  -The analysis of side-by-side growth and dissolution kinetics are interesting and a novel view into the non-equilibrium aspects of phase separation in cells.
  
  -Purification of PFK-1 and in vitro reconstitution of condensates would be supportive of liquid-like characteristics although I don't think it is necessary however it would add a lot to the relevance to show enzyme activity is different +/- condensate state but I am not sure if an easy enzymatic assay exists in vitro.
  
  We agree. But the significance of this particular paper, specifically in the context of the in vitro enzymatic work on glycolytic proteins, is to examine the dynamic in vivo localization and the biophysical characteristics of the condensates. To better underscore this in the context of the field, we add discussion of a recent in vitro manuscript demonstrating that liquid droplet formation of glycolytic proteins affect their enzymatic activity (Ura et al., 2020) (lines 444-464; 484-492). While we see the value of future studies reconstituting the glycolytic particles, we believe that is beyond the scope of this particular in vivo study.
  
  **Minor comments:**
  
  -Stress granules in other organisms (yeast paper) have different composition depending on stress type. To make the claim that the PFK-1 compartments are independent of SGs one would ideally test multiple different SG markers.
  
  We selected the stress granule protein TIAR-1 because it is one of the most studied stress granule markers in C. elegans and it is reportedly one of the core proteins and universal components of stress granules irrespective of a stress type (Buchan et al., 2011; Gilks et al., 2004; Huelgas-Morales et al., 2016; Kedersha et al., 1999). Although we did not include images in the manuscript, we had tested a total of three stress granule markers: TIAR-1, TDP-43, and G3BP1 with similar results. We now added that as data not shown (lines 193-194).
  
  -it should be stated in the main text that the microfluidic-hydrogel device was fabricated following previously published protocols
  
  We have added the reference in the main text (line 170) to supplement what we had written in the Methods section: “A reusable microfluidic PDMS device was fabricated to deliver gases through a channel adjacent to immobilized animals, following protocols as previously described (Lagoy and Albrecht, 2015)”.
  
  -Figure 4b: Y-axis should be changed from probability to fraction of occurrence
  
  We have corrected this in both the figure and the figure legends (Figure 4B).
  
  -The discussion should be less speculative concerning any effects seen in PFK1-Cry2 expressing C. elegans
  
  We have modified the discussion as suggested.
  
  -it is perplexing that a protein known to tetramerize with no disordered or RNA-binding domains forms condensates like this. Is there anything known from other systems of additional interacting proteins that may have features that promote liquidity and serve to fluidize these assemblies?
  
  Condensates can form via multivalent interactions, which include, but is not exclusive, to disordered or RNA-binding domains. Because glycolytic proteins have dihedral symmetries that can facilitate multivalent interactions, we believe these structural properties, in combination with regulated conformational changes, promote multivalent interactions leading to their condensation. We had a statement in the discussion (lines 494-519) now add this more clearly in the results (lines 395-398).
  
  Reviewer #2 (Significance (Required)):
  
  Stimulus-induced phase separation has been observed for dozens of metabolic enzymes from various different pathways (reviewed in Prouteau, 2018). Several studies have published the formation of condensates through PFK-1 in diverse organisms (C. elegans, Yeast, human cancer cells) in response to hypoxia or in some cancer lines also without hypoxia (Jin, 2017, Jang, 2016, Kohnhorst 2017, etc.). A yeast study showed that PFK-1 condensates contain various other glycolytic enzymes and that condensate formation enhances glycolytic rates (Jin, 2017).
  
  This study gives the advance of analyzing the dynamics of PFK-1 condensate formation in vivo in the context of a live animal using a microfluidic-hydrogel device and showing that PFK-1 relocalizes to reversible condensates within minutes of hypoxia. If further appropriate experiments (as mentioned above) are performed, this study would strongly suggest that the underlying process of PFK-1 condensate formation is liquid-liquid phase separation. Ideally, if at all feasible, it would be strengthened if there was some insight into the functional consequences of the condensed assemblies formed in hypoxia. These findings may be interesting to researchers working on glycolysis and metabolism in different cells but particularly in neurons.
  
  Field of expertise
  
  -Phase separation, microscopy, in vitro reconstitution
  
  -no experience with C. elegans biology and do not have a practical handle on ease or difficulties of genetic manipulation of C. elegans or metabolic assays for PFK-1
  
  Reviewer #3 (Evidence, reproducibility and clarity (Required)):
  
  **Summary:**
  
  In this manuscript, the authors focus on the subcellular localization of the key glycolytic enzyme PFK-1.1 in C. elegans, initially in whole animals through GFP tagging of the endogenous locus and subsequently in single cells/tissues using a clever genome editing strategy that permitted tissue-specific expression of GFP-tagged PFK-1.1 from its endogenous locus. They observe that PFK-1.1 localization differs from cell-type to cell-type and can be dynamically reorganized in response to exogenous cues. Focusing on hypoxia, they observe that PFK-1.1 forms foci near synapses in neurons under this stress condition. These foci are not stress granules and they are dissolved upon re-oxygenation. These condensates have properties of liquid droplets and can mature (harden) over time. PFK-1.1 fused to the CRY domain can trigger condensate formation under normoxic conditions, which can co-recruit WT PFK-1.1 as well as aldolase.
  
  **Major comments:**
  
  The conclusions are convincing but the impact could be increased if the authors were able to demonstrate the physiological role that the observed phase separation plays in this stress response. Would it be possible to assess glycolytic flux under hypoxia vs normoxia?
  
  It is currently not possible to assess glycolytic flux in vivo in our system, as we lack metabolic sensors (an active area of work we are trying to address, but will take several years to perform correctly). We have added discussion of new in vitro studies examining the consequences of metabolic flux due to glycolytic compartmentation into liquid droplets (Ura et al., 2020), and the significance of those findings in the context of our in vivo studies (lines 444-464; 484-492).
  
  The authors should comment on viability during the hypoxia time course.
  
  C. elegans can survive anoxic condition for a day (Powell-Coffman, 2010). Our hypoxic conditions last minutes, and we can rescue live C. elegans upon completion of the assays. We now include a description of this in the Methods (lines 1216-1218).
  
  The co-clustering of ALDO-1 and PFK-1.1::mCh::CRY2 in Figure 7 should be properly quantified/statistically analyzed
  
  We quantified the fraction of animals that displays ALDO-1 clustering in PFK-1.1::mCh::CRY2 co-expressing animals, as suggested (Figure S7C).
  
  A control of mCh::CRY2 + ALDO-1::EGFP is missing from the experiments shown in Figure 7. Is the presence of mCh::CRY2 sufficient to drive ALDO-1::EGFP clustering?
  
  As a control for the CRY2 tag promoting the formation of glycolytic condensates, we had co-expressed mCh::CRY2 with PFK-1.1::EGFP, which is insufficient to cause the formation of the condensate (Figure 7C). We have now added a new data where we show that in pfk-1.1 deletion mutants, ALDO-1 condensate formation is suppressed, which further demonstrates the dependency between PFK-1.1 and ALDO-1 (Figures 7H and S7C).
  
  Does hypoxia trigger co-clustering of ALDO-1 and PFK-1.1?
  
  To answer this question, we examined the dynamic formation of ALDO-1 and PFK-1.1 condensates by co-expressing the two proteins together and observed that hypoxia triggers their co-clustering. We now include this in Figure 7E and Video 8.
  
  The authors speculate that hypoxia acts via diminished energy (altered ATP AMP ratios). Can this be measured? To support this hypothesis, the authors may wish to test if similar phase separation is triggered by mitochondrial poisons.
  
  We currently lack sensors that can reliably measure, in vivo, the subcellular changes in energy or metabolic flux in C. elegans neurons. However, we previously did test mitochondrial mutants and observed that in those mutants we observe glycolytic condensates (Jang et al., 2016), supporting the idea that defects in energy production promotes the formation of glycolytic condensates.
  
  **Minor comments:** Is 21% O2 not hyperoxic for worms?
  
  While C. elegans are known to prefer lower percentage of oxygen than those in air, in the lab animals are reared in normal air. We therefore used 21% oxygen present in air as our normoxic conditions.
  
  Can the authors speculate more on how do these condensates exhibit "memory" (how they're able to cluster in the same place repeatedly)? Is there any role for the cytoskeleton in mediating nucleation and/or condensation of PFK and glycolytic enzymes?
  
  When we were testing the reversibility of PFK-1.1 condensates, we were not expecting the reappearance of PFK-1.1 condensates in the same place repeatedly. Our current speculation is that, because many glycolytic enzymes, such as PFK-1.1, are allosterically regulated by nucleotides, AMP/ATP ratio may play a role on where glycolytic condensates appear. In other words, the specific synaptic areas, where PFK-1.1 condensate repeatedly reappeared, may have different AMP/ATP ratio that may trigger the condensation of the glycolytic proteins in those locationsupon conformational changes in PFK-1. We can’t exclude, currently, the presence of nucleating factors at synapses that facilitate PFK-1 clustering, but we have not yet identified them. We now include a discussion of this (lines 494-519).
  
  Do the authors think that these clusters are effectively G-bodies from yeast?
  
  G-bodies from yeast also shows glycolytic proteins changing from its diffuse localization to punctate localization in response to hypoxia (Jin et al., 2017). G-bodies, like C. elegans glycolytic condensates, are forms of subcellular glycolytic organization within eukaryotic cells. Yet, G-bodies take 24 hours to form, while we observe the glycolytic clusters in C. elegans within minutes of hypoxic conditions. We will need to understand the composition and function of both to determine if these forms of glycolytic subcellular organization represent the same structure. We note that glycolytic clusters have also been observed in some human cancer cell lines (Kohnhorst et al., 2017). Observation of glycolytic compartments in multiple different species and cell types suggest that, although the regulation, composition and formation kinetics of the glycolytic condensates may differ, compartmentalization of glycolytic enzymes may be a conserved feature. We now add a sentence discussing this (line 535-537).
  
  Reviewer #3 (Significance (Required)):
  
  It is much appreciated that this study tackles the cell biology of signaling and metabolism, which is a fascinating but difficult to study aspect of molecular biology. This work conclusively documents the dynamic reorganization of metabolic enzymes in vivo in response to physiological stimuli. Such reorganization had been proposed previously but was controversial and difficult to study in a controlled way. This work not only confirms previous observations but further demonstrates that the dynamic reorganization is mediated by a liquid-liquid phase separation. What is lacking is a demonstration that this phase separation is physiologically important. Such observations would generate interest from a much broader audience; the present audience presently targeting people specifically interested in non-membrane organelles per se. The reviewer has expertise in cell signalling and its regulation by phase separation.
  
  As we explain for Reviewer 1, we focus this study on the biophysical characterization of the condensates, and how that results in compartmentalized enrichment of glycolytic proteins. Examination of the functional significance of the phase separation to the enzymatic reactions in vivo is not currently possible because we lack probes we can use in vivo to measure the metabolites resulting from the reaction. We have now added discussion acknowledging this and framing its significance in the context of what has been published in the field (lines 484-492). For example, a recent manuscript in ChemRxiv demonstrated, in vitro, that the enzymatic activity of glycolytic proteins, hexokinase and glucose-6 phosphate dehydrogenase, promote these enzymes condensing into liquid droplets. The authors further found that the condensation accelerated the glycolytic reactions (Ura et al., 2020). This raises the question whether glycolytic proteins compartmentalize, and form condensates, in vivo, which we address in this manuscript. We capture this point in (lines 444-464) where we explain that, while it has long been hypothesized that glycolytic proteins like PFK-1 could be compartmentalized, this remained controversial due to lack of dynamic in vivo imaging. In our study, and through a systematic examination of endogenous PFK-1.1 via the use of a hybrid microfluidic-hydrogel device, we conclusively determine that PFK-1.1 indeed displays distinct patterns of subcellular localization in specific tissues in vivo.
  
  **REFEREES CROSS-COMMENTING** Globally it seems that all reviewers feel that impact would be increased if the physiological consequence of PFK-1.1 condensates was examined. Other, specific comments seem fair.
  
  PeerReviewed
2. EMBOpress 20 Apr 2020
  
  in Review Commons
  
  Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.
  
  Learn more at Review Commons
  
  Referee #2
  
  Evidence, reproducibility and clarity
  
  This paper reports on the condensation of the glycolytic enzyme PFK-1 in response to hypoxic conditions in neurons of C. elegans. The authors employ a microfluidic-hydrogel device to dynamically monitor the relocalisation of PFK-1 from a mostly diffuse state to clusters in response to hypoxia and show that PFK-1 can undergo multiple rounds of PFK-1 clustering and dissolution. The authors work through the key features of a liquid-like compartment (sphericity, fusion, fast internal rearrangements) and give evidence that PFK-1 may have all three. Finally, the authors tag PFK-1 with the light-inducible multimerization domain Cry2 and find that even without light PFK-1 will constitutively form clusters that sequestrate endogenous PFK-1 as well as other glycolytic proteins. The strength of this work is that it is characterizing what appears to likely be phase separation in the context of a whole animal experiencing a stress that it could encounter in the natural world. A limitation of the work is that it is unclear what the functional implications are of condensates of PFK-1 at the molecular or cell scale.
  
  Major comments:
  
  -All experiments were performed using fluorescently tagged PFK-1 expressed from endogenous promoter or from the native genetic locus which is important for excluding overexpression artifacts. However, there is still risk that the GFP tag is driving the assembly process. In order to exclude tag-specific effects that may cause aggregation of the tetrameric PFK-1, ideally a control would be done in which PFK-1 is visualized through immunofluorescence experiments of WT cells. Alternatively, a short tag (e.g HA, His) as epitope for is an alternative .
  
  -For the Cry2-section, the complementation of the pfk-1 mutant supports functionality of the synaptic clustering phenotype. Are there other features of function that can be evaluated or could you look at how Cry-2 vs wt worms recover from different durations of stress or frequencies. Could you see if the Cry-2-fusion will rescue function to a partial-loss-of-function allele or a tetramerization deficient allele? A detailed analysis of the effects of constitutive presence of PFK-1-Cry2 clusters would be necessary to bolster claims that this is fully functional construct. Can enzyme activity be somehow monitored?
  
  -The analysis of the sphericity of clusters (4A) is limited due to the diffraction limit of light which limits an analysis of a compartment of this size. While this is a limitation of the live organism, this should be more clearly acknowledged.
  
  -Fusion experiments (4C) do not fully exclude that clusters overlap instead of merging. It would be beneficial to show the foci for several subsequent frames. One would expect that upon fusion, the condensate size would increase, but video 3 suggests the opposite. It would be useful to quantify condensate size before and after fusion for several separate fusion events.
  
  -an alternative possible experiment would be the tagging of PFK-1 with a photoconvertible fluorophore (e.g. Dendra2) and subsequent analysis of fusion events
  
  -4D). It is unclear if foci are indeed undergoing fission or if two clusters next to each other are moving apart.
  
  -The analysis of side-by-side growth and dissolution kinetics are interesting and a novel view into the non-equilibrium aspects of phase separation in cells.
  
  -Purification of PFK-1 and in vitro reconstitution of condensates would be supportive of liquid-like characteristics although I don't think it is necessary however it would add a lot to the relevance to show enzyme activity is different +/- condensate state but I am not sure if an easy enzymatic assay exists in vitro.
  
  Minor comments:
  
  -Stress granules in other organisms (yeast paper) have different composition depending on stress type. To make the claim that the FPK-1 compartments are independent of SGs one would ideally test multiple different SG markers.
  
  -it should be stated in the main text that the microfluidic-hydrogel device was fabricated following previously published protocols
  
  -Figure 4b: Y-axis should be changed from probability to fraction of occurrence
  
  -The discussion should be less speculative concerning any effects seen in PFK1-Cry2 expressing C. elegans
  
  -it is perplexing that a protein known to tetramerize with no disordered or RNA-binding domains foms condensates like this. Is there anything known from other systems of additional interacting proteins that may have features that promote liquidity and serve to fluidize these assemblies?
  
  Significance
  
  Stimulus-induced phase separation has been observed for dozens of metabolic enzymes from various different pathways (reviewed in Prouteau, 2018). Several studies have published the formation of condensates through PFK-1 in diverse organisms (C. elegans, Yeast, human cancer cells) in response to hypoxia or in some cancer lines also without hypoxia (Jin, 2017, Jang, 2016, Kohnhorst 2017, etc.). A yeast study showed that PFK-1 condensates contain various other glycolytic enzymes and that condensate formation enhances glycolytic rates (Jin, 2017).
  
  This study gives the advance of analyzing the dynamics of PFK-1 condensate formation in vivo in the context of a live animal using a microfluidic-hydrogel device and showing that PFK-1 relocalizes to reversible condensates within minutes of hypoxia. If further appropriate experiments (as mentioned above) are performed, this study would strongly suggest that the underlying process of PFK-1 condensate formation is liquid-liquid phase separation. Ideally, if at all feasible, it would be strengthened if there was some insight into the functional consequences of the condensed assemblies formed in hypoxia. These findings may be interesting to researchers working on glycolysis and metabolism in different cells but particularly in neurons.
  
  Field of expertise
  
  -Phase separation, microscopy, in vitro reconstitution
  
  -no experience with C. elegans biology and do not have a practical handle on ease or difficulties of genetic manipulation of C. elegans or metabolic assays for PFK-1
  
  PeerReviewed
3. EMBOpress 20 Apr 2020
  
  in Review Commons
  
  Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.
  
  Learn more at Review Commons
  
  Referee #1
  
  Evidence, reproducibility and clarity
  
  Summary
  
  Jang et al., address the important question of spatially localized or compartmentalized metabolic enzymes with a focus on the glycolytic enzyme PFK1. Using a good strategy of inserting a fluorescent tag at the endogenous PFK1 locus with tissue-specific inducible expression in C. elegans, combined with strong quantitative longitudinal imaging and innovative bioengineered microfluidic-hydrogels to control oxygen availability as well as optogenetic approaches, they show PFK1 condensates, which are not stress granules and not seen in normoxia, assemble with hypoxia. PFK1 condensates are dynamic, reversible, localized at the synapse in neurons, and recruit aldolase, another glycolytic enzyme. Although glycolytic proteins were previously shown to compartmentalize near the plasma membrane, and PFK1 was previously shown to assemble into filaments in vitro and be punctate at the plasma membrane in mammalian cells, evidence for cellular localized PFK1 condensates in animals is highly significant. The work includes strong biophysical characterization of PFK1 phase-separated condensates, but no clear indication of the composition of condensates. More significantly, the findings lack functional significance related to PFK1 activity or glycolytic flux with hypoxia vs normoxia. Despite previous work by this group showing that disrupting subcellular localization of glycolytic enzymes impairs neuronal activity in response with hypoxia, the reader is left with questions on the importance of localized and PFK1 condensates and their make-up .
  
  Major comments:
  
  Key conclusions are convincing, and most experimental approaches, biophysical characterization including thermodynamic principles, and data analysis are exemplary and well described. However, as indicated above, the work is limited to a descriptive analysis of cellular localization of PFK1 condensates and their biophysical properties without insights on functional significance relative to enzyme activity - or at least glycolytic flux or metabolic reprogramming with hypoxia. At best, only correlations can be drawn from hypoxia-induced localized PFK1 condensates and the authors' previous report (Jang et al., 2016) on hypoxia-regulated neuronal activity. Some insight or at least prediction in the discussion on the differences in spatially localized PFK1 in muscle vs neurons with regard to metabolic or energy distinctions should be included.
  
  Despite the strong biophysical analysis of condensates, several important features are not determined. First is at best a rudimentary analysis of the composition of condensates and also how PFK1 is assembled into these structures. For the former, is the core of the condensate predominantly PFK1 with perhaps aldolase only recruited to the periphery or is aldolase an integral component of the structure. Hence, is it a PFK1 condensate or a glycolytic condensate? For the latter question, is there a particular orientation for PFK1 in condensates, i.e a collection of filaments as previously reported, which might provide insight on assembly? Finally, and less critical but also important is the criterion for spherical, which is not well defined, and at least some idea or speculation on determinants for a spherical morphology - compared with filaments that have been reported for other non-glycolytic metabolic enzymes.
  
  Significance
  
  The work is an important advance in our understanding on the self-assembly of metabolic enzymes by showing hypoxia-induced PFK1 condensates in vivo, their spatially-restricted subcellular localization in muscle cells and neurons, and their biophysical properties, the latter being distinct from those of stress granules. Taken together, these findings are more extensive than many previous reports on the assembly of metabolic enzymes into filaments or condensates, but fall short for new insights on functional significance.
  
  Expertise is published on topic
  
  PeerReviewed
Visit annotations in context

Tags

PeerReviewed

Annotators

EMBOpress

URL

biorxiv.org/content/10.1101/636449v1
osf.io osf.io

ten-simple-rules-dockerfiles.pdf

1
1. davclark 17 Apr 2020
  
  in Public
  
  You should regularly re-build the image using the--no-cacheoption
  
  And perhaps make sure to tag your good / working container before you do!
Visit annotations in context

Annotators

davclark

URL

osf.io/fsd7t
daryl.wakatara.com daryl.wakatara.com

Why I Switched Back to Taskpaper

1
1. Eejain 16 Apr 2020
  
  in Public
  
  Perhaps it’s just the ability to see everything at once and then filter it down to what I need ot focus on, but also, it’s forcing me to plan and move things into realistic periods when I can get them done.
  
  that's a good point, relates to the bigger idea of project-based (in one file) or tag-based (in different files) todo management
Visit annotations in context

Annotators

Eejain

URL

daryl.wakatara.com/why-i-switched-back-to-taskpaper/
medium.com medium.com

Local e-commerce redesign

1
1. kathryn.mcclure 13 Apr 2020
  
  in Public
  
  we
  
  Instead of just saying we, say my team. This gives you the opportunity to tag your partner(s) and give an over view of the project.
  
  I recommend beginning case studies with a description of the project, your roles, and the timeline.
  
  Project: E-commerce (Insert DEVICE here) Website Concept Roles: UX/UI Designer Timeline: 2 Week Sprint
Visit annotations in context

Annotators

kathryn.mcclure

URL

medium.com/@vladatkach2/dcae7832b022
www.makeuseof.com www.makeuseof.com

New PC? 15 Must-Have Windows Applications You Should Install First

1
1. ch3st3rnguy3n 13 Apr 2020
  
  in Public
  
  Whether you’ve just purchased a new PC or reinstalled Windows, the first task you’ll likely do is installing apps.
  
  True that
Visit annotations in context

Annotators

ch3st3rnguy3n

URL

makeuseof.com/tag/getting-a-new-pc-12-must-have-applications-to-install-first/
theconversation.com theconversation.com

Pourquoi le temps semble filer plus vite avec l’âge

1
1. luizauerj 11 Apr 2020
  
  in Public
  
  Plusieurs théories tentent d’expliquer cette sensation d’accélération du temps avec l’âge. L’une d’elle évoque une dégradation progressive de notre horloge biologique, due au ralentissement naturel de notre métabolisme au fil des ans : quand nous vieillissons, notre respiration et nos battements de cœur ralentissent. Chez les enfants, au contraire, le cœur bat plus vite et les poumons s’activent davantage. C’est cette plus grande intensité de l’activité biologique qui leur donne l’illusion d’un temps dilaté.
  
  argument epistémique abductif. Tag: IED_QA 3 L'auteur utilise l'argument de la vitesse de métabolisme (fait connu) comme une possible hypothese pour expliquer la perception différente du temps entre l'âge adulte et l'enfance.
Visit annotations in context

Annotators

luizauerj

URL

theconversation.com/pourquoi-le-temps-semble-filer-plus-vite-avec-lage-64030
www.weforum.org www.weforum.org

8 digital life skills all children need – and a plan for teaching them

1
1. hkumar 10 Apr 2020
  
  in Public
  
  If ocean plastic pollution was one of the major environmental challenges we finally woke up to in 2018, the ebb and flow of public opinion could and should turn to electronic waste in 2019.
  
  I think it is important that this issue gets more publicity. However, we cannot shift the focus to ewaste and loss sight of plastic pollution. I believe we need to effectively tackle one issue at a time. Ocean plastic pollution has gotten more publicity since 2018, but nowhere near enough. Furthermore, drastic enough measures still have not been taken to address ocean plastic pollution. https://www.e-cycle.com/tag/e-waste-effects-to-the-human-body/
Visit annotations in context

Annotators

hkumar

URL

weforum.org/agenda/2019/01/how-a-circular-approach-can-turn-e-waste-into-a-golden-opportunity/
www.multiplikatoren-projekt.peoplemanagement.uni-muenchen.de www.multiplikatoren-projekt.peoplemanagement.uni-muenchen.de

using-hypothesis-for-readings

1
1. nathanpgibson 06 Apr 2020
  
  in Public
  
  adding tags
  
  When you add tags, you can see all the content for a certain tag on its own web page, like this one: https://hypothes.is/search?q=tag:%22e-Learning%20Guides%22
  
  e-Learning Guides
Visit annotations in context

Tags

e-Learning Guides

Annotators

nathanpgibson

URL

multiplikatoren-projekt.peoplemanagement.uni-muenchen.de/downloads/using-hypothesis-for-readings.pdf
www.makeuseof.com www.makeuseof.com

Raspberry Pi Media Center: How to Install Kodi on Raspbian

1
1. diegodlh 05 Apr 2020
  
  in Public
  
  Configuring Your Raspberry
  
  You also have to enable the gui in boot settings
Visit annotations in context

Annotators

diegodlh

URL

makeuseof.com/tag/install-kodi-raspbian-media-center/
kdemoranville18.edublogs.org kdemoranville18.edublogs.org

Major Assignment #2 Second Draft – Kayla's Blog

1
1. eng101 03 Apr 2020
  
  in Public
  
  but either way there is no price tag on the value of education.
  
  shouldn't this be moved later when you discuss value for the money? Here you are just talking about the rise in cost, right?
Visit annotations in context

Annotators

eng101

URL

kdemoranville18.edublogs.org/2020/03/29/major-assignment-2-second-draft/
www.makeuseof.com www.makeuseof.com

Understanding How Open Source Software Developers Make Money

4
1. Ja1037 02 Apr 2020
  
  in Public
  
  Offering your product in a free and paid version is nothing new and it’s entirely legitimate for OSS products,
  
  I feel as if this has been popular in the whole entire software industry!
2. Ja1037 02 Apr 2020
  
  in Public
  
  By far the most common method of income is to provide a service alongside the OSS product. Pick any OSS project from random and there’s a good chance that they utilize this method in one way or another.
  
  This makes sense
3. hsv1000 01 Apr 2020
  
  in Public
  
  self-taught lone wolves
  
  I wonder why this is
4. hsv1000 01 Apr 2020
  
  in Public
  
  open source and profit are mutually exclusiv
  
  wow I did not know this
Visit annotations in context

Annotators

Ja1037

hsv1000

URL

makeuseof.com/tag/understanding-how-open-source-software-developers-make-money/
Mar 2020
www.iubenda.com www.iubenda.com

Introduction to the Prior Blocking of Cookie Scripts

1
1. TylerRick 30 Mar 2020
  
  in Public
  
  This method has the advantage of being quite fast but with the limit of working only for scripts that don’t require a specific position. Google Tag Manager is therefore not effective for all scripts that display a specific element in a specific position of the page (such as the Facebook Like button).
  
  Google Tag Manager limitations cookie blocking
Visit annotations in context

Tags

limitations

cookie blocking

Google Tag Manager

Annotators

TylerRick

URL

iubenda.com/en/help/3081-prior-blocking-of-cookie-scripts
lelon.io lelon.io

Andy Matuschak’s Evergreen Note Taking System On Roam

1
1. RobHaisfield 30 Mar 2020
  
  in Public
  
  If a note is broad (i.e. contains more than one concept), it makes finding any concept contained in that note harder (since you have to sort through concepts). (This is “solvable” by storing an additional copy of just that one concept somewhere else, but if you do that, why not store just the concept to begin with and refer to it from within the broad note to avoid duplication and extra note maintenance?)
  
  I haven’t thought about reducing [[note maintenance]] yet. I largely just use the [[develop]] tag for that. Interesting.
Visit annotations in context

Annotators

RobHaisfield

URL

lelon.io/productivity/andy-matuschaks-evergreen-note-taking-system-on-roam/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

The Adams Administration

2
1. cushing_j 29 Mar 2020
  
  in Public
  
  Adams administration!
  
  I decided to tag the "Adams administration" as an orgName because it describes all of the politicians involved in Adams's presidency.
2. dtd006 29 Mar 2020
  
  in Public
  
  Vice President
  
  I would tag Vice President as a "concept"
Visit annotations in context

Annotators

dtd006

cushing_j

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/the-adams-administration/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Guns and Ships

1
1. johnbaber8 29 Mar 2020
  
  in Public
  
  British henchmen
  
  I did not tag soldiers or henchman as people because I did not find it necessary to tag individuals who are not important or directly mentioned
Visit annotations in context

Annotators

johnbaber8

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/guns-and-ships/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

The Story of Tonight

2
1. johnbaber8 29 Mar 2020
  
  in Public
  
  tonight
  
  I decided not to tag words talking about a time as an event. This is because there would be too many things considered events then. I believe events are something that is important
2. cushing_j 29 Mar 2020
  
  in Public
  
  the story of tonight
  
  I chose to tag "the story of tonight" as an event because the characters are talking about how what they do tonight will be leave a mark in their legacy and will be so influencial that their children will tell "the story of tonight."
Visit annotations in context

Annotators

cushing_j

johnbaber8

URL

hamiltondiged.blogs.bucknell.edu/2020/03/26/the-story-of-tonight/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Blow Us All Away

1
1. johnbaber8 29 Mar 2020
  
  in Public
  
  father
  
  I did not tag relationship names such as father, mother, brother, or sister as person names
Visit annotations in context

Annotators

johnbaber8

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/blow-us-all-away/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Cabinet Battle #1

2
1. johnbaber8 29 Mar 2020
  
  in Public
  
  President
  
  I did not tag president as a person because i only decided to tag names for people
2. johnbaber8 29 Mar 2020
  
  in Public
  
  South
  
  I decide to tag the South as a place because during this time the North and the South were very specific and seperate
Visit annotations in context

Annotators

johnbaber8

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/cabinet-battle-1/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Yorktown (The World Turned Upside Down)

3
1. johnbaber8 29 Mar 2020
  
  in Public
  
  The bullets out your gun!
  
  I decided to tag all objects that you can physically grasp and touch. Things that are very large i did not consider objects
2. johnbaber8 29 Mar 2020
  
  in Public
  
  Immigrants:
  
  I did not tag immigrants as a person because i decided to only tag names for people
3. chrislanciano 29 Mar 2020
  
  in Public
  
  Immigrants:
  
  I would definitely consider immigrants to be a group of people Orr organization as denoted by a pink tag. Obviously immigrants are a group of people, which would aid me in deciding that this was a group and an organization.
Visit annotations in context

Annotators

chrislanciano

johnbaber8

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/yorktown-the-world-turned-upside-down/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

My Shot

2
1. WillOrzell 29 Mar 2020
  
  in Public
  
  my shot
  
  I think we could potentially tag 'my shot' as a concept as it's less of a physical/tangible thing but a very important concept that is partially behind Hamilton's work ethic and drive.
2. bjb019 29 Mar 2020
  
  in Public
  
  diamond in the rough, a shiny piece of coal
  
  I thought that this was an interesting thing to tag, not only because it is an object but also is a reflection on Hamilton being unique.
Visit annotations in context

Annotators

bjb019

WillOrzell

URL

hamiltondiged.blogs.bucknell.edu/2020/03/26/my-shot/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Washington On Your Side

1
1. WillOrzell 29 Mar 2020
  
  in Public
  
  Washington
  
  I think we should tag Washington as a persName.
Visit annotations in context

Annotators

WillOrzell

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/washington-on-your-side/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

It’s Quiet Uptown

1
1. dtd006 29 Mar 2020
  
  in Public
  
  church
  
  I would tag church as a "placeName"
Visit annotations in context

Annotators

dtd006

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/its-quiet-uptown/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Your Obedient Servant

1
1. chrislanciano 29 Mar 2020
  
  in Public
  
  Obedient Servant
  
  I really like this tag as a person. I feel like the class would be a 50/50 split on whether or not to tag this as a person or even tag it at all, but it is obvious that the Obedient Servant that is being mentioned is Burr himself, as he is promising to abide by the duel that he is proposing to Hamilton.
Visit annotations in context

Annotators

chrislanciano

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/your-obedient-servant/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Alexander Hamilton

1
1. symoneryans 29 Mar 2020
  
  in Public
  
  mainland
  
  I chose to tag this because the whole point of this song is to give background on Hamiltons life and the reference to mainland is Burr talking about the colonies as Hamilton was from a British island
Visit annotations in context

Annotators

symoneryans

URL

hamiltondiged.blogs.bucknell.edu/2020/03/27/alexander-hamilton/
hamiltondiged.blogs.bucknell.edu hamiltondiged.blogs.bucknell.edu

Helpless

1
1. bjb019 29 Mar 2020
  
  in Public
  
  to form a harem
  
  https://www.merriam-webster.com/dictionary/harem Very interesting tag. This is what it means
Visit annotations in context

Annotators

bjb019

URL

hamiltondiged.blogs.bucknell.edu/2020/03/26/helpless/
outline.com outline.com

Copyright © 2020 Deutsches Netzwerk Evidenzbasierte Medizin e.V.

1
1. autschn 29 Mar 2020
  
  in Public
  
  Die Nennung von Fällen ohne Bezugsgrößen ist irreführend. So werden beispielsweise für die einzelnen Länder, Bundesländer oder Regionen lediglich Rohdaten berichtet, ohne Bezug zur Bevölkerungsgröße. Die Angaben könnten sich jeweils auf 100.000 Einwohner beziehen. Auch werden keine zeitlichen Bezugsgrößen genannt. So heißt es etwa "bisher gibt es 10.000 Fälle". Die Nennung von Rohdaten ohne Bezug zu anderen Todesursachen führt zur Überschätzung des Risikos. In Deutschland versterben etwa 2.500 Personen pro Tag. Die Angaben zu den Todesfällen durch Covid-19 sollten daher entweder die täglich oder wöchentlich verstorbenen Personen mit Angabe der Gesamttodesfälle in Deutschland berichten. Auch ein Bezug zu Todesfällen durch andere akute respiratorische Infektionen wäre angemessen.
  
  hä?! Macht RKI doch ... was können die dafür dass die meisten Reporter zu dumm sind das so weiterzugeben?
Visit annotations in context

Annotators

autschn

URL

outline.com/vwCK5s
www.buzzfeednews.com www.buzzfeednews.com

A Woman With Lupus Said Her Health Care Provider Is Stopping Her Chloroquine Prescription And Thanked Her For The “Sacrifice”

1
1. golombb 27 Mar 2020
  
  in Public
  
  "Extensive experience and research show that hydroxychloroquine builds up in the body and continues to work for an average of 40 days even after the last dose is taken. By then, we expect the drug manufacturers to have ramped up production to meet the increased demand. Until then, we are no longer refilling routine prescriptions to ensure we have adequate supply to care for our sickest patients," Gin said.{"adPos":"promo-inline3","adType":"ex","isInfinite":true,"platform":"autodetect","position":3,"renderLookahead":"x0.25","size":[[5,5],[728,90],[300,250],"fluid",[320,50]],"targeting":{"badges":["viral","coronavirus"],"bid":"5376488","brain_tags":["adult-0","crime-30","safe-70","non_profane"],"nbs":0,"nsfw":0,"pos":["promo-inline3"],"sensitive":0,"tag":[],"trending":0,"user":"tanyachen","wid":"212-1","infinite_index":1},"viewability":"high","wid":"212-1","zone1":"bfnews"} Advertisement [tl-ut] { content: ""; } div#BuzzfeedNews_Desktop_inarticle_MPU_Flex_container { position: relative; box-sizing: border-box; margin: 0px auto; display: flex; flex-flow: row nowrap; align-items: initial; max-width: 100%; max-height: none; background-color: rgb(255, 255, 255); border: 1px solid rgba(0, 0, 0, 0.1); width: 100%; height: auto; padding: 0px; font-size: 16px; font-weight: 400; font-style: normal; font-family: Avenir; color: rgb(34, 34, 34); letter-spacing: normal; line-height: 1.5em; text-decoration: none solid rgb(34, 34, 34); text-align: start; border-radius: 3px; background-image: none; -webkit-box-orient: horizontal; min-width: 728px; max-width:1100px;} [id^='BF_WIDGET'] div#BuzzfeedNews_Desktop_inarticle_MPU_Flex_container{ left:-44px; } div#BuzzfeedNews_Desktop_inarticle_MPU_Flex_container_left { position: relative; } div#BuzzfeedNews_Desktop_inarticle_MPU_Flex_container_right { position: relative; padding: 10px 10px 10px 20px; text-align: left; width: 100%; box-sizing: border-box; flex-grow: 1; } div#BuzzfeedNews_Desktop_inarticle_MPU_Flex_textcontent { position: relative; flex-direction: column; height: 100%; display: flex; } div#BuzzfeedNews_Desktop_inarticle_MPU_Flex_container_top { position: relative; flex-grow: 1; } div#BuzzfeedNews_Desktop_inarticle_MPU_Flex_container_bottom { position: relative; } img#BuzzfeedNews_Desktop_inarticle_MPU_Flex_media { display: block; box-sizing: border-box; width: initial; max-width: unset; } a#BuzzfeedNews_Desktop_inarticle_MPU_Flex_sponsor { display: block; font-size: 14px; font-family: "Proxima Nova", Helvetica, Arial, sans-serif; text-transform: none; color: rgb(238, 51, 34); text-decoration: none solid rgb(238, 51, 34); padding: 0px 0px 5px; font-weight: 400; font-style: normal; letter-spacing: normal; line-height: 14px; text-align: left; border-radius: 0px; border-width: 0px; border-color: rgb(238, 51, 34); border-style: none; background-color: rgba(0, 0, 0, 0); background-image: none; -webkit-box-orient: horizontal; } span#BuzzfeedNews_Desktop_inarticle_MPU_Flex_sponsor_name { font-weight: 700; padding: 0px; font-size: 14px; font-style: normal; font-family: "Proxima Nova", Helvetica, Arial, sans-serif; color: rgb(238, 51, 34); letter-spacing: normal; line-height: 0em; text-decoration: none solid rgb(238, 51, 34); text-align: left; border-radius: 0px; border-width: 0px; border-color: rgb(238, 51, 34); border-style: none; background-color: rgba(0, 0, 0, 0); background-image: none; -webkit-box-orient: horizontal; } span#BuzzfeedNews_Desktop_inarticle_MPU_Flex_sponsor:hover { } h3#BuzzfeedNews_Desktop_inarticle_MPU_Flex_heading { font-size: 22px; color: rgb(17, 17, 17); font-weight: 600; font-family: "Proxima Nova", Helvetica, Arial, sans-serif; line-height: 1.18182em; text-align: left; text-decoration: none solid rgb(17, 17, 17); margin: 5px 0px 0px; padding: 0px; font-style: normal; letter-spacing: normal; border-radius: 0px; border-width: 0px; border-color: rgb(17, 17, 17); border-style: none; background-color: rgba(0, 0, 0, 0); background-image: none; -webkit-line-clamp: 2; -webkit-box-orient: vertical; } h3#BuzzfeedNews_Desktop_inarticle_MPU_Flex_heading:hover { text-decoration: none; } p#BuzzfeedNews_Desktop_inarticle_MPU_Flex_caption { color: #222; font-family: 'Pensum','Times New Roman',times,serif; font-size: 16px; font-weight: 400; line-height: 30px; display: -webkit-box; -webkit-line-clamp: 1; -webkit-box-orient: vertical; overflow: hidden;} a#BuzzfeedNews_Desktop_inarticle_MPU_Flex_cta { border: 0px solid rgb(255, 100, 24); width: 100%; text-align: center; text-decoration: none; padding: 5px; font-weight: bold; font-size: 10px; font-family: Helvetica, Arial, sans-serif; color: rgb(255, 255, 255); letter-spacing: normal; line-height: 1; display: none; box-sizing: border-box; background-color: rgb(245, 166, 35); max-width: 200px; } a#BuzzfeedNews_Desktop_inarticle_MPU_Flex_cta:hover { text-decoration: none; } a#BuzzfeedNews_Desktop_inarticle_MPU_Flex_media_link, a#BuzzfeedNews_Desktop_inarticle_MPU_Flex_heading_link, a#BuzzfeedNews_Desktop_inarticle_MPU_Flex_caption_link { font-size: inherit !important; font-weight: inherit !important; font-style: inherit !important; line-height: inherit !important; color: inherit !important; text-decoration: none !important; } Quinlan School of Business Our top-ranked, flexible MBA program can help you take your career to the next level at whatever pace works for you. Learn more! Sponsored by Loyola University Chicago See More "Kaiser Permanente physicians and pharmacists are also working together on an evidence-based approach to identify alternative therapies for patients with lupus," Gin added in a follow-up statement.Dale said she immediately called her doctor and has been scheduled for a phone call next week. { "id": "1241213117872234496", "params": { "conversation": "none" } } { "id": 124409135 } Despite thin evidence for the drug’s effectiveness against coronavirus infections, shortages of chloroquine have erupted since Trump called it a “game changer” at a White House news conference late last week. The drug, a derivative of an antimalarial drug, has been added to the regimen for treating COVID-19 infections in China and South Korea and is being tested in clinical trials in the US.However, experts on drug testing have been skeptical of the evidence for its benefits. A frequently cited French study of 20 patients saw several drop out of the trial to instead go into intensive care.{"adPos":"promo-inline4","adType":"ex","isInfinite":true,"platform":"autodetect","position":4,"renderLookahead":"x0.25","size":[[5,5],[728,90],[300,250],"fluid",[320,50]],"targeting":{"badges":["viral","coronavirus"],"bid":"5376488","brain_tags":["adult-0","crime-30","safe-70","non_profane"],"nbs":0,"nsfw":0,"pos":["promo-inline4"],"sensitive":0,"tag":[],"trending":0,"user":"tanyachen","wid":"213-1","infinite_index":1},"viewability":"high","wid":"213-1","zone1":"bfnews"} Advertisement An Arizona man died on Monday after self-medicating with a related drug, chloroquine phosphate, where chloroquine was also touted at White House news conferences.“For many people with lupus there are no alternatives to these medications,” the Lupus Foundation of America said in a statement on Monday, warning of shortages. “Hydroxychloroquine or chloroquine are the only methods of preventing inflammation and disease activity that can lead to pain, disability, organ damage, and other serious illness.”Dale, who's been calling multiple pharmacies in her local area, said, "I have learned that all area pharmacies are completely out of hydroxychloroquine." { "id": 124409399 } <img src="https://img.buzzfeed.com/buzzfeed-static/static/2020-03/25/15/asset/705a16f1d755/sub-buzz-3668-1585151028-3.jpg" alt="" class="xs-block"/> healthy.kaiserpermanente.org { "id": 124409135 } "In their mission statement, Kaiser says that they aim 'to provide high-quality, affordable health care services and to improve the health of our members and the communities we serve,'" Dale said. "How is denying medication for a chronically ill, immunocompromised patient during a pandemic improving my health?""I want Kaiser to follow their own mission statement and reverse the decision to withhold my medication." UPDATE March 25, 2020, at 12:48 p.m. This story has been updated to include a follow-up statement from Kaiser Permanente. More on this Chloroquine Is Being Touted As A Miracle Drug For Coronavirus, But There Are Reasons To Be Wary Dan Vergano · March 23, 2020 A Man Died After Self-Medicating With A Form Of A Drug That Trump Promoted As A Potential Treatment For The Coronavirus Brianna Sacks · March 23, 2020 Trump Said He Wants To Give Coronavirus Patients An Experimental Drug Called Chloroquine Dan Vergano · March 19, 2020 Tanya Chen is a social news reporter for BuzzFeed and is based in Chicago. Contact Tanya Chen at tanya.chen@buzzfeed.com. Got a confidential tip? Submit it here. Dan Vergano is a science reporter for BuzzFeed News and is based in Washington, DC. Contact Dan Vergano at dan.vergano@buzzfeed.com. { "ads": {"density": 1, "placements": [], "skin": "news", "units": [{"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar1-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar1-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1301}, "viewability": "high", "wid": 1301, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar2-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar2-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1302}, "viewability": "high", "wid": 1302, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar3-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar3-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1303}, "viewability": "high", "wid": 1303, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar4-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar4-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1304}, "viewability": "high", "wid": 1304, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar5-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar5-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1305}, "viewability": "high", "wid": 1305, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar6-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar6-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1306}, "viewability": "high", "wid": 1306, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar7-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar7-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1307}, "viewability": "high", "wid": 1307, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar8-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar8-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1308}, "viewability": "high", "wid": 1308, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar9-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar9-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1309}, "viewability": "high", "wid": 1309, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}, {"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar10-bp", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [5, 5], [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar10-bp"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 1310}, "viewability": "high", "wid": 1310, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}], "unitsRepeated": [{"companions": [], "is_enabled": true, "programmatic_wireframes": true, "slot": {"adPos": "sidebar-bp-infinite", "adType": "post", "isInfinite": true, "platform": "desktop", "position": null, "renderLookahead": "x0.25", "size": [[300, 250], "fluid", [300, 600]], "targeting": {"badges": ["viral", "coronavirus"], "bid": "5376488", "brain_tags": ["adult-0", "crime-30", "safe-70", "non_profane"], "nbs": 0, "nsfw": 0, "pos": ["sidebar-bp-infinite"], "sensitive": 0, "tag": [], "trending": 0, "user": "tanyachen", "wid": 13000}, "viewability": "high", "wid": 13000, "zone1": "bfnews"}, "template_path": "/adlib/skins/news/sidebar.html", "wireframe_height": 600, "wireframe_width": 300, "with_companions": false}]} } {"adPos":"sidebar3-bp","adType":"post","isInfinite":true,"platform":"desktop","position":null,"renderLookahead":"x0.25","size":[[300,250],"fluid",[5,5],[300,600]],"targeting":{"badges":["viral","coronavirus"],"bid":"5376488","brain_tags":["adult-0","crime-30","safe-70","non_profane"],"nbs":0,"nsfw":0,"pos":["sidebar3-bp"],"sensitive":0,"tag":[],"trending":0,"user":"tanyachen","wid":"1303-1","sidebarPage":3,"infinite_index":1},"viewability":"high","wid":"1303-1","zone1":"bfnews","observeDimensions":true} Advertisement
  
  This may be true but I am still confused about how people know this helps fight COVID-19
Visit annotations in context

Annotators

golombb

URL

buzzfeednews.com/article/tanyachen/kaiser-permanente-lupus-chloroquine
addvaluerenovations.co.nz addvaluerenovations.co.nz

Affordable Bathroom Renovations Auckland, NZ | Add Value

1
1. leroy30 26 Mar 2020
  
  in Public
  
  Change title tag:
  
  Affordable Bathroom Renovations, Auckland - Add Value Renovation
Visit annotations in context

Annotators

leroy30

URL

addvaluerenovations.co.nz/bathroom-renovations
europepmc.org europepmc.org

Precision nomenclature for the new genomics

1
1. GigaScience 25 Mar 2020
  
  in Gigascience Annotations
  
  hypothes.is (use the hashtag/tag #chromosomenomenclature)
  
  Please add comments directly on the key parts of the commentary you would like to raise any issues with.
  
  chromosomenomenclature
Visit annotations in context

Tags

chromosomenomenclature

Annotators

GigaScience

URL

europepmc.org/article/MED/31437278
fortelabs.com fortelabs.com

Tagging is Broken

4
1. RobHaisfield 25 Mar 2020
  
  in Public
  
  But everything’s changed. Notes are stored locally so there is no lag, search is blazing fast, and the search bar provides recommendations and suggestions that are far more accurate and helpful than tags
  
  I would like less lag time in adjusting and reformulating my searches as I go. The lag time at this point has more to do with how slow Roam is to render things than how slow the process could be, since the sidebar should allow you to pull up many things, explore new threads of thought, and change your question as you go. Thing is, text search with filtration is still powerful as a tag based search if you’re looking at purely functionally, but tagging also gears you towards asking, “when do I want this to resurface?” So you end up coming to more relevant stuff than simply what is written in the base text. This means you're more likely to resurface notes when you take a tagging approach because it gears you to behave differently towards your notes.
  
  That being said, I don't see this as one or the other. Sometimes I think it's better to just search for a note with text. Only problem is that primarily works for searches where you can clearly formulate your question/goal, you know what you're looking for, and you know where to find it. That essentially turns search into something that requires memory, whereas tag based searches allow for exploratory browsing.
2. RobHaisfield 25 Mar 2020
  
  in Public
  
  So which behaviors are desired and undesired when it comes to organization? In any organizational system, the constant temptation is to overorganize, i.e. to create too many categories, too many subdivisions that are too specific. As the number of tags grows arithmetically, their complexity grows geometrically, for multiple reasons, both technological and cognitive (see memory fatigue above). This phenomenon is all the more problematic with unlimited digital information that never runs into physical constraints.
  
  This could be true, it’s a good point. I come back to Conor’s framework of reduce, filter, and map though. If the user is doing behaviors along the way that reduce the amount of information they’ll see when they look in the future, filter to specify what you want, and then map it out to new contexts, then this bug becomes a feature. Progressive summarization helps in that process, and filter specific tags also helps, as well as straight up deleting things or applying an archive tag that you can always filter out.
  
  This problem also gets reduced when you have aliases and hierarchy abilities along with the tags, because then you’re able to better express what you mean in fewer words/tags. Problem is that Roam doesn't have aliases yet to let the user reduce clutter in their pages and with the different ways that users express the same thing. Tags with bidirectional and unidirectional aliases allow us to express meaning, rather than remember specific words.
3. RobHaisfield 25 Mar 2020
  
  in Public
  
  Thus the most problematic behavior is implicitly encouraged and enabled. Grrrrrrr. It could be a misspelling. It could be a slip of the finger. It could be a different capitalization, punctuation, or tense, whatever. No warning or indication is given, and a divergent tag is created, for you to hopefully notice and fix later, hopefully before you rely on it.
  
  Aliases and hierarchy! Would also be helpful if the app suggested aliases to you based on frequent usages together or similar letter combinations.
4. RobHaisfield 25 Mar 2020
  
  in Public
  
  As good as your brain is at recognizing patterns, it is terrible at storing and recalling multiple patterns precisely, since the patterns of neuronal activation interfere with each other. Yet this is exactly what you’re doing with tags. What a terrifically unnecessary expenditure of mental resources
  
  See my comment about pattern recognition above. The functionality of tagging can be set up so you have frequent/spaced repetition. Also sort of a feature, not a bug. If something doesn’t get consistently reinforced, then it should sink away. You just need to balance search strategies. Don’t always go in with one predefined query with filters. Let yourself follow train of thoughts, and if it resurfaced, then tag it up more then or build on it. If not, it probably wasn’t too valuable anyway.
  
  You can also program your attention so some tags never go away by making index pages. I’d say this all just depends on your behavior with notes more than anything, I don’t think about it so much as tags vs. folders.
  
  However, I would totally say that you need to behave a certain way with your notes or it does become unnecessary expenditure. If you aren’t searching your notes and conversing with them, you won’t see the benefits.
Visit annotations in context

Annotators

RobHaisfield

URL

fortelabs.com/blog/tagging-is-broken/
hypothes.is hypothes.is

Hypothesis

1
1. lmichan 23 Mar 2020
  
  in Public
  
  unidad_COVID2019 query
Visit annotations in context

Tags

unidad_COVID2019

query

Annotators

lmichan

URL

hypothes.is/search
protonmail.com protonmail.com

How to avoid receiving spam - ProtonMail Support

1
1. earthworm 23 Mar 2020
  
  in Public
  
  Using + to trace Spam Some site collect your information to sell to other people. To detect this, you can use a custom tag in your email address for every site you sign up for. For instance, if you were joining the Washington Post email list, you could sign up with your_username+washingtonpost@protonmail.com. This will deliver the email into your account, while allowing you to identify where you inputted the email address. If you receive email to this address from anyone other than the Washington Post you will know they either sold your data or experienced a data breach. To learn more about using + in your email address, please see: Addresses and Aliases.
  
  spam
Visit annotations in context

Tags

spam

Annotators

earthworm

URL

protonmail.com/support/knowledge-base/avoid-spam/
shoup.net shoup.net

std6.pdf

1
1. blipp 17 Mar 2020
  
  in Public
  
  DEM.Encrypt(K, L, M)
  
  Here, in contrast to the DEM used in the Tag-KEM paper, we have a label (additional data).
  
  hpke
Visit annotations in context

Tags

hpke

Annotators

blipp

URL

shoup.net/iso/std6.pdf
eprint.iacr.org eprint.iacr.org

027.pdf

2
1. blipp 17 Mar 2020
  
  in Public
  
  Theorem 3.1[Tag-KEM/DEM Composition Theorem] If the Tag-KEM is CCA secure and theDEM is one-time secure then the Hybrid PKE scheme in Section3is CCA secure. In particular,²pke<2²tkem+²dem.
  
  Could we reuse this theorem? Or at least replicate this proof in CryptoVerif?
  
  hpke
2. blipp 17 Mar 2020
  
  in Public
  
  Notethat, in the above syntactic definition,τis not included inψand explicitly given toTKEM.Dec
  
  If we change DHKEM to use a context when deriving zz, does this make DHKEM a Tag-KEM?
  
  hpke
Visit annotations in context

Tags

hpke

Annotators

blipp

URL

eprint.iacr.org/2005/027.pdf
docs.aws.amazon.com docs.aws.amazon.com

Tagging Your Amazon EC2 Resources - Amazon Elastic Compute Cloud

1
1. sportebois 17 Mar 2020
  
  in Public
  
  You can tag public or shared resources, but the tags you assign are available only to your AWS account and not to the other accounts sharing the resource.
  
  tags on AMI cannot be used for search across accounts
Visit annotations in context

Annotators

sportebois

URL

docs.aws.amazon.com/AWSEC2/latest/UserGuide/Using_Tags.html
Local file Local file

2020 TOY packet.pdf

1
1. embartra 13 Mar 2020
  
  in Public
  
  Lydia Sigourney
  
  You could, if you wanted, tag all of the Connecticut authors or locations in a document.
  
  connecticut
Tags

connecticut

Annotators

embartra
alexberenfeld.com alexberenfeld.com

Project 2

1
1. sc_geier 11 Mar 2020
  
  in Public
  
  sometimes called
  
  The HTML5 audio broken for several reasons: you can't use the same name for a class and ID ('audio'), and you can't use an already reserved tag name (<audio> and 'audio'). Also, it looks like the path to the audio file is wrong, or the file is missing.
Visit annotations in context

Annotators

sc_geier

URL

alexberenfeld.com/project2/ch3.html
my.onetrust.com my.onetrust.com

OneTrust Cookie Auto-Blocking™

1
1. TylerRick 11 Mar 2020
  
  in Public
  
  Cookies may not be detected by scanner if the related tag is triggered by actions such as form submission, scroll depth, timing delay, etc. These tags will need to be controlled by manual methods.
  
  With all these caveats listed, it makes me wonder for which tags auto-blocking does work. Only script tags inside of head?
  
  They are a bit vague in their "how it works" description...
  
  automation cookie blocking caveat
Visit annotations in context

Tags

caveat

automation

cookie blocking

Annotators

TylerRick

URL

my.onetrust.com/s/article/UUID-c5122557-2070-65cb-2612-f2752c0cc4aa
www.biorxiv.org www.biorxiv.org

The interaction between Sec22b-Stx complexes and E-Syts promotes neurite growth and ramification

1
1. EMBOpress 11 Mar 2020
  
  in Review Commons
  
  Note: This rebuttal was posted by the corresponding author to Review Commons. Content has not been altered except for formatting.
  
  Learn more at Review Commons
  
  Reply to the reviewers
  
  __Reviewer #1 (Evidence, reproducibility and clarity (Required)):
  
  In this article, the authors characterize a complex formed by sec22b-stx1-Esyt2. They investigate how such interactions are involved in the modulation of dynamics of the plasma membrane in the context of neuritogenesis. They conclude that the contact sites between the ER and the plasma membrane, mediated by the afore-mentioned complex, contribute the expansion of the plasma membrane.
  
  **Major comments:**
  
  Overall, the article clearly shows that in mammalian cells there is an interaction between sec22b-stx1-Esyt2 which appears to be important for filopodia formation and possibly neuritogenesis in neurons. However, performing additional experiments to better clarify some links and assumptions made by authors could strengthen the article.
  
  The manuscript relies on work performed either on cell lines (HeLA, PC12) or primary neuronal cultures. Although it is clear the value of the findings obtained using cell lines, they should be seen as a complementary rather than an exclusive approach. This is particularly important as the authors often make claim about neuron-related cellular biology.
  
  For instance, the biochemistry-based findings on the interaction and characterization of the protein complex (Figure 1) are all derived from experiments perfomed in Hela or PC12. As the authors have the capacity to culture and manipulate primary neuronal cultures, such findings should be validates in neuronal cells. The authors could also consider performing biochemical experiments (i.e. co-ip) of the endogenous proteins in neuronal cultures or brain tissue.*
  
  ->Endogenous Co-IP has been tried in E18 brain tissue. One experiment using brain tissue demonstrated co-immunoprecipitation of endogenous Sec22b and E-Syt2. Unfortunately, repetitions of this experiment failed due to high background in negative control (naïve Rabbit IgG). We agree with the reviewer that this data is worth trying again. We will carry out this co-immunoprecipitation experiment from cultured neurons to answer the reviewer’s request.
  
  The authors do show some evidence regarding the complex in neuronal cells using PLA (proximity ligation assay, figure 2) or super resolution microscopy, however, these findings should be corroborated by stronger findings targeting interaction and not based on simple proximity.
  
  ->We agree with this reviewer that PLA is limited in demonstrating the occurrence of a protein complex. We would like to stress that we have used PLA complementarily to immunoprecipitation and that we already have shown STED super-resolution data (Figure 3). In order to strengthen the STED data, we will include more details in the figure, as a supplementary movie and a supplementary spreadsheet with the quantification of the distance between the E-Syt2/Sec22b clusters to the plasma membrane stained using WGA. The STED data demonstrate that 50% of the clusters are closer than 33.6nm to the plasma membrane, a distance in the range of ER-PM contact sites.
  
  A similar critique regards the experiments using RNA-interference of Figure 4. Performing loss-of-function experiments in neuronal cultures would strengthen and complement the results obtained via over-expression approaches shown in Figure 5.
  
  ->The loss-of-function experiments in neuronal cultures using siRNA were attempted unsuccessfully. The three E-Syts have largely different cDNA sequences thus three distinct siRNAs must be transfected in order to silence all three simultaneously. This is quite challenging in neuronal cultures and we were never able to get strong silencing of the three E-Syts. In the following points, we plan to carry out further experiments using expression of a fragment of Sec22b (Longin domain). We are confident that this is a better approach to demonstrate the importance of Sec22b/E-Syt interaction.
  
  *Given that the authors have already in place all the necessary technology for the suggested biochemical and morphological-related experiments, these could be carried out swiftly within 3-4 months.
  
  **Minor comments:**
  
  The manuscript is really technical and at times tough to follow; it could benefit from key sentences to better guide the reader, particularly if not coming from the specialist field, in the appreciation of the experiments and results.
  
  Authors should submit the manuscript to a severe round of proofreading. There are several inconsistencies and sometimes what looks like internal comments: i.e. in the methods "STED Missing" or the fact that "LTP" is used everywhere but not defined and considering that the targeted audience is most likely neuroscience-based could easily lead to confusion.
  
  *
  
  ->We fully agree with this reviewer and apologize for leaving behind such errors. We will carefully proofread the revised ms.
  
  *The experiments appear to have been repeated a sufficient number of times and the statistics seem adequate. It would be advisable to show in dot-plots the findings rather than in bar graphs all findings and not just the morphometrics-relative ones.
  
  *
  
  ->We will modify the figures according to this reviewer’s suggestion.
  
  Reviewer #1 (Significance (Required)):
  
  *This work closely follows the excellent previous work from the Galli laboratory. As such, it is mostly incremental from a technical perspective and does not present particularly novel findings. An interesting aspect would be in addressing directly the influence of the described interactions in the lipid transfer between ER and the plasma membrane but in that sense the manuscript falls short. Although it is to be appreciated the functional readouts in terms of neuritogenesis, in the present state the manuscript features findings suitable for a very specific audience.
  
  I believe that the appropriate audience for the present manuscript lies within the neuroscientific community interested in development, specifically neuritogenesis, and/or membrane dynamics. Additionally, it might be interesting also for researchers outside of the neuroscience community and interested in the dynamics between ER and plasma membrane.
  
  *
  
  ->We are happy to read the comments of this reviewer. Nevertheless, we would like to stress the importance of deciphering precise molecular mechanisms in any biological process. Here, we are the first to demonstrate an interaction between lipid-transfer proteins E-Syts and ER v-SNARE Sec22b. As an example, the molecular mechanism connecting synaptic SNAREs and synaptotagmin has been the topic of more than 500 publications since seminal articles in the early 1990’s. We think that the first article linking E-Syts to SNAREs cannot be considered as a mere increment from our previous work.
  
  The activity of E-Syts to transfer lipids in vitro has been well established __(1–3) In addition, recent work by the De Camilli lab using Origami showed that reducing the distance between liposomes enhanced the lipid transfer mediated by E-Syt2 (3). Therefore, we did not carry out experiments such as combining SNAREs and E-Syt2 in artificial membranes in vitro because we considered that there would not be much more to demonstrate than what has already been done. Furthermore, we considered the experiments in cells, particularly neurons, much more critical at this point. Demonstrating transfer of glycerophospholipid between ER and PM in cells cannot be performed like other lipids’ transfer at other membrane interfaces for the following reasons: phospholipids are very abundant (4) and they are not modified upon transfer (1)__, there are no specific dyes to detect glycerophospholipids (unlike phosphoinositides), and ER and PM are too close to distinguish if a glycerophospholipid is in one or the other membrane. Such a challenging experiment would require the ability to setup a specific biochemical assay circumventing these constraints. We think that this is out of the scope of the present study focused on the role of E-Syt/Sec22b complex.
  
  Nevertheless, in order to get further insights on this question, we will express WT and mutant E-Syt2, purify the PM using the protocol of Figure 4 in Saheki et al __(1)__, followed by lipidomics analysis. We hope that this approach further supports our idea that E-Syts mediate an important lipid transfer mechanism towards the PM.
  
  * Keywords regarding my expertise: Molecular and Cellular Neuroscience, Morphometrics, Dendrite, Neurons, Dendritogenesis, Biochemistry, Imaging, Microscopy.
  
  __
  
  Reviewer #2 (Evidence, reproducibility and clarity (Required)): *This manuscript identifies and characterises a novel interaction between E-Syts and Sec22b and demonstrates that lipid transfer between the ER and PM contributes to the development of filopodia and neuronal expansion. This interaction with E-Syt2 occurs through the Longin domain of Sec22b Sec22b association. The authors suggest a continuum with further interactions with syntaxin1, that mediates neurite outgrowth. Overall I find this study very interesting and convincing. The experimental analysis is well carried out and the claims are well aligned with their results.
  
  I only have minor issues:
  
  Figure 1. Some of the western blots have several bands and it is difficult to know which band is the relevant one. They should be indicated in the fig panel. Further panel E and F are barely readable and should be redrawn with the appropriate line and font size.*
  
  ->We will make the changes requested by this reviewer in Figure 1.
  
  *
  
  Figure 2: is there a difference between the number of dots in axons and dendrites? Can the author elaborate on this aspect as it is not clear from the image presented.
  
  ->We could not combine PLA with further staining of MAP2 and TAU. Indeed, to perform PLA, neurons are already double labelled to detect the proteins of interest. At the stage of the neurons used in this study, both axons and dendrites are growing. Therefore, we did not invest in distinguishing between axons and dendrites. Because growth cones are known to be the major sites of membrane growth, we instead distinguished dots within neurites and in growth cones. We will make the other changes requested by this reviewer in Figure 2.
  
  Figure 7: statistical analysis should be indicated from the BoNT/A and BoNT/C as BoNT/A represent an appropriate control cleaving SNAP25 but not Syntaxin.
  
  ->We agree with this request and we will add statistical analysis as suggested, using BoNT/A as an additional control.
  
  On top of controlling fusion and neuronal outgrowth, syntaxin has a role in survival and its cleavage leads to neuronal death. Is this pathway mediated by E-Syts interactions?
  
  ->We have stated in the ms: “Since exposure to BoNT/C1 at high concentrations and for long incubation periods causes degeneration of neurons in culture __(5,6)__, various concentrations and incubation times were tested, and a 4-hour treatment of neurons with 1nM BoNTs was chosen to avoid such deleterious effects.” Accordingly, we did not see any degeneration in our experimental conditions.__ __
  
  Reviewer #2 (Significance (Required)): This papers identifies the molecular mechanism of neuronal outgrowth. It is highly significant. ->We are very grateful to this reviewer for pointing out the high significance of our article.
  
  __Reviewer #3 (Evidence, reproducibility and clarity (Required)):
  
  __*1. The evidence for the claim that the Sec22b/Stx1 complex and E-Syts colocalize in native cells (neurons) and bind in heterologous cells is strong (3 independent lines of evidence: co-immunoprecipitation, Proximity Ligation (PLA) and STED super-resolution microscopy) However, the current title of the paper makes a claim beyond this interaction/proximity, based on evidence that is obtained with E-Syt over-expression in wildtype cells. The physiological relevance of the effects remain elusive with over-expression in wildtype cells only.
  
  Furthermore, it is plausible that overexpression of membrane binding/bending C2-domains promotes neurite outgrowth and ramifications by a non-specific effect (as shown for copine C2 domains, PMID:25450385 and indirect evidence for synaptotagmins1,2,7).*
  
  * This issue is especially relevant in the light of the fact that loss of all 3 Extended Synaptotagmins does not affect normal mouse development and viability (PMID: 27399837)
  
  It would be more appropriate to choose a more descriptive title*
  
  *
  
  ->We agree with this reviewer that the original title may be too strong and are now proposing the following, more descriptive title:
  
  Role of the Sec22b/E-Syt complex in neurite growth and ramification
  
  We are fully aware that proteins harbouring C2 domains could potentially promote non-specific effects when overexpressed. However, we do not think this is the case here because none of the morphological effects of E-Syt2 expression in neurons and HeLa cells were reproduced by mutants lacking the SMP or the membrane-anchoring domains. Based on work on Copine __(7)__, a cytosolic protein, E-Syt2 still containing 3xC2 domains but lacking the membrane-anchoring domain should have shown a morphological effect if non-specific binding to phosphoinositides was the mechanism of action. We will discuss this point in the ms.
  
  The evidence for the working model that the morphological effects of E-Syt2 depends on the Sec22b/Stx1 complex is not strong. Although plausible, the positive effect on neurite outgrowth (E-Syt2 overexpression) and the negative effects (inhibition by Stx1 cleaveage, Sec22b-Longin or Sec22b extended linker expression) may in fact be independent
  
  The evidence could be strengthened by PLA measurements in neurons over-expressing Myc-E-Syt2 and Sec22b to assess MSC density. It is predicted that in both conditions, MCS density increases. MCS density by PLA measurements could also be performed in Sec22b-P33 and DLongin overexpressed and BoNT/C1 treated neurons. According to the model, the number of MCS should go down. This is of special interest for BoNT/C1 treatment, as it is important to show that the altered morphology is not purely caused by a pre-state of degeneration that is known to be induced by BoNT/C1. In addition, EM measurements of ER-PM distances might provide an independent line of evidence.*
  
  ->We agree with this reviewer that additional experiments could strengthen the description of the molecular mechanism. To this end, we will carry out the following experiments:
  
  1/Co-immunoprecipitation experiments of endogenous Syntaxin, Sec22b and E-Syt2 in cells expressing GFP as control or Longin-GFP to demonstrate that expression of the Longin domain perturbs the association of Sec22b with E-Syt2 and Syntaxin.
  
  2/PLA measuring the association between E-Syt2 and Syntaxin in cells expressing GFP as control or Longin-GFP to demonstrate that expression of the Longin domain perturbs the association between E-Syt2 and Syntaxin using a complementary approach.
  
  Unfortunately, membrane-associated, BoNTC1-cleaved syntaxin corresponds to a short fragment undetectable by available antibodies whereas the fragment detected by the antibody after BoNTC1 cleavage lacks the transmembrane domain (Figure 7a). Therefore, we cannot perform PLA in BoNTC1-treated neurons.
  
  We are confident that further exploring the mechanism of action of the Longin domain, together with the data already in the ms, will make it very clear that the morphological effects of E-Syt2 depends on the Sec22b/Stx1 complex.
  
  Link between neurite outgrowth and lipid transfer is weak. The authors argue that functional E-syt/Sec22b/Stx interaction is important for neurite outgrowth by mediating lipid transfer. The only line of evidence they provide is the absence of outgrowth effects in E-syt mutants lacking SMP or membrane spanning domains. However, from the data it is unclear whether these mutants are correctly folded, expressed and/or localized. Additional ICC stainings of the mutants in neurons are necessary to drive this point home. *
  
  *
  
  ->The mutants and siRNA have been already used and validated in Giordano et al. 2013 __(8)__, therefore we did not carry out experiments aiming at basic characterization of these reagents. To answer this request, we will show images of the subcellular localization by ICC of WT and mutant E-Syt2 in the revised Figure 6 or in a Supplementary Figure.
  
  In addition, the authors might make the link between neurite outgrowth and lipid transfer stronger by examining PM lipid levels and distribution in control, Myc-E-Syt2 and E-Syt2 mutant neurons.
  
  ->We agree with this reviewer that this question is of high relevance. In order to answer this request, we will express WT and mutant E-Syt2, purify the PM using the protocol of Figure 4 in Saheki et al __(1)__, followed by lipidomics analysis. We hope that this approach further supports our idea that E-Syts mediate an important lipid transfer mechanism towards the PM.
  
  There is no clear evidence that E-syt first binds to Sec22b, after which Stx1 leaves SNAP25 and joins the interaction. This should be indicated as speculation.
  
  * ->We will make it clear that our model in Figure 9 is a hypothetical model.
  
  An apparent discrepancy exists between the TKD E-syts effects (i.e. reduced MSC density, Fig 4) and the lack of neurite outgrowth defects in TKO E-syts. According to the proposed model, the levels of E-syt correlate with the number of MSCs and thereby neurite outgrowth. Furthermore, to knock down E-Syts, single siRNAs against the three E-syts were used in Fig4. Off-target effects are not controlled in this approach. Using multiple siRNAs and/or siRNA resistant rescues would be required for robust conclusions.
  
  *
  
  ->The mutants and siRNA have already been used and validated in Giordano et al. 2013 __(8)__, therefore we did not carry out experiments aiming at basic characterization of these reagents. In addition, we would like to stress the complexity of carrying out a rescue experiment of a triple KD of proteins.
  
  Statistical analysis is incomplete. It is not clear whether statistical assumptions (e.g. normal distribution) were checked before performing the tests, and whether non-parametric alternatives where used if assumptions were not met.
  
  ->We thank this reviewer for making this important alert. We would like to stress that we have always checked whether samples followed the normal distribution and made non-parametric tests__. We will include this comment in the methods.__
  
  In Fig4, a T-test is used between multiple groups. This test can only be used when comparing two groups. Number of (independent) measurements is not clear in Fig1, 2, 4
  
  ->In all the figure legends the number of repetitions is specified
  
  All figures: display all individual data points in all bar graphs (as shown in 5c)
  
  *
  
  *
  
  \*Minor comments:**
  
  Inconsistencies on distances in model. Syts are enlongated proteins and thought to be found in MSCs of ~20 nm (Fernandez-Busnadiego, 2015). Trans-SNARE complexes start to interact when the distance between membranes is ~8 nm (Liu, 2007). In the introduction, the authors suggest that incomplete zippering might occur between Stx and Sec22b, resulting in a distance between 10 and 20 nm, which would allow E-Syt localization. In the discussion, however, the authors suggest a model where Sec22b/Stx interaction is important to bring the membranes in ~10 nm distance to enhance LTP activity. Proof for either model is lacking. Please clarify.*
  
  Fig1A: Please clarify the multiple bands? for Stx3 (anti-eGFP).
  
  *
  
  ->These additional bands are recognised by the anti-GFP antibody, the tag being N-terminal, thus they represent proteolytic fragments. We consistently observe these in our experiments.
  
  Fig2: There is no size marker for panels C1-C6
  
  *
  
  ->We will make the appropriate correction.
  
  Fig3: Both proteins seem to show a diffuse pattern. Please specify the validity of measuring average distance. A higher magnification zoom of staining pattern in the growth cone and visualization of the calculation could benefit interpretation.
  
  *
  
  ->We agree with this reviewer that Figure 3 was not optimal to show all the extent of our STED data. In order to strengthen this part, we will include more details in both the figure and as a supplementary movie and supplementary spreadsheet with the quantification of the distance between the E-Syt2/Sec22b clusters to the plasma membrane stained using WGA. The STED data demonstrate that 50% of the clusters are closer than 33.6nm to the plasma membrane, a distance in the range of ER-PM contact sites.
  
  E-Syt2 and E-Syt3 are used interchangeably throughout the manuscript and E-Syt1 is left out completely. It would help the reader if the authors could elaborate on their interpretation on the similarities and differences in structure and functionality between the three E-Syts.
  
  Why is there a red line in Fig 7b?*
  
  __->We added the red line to highlight the shift of SNAP25 band in BoNTA samples. If misleading, it can be removed
  
  Reviewer #3 (Significance (Required)):__
  
  A growing body of literature recognizes the importance of close proximities between membranes, facilitating direct interaction between organelles (Scorrano et al., 2019). Membrane Contact Sites (MCSs) are shown to be important for a wide range of cellular functions, such as lipid and calcium transfer. E-Syts have been recognized as one of the key players in neuronal MCSs, mediating lipid transport (Fernández-Busnadiego et al., 2015). A study published in 2014 by the authors of the current study revealed another two proteins important for MSCs in neurons (Petkovic et al., 2014). ER protein Sec22b and PM SNARE Syntaxin1 were shown to form a non-fusogenic trans-SNARE complex, important for lipid-transfer mediated neurite outgrowth. Gallo and colleagues have now provided important new evidence that these two components (E-Syts and Stx1/Sec22b) are together and may work together at MSCs.
  
  ->We thank this reviewer for stressing the importance of our article and agree with the conclusion of __Fernández-Busnadiego et al. (9) on E-Syts being one of the key players in neuronal MCSs, mediating lipid transport. We think that our work is a further key piece of evidence in the demonstration of the importance of E-Syts in neuronal development.__
  
  Bibliography
  
  Saheki Y, Bian X, Schauder CM, Sawaki Y, Surma MA, Klose C, et al. Control of plasma membrane lipid homeostasis by the extended synaptotagmins. Nat Cell Biol. 2016 Apr 11;18(5):504–515. Yu H, Liu Y, Gulbranson DR, Paine A, Rathore SS, Shen J. Extended synaptotagmins are Ca2+-dependent lipid transfer proteins at membrane contact sites. Proc Natl Acad Sci USA. 2016 Apr 19;113(16):4362–4367. Bian X, Zhang Z, Xiong Q, De Camilli P, Lin C. A programmable DNA-origami platform for studying lipid transfer between bilayers. Nat Chem Biol. 2019 Jul 18;15(8):830–837. Alberts B, Johnson A, Lewis J, Raff M. The lipid bilayer. Molecular Biology of …. 2002; Osen-Sand A, Staple JK, Naldi E, Schiavo G, Rossetto O, Petitpierre S, et al. Common and distinct fusion proteins in axonal growth and transmitter release. J Comp Neurol. 1996 Apr 1;367(2):222–234. Igarashi M, Kozaki S, Terakawa S, Kawano S, Ide C, Komiya Y. Growth cone collapse and inhibition of neurite growth by Botulinum neurotoxin C1: a t-SNARE is involved in axonal growth. J Cell Biol. 1996 Jul;134(1):205–215. Park N, Yoo JC, Lee Y-S, Choi HY, Hong S-G, Hwang EM, et al. Copine1 C2 domains have a critical calcium-independent role in the neuronal differentiation of hippocampal progenitor HiB5 cells. Biochem Biophys Res Commun. 2014 Nov 7;454(1):228–233. Giordano F, Saheki Y, Idevall-Hagren O, Colombo SF. PI (4, 5) P2-dependent and Ca2+-regulated ER-PM interactions mediated by the extended synaptotagmins. Cell. 2013; Fernández-Busnadiego R, Saheki Y, De Camilli P. Three-dimensional architecture of extended synaptotagmin-mediated endoplasmic reticulum-plasma membrane contact sites. Proc Natl Acad Sci USA. 2015 Apr 21;112(16):E2004–13.
  
  PeerReviewed
Visit annotations in context

Tags

PeerReviewed

Annotators

EMBOpress

URL

biorxiv.org/content/10.1101/2019.12.24.886648v1
outilstice.com outilstice.com

Resoomer. Créer un résumé de n'importe quel texte

1
1. sotest 11 Mar 2020
  
  in Public
  
  vec des articles de presse, des textes scientifiques, des textes d’histoire ou des commentaires d’œuvres
  
  Dommage, je ne peux pas choisir la couleur du surlignage, mais je peux mettre un tag
Visit annotations in context

Annotators

sotest

URL

outilstice.com/2017/06/resoomer-creer-un-resume-de-nimporte-quel-texte/
ecmoed.blog ecmoed.blog

COVID19 – ECMOed

1
1. ikoln16mari 11 Mar 2020
  
  in Public
  
  unidad_COVID2019
  
  unidad_COVID2019 UCOVID19_infografías schema.org/infographic difusion CIE-10B97(COVID-19)
Visit annotations in context

Tags

unidad_COVID2019

difusion

UCOVID19_infografías

CIE-10B97(COVID-19)

schema.org/infographic

Annotators

ikoln16mari

URL

ecmoed.blog/tag/covid19/
gacetamedica.com gacetamedica.com

Covid-19 Archives - Gaceta Médica

1
1. lmichan 10 Mar 2020
  
  in Public
  
  unidad_COVID2019,favorito
  
  es unidad_COVID2019
Visit annotations in context

Tags

es

unidad_COVID2019

Annotators

lmichan

URL

gacetamedica.com/tag/covid-19/
www.analyticsmania.com www.analyticsmania.com

Setup GDPR Cookie Consent Notification with Google Tag Manager - Analytics Mania

5
1. TylerRick 10 Mar 2020
  
  in Public
  
  To be fully compliant with GDPR, you would also need to enable Show Reject All Button setting.
  
  GDPR cookie consent
2. TylerRick 09 Mar 2020
  
  in Public
  
  Consent Model. In the case of GDPR, you must choose the Opt-in. This means that you cannot start tracking people before the consent was given.
  
  cookie consent tracking (Internet) GDPR
3. TylerRick 03 Mar 2020
  
  in Public
  
  This cookie consent notification is just a tool for getting consent, it’s not capable of managing your tracking tags because every website and every GTM container is unique, therefore there is no universal solution. As a result, you will have to manually update all your tracking tags with additional firing rules.
  
  cookie consent
4. TylerRick 03 Mar 2020
  
  in Public
  
  Configuring OneTrust’s cookie consent solution is just half of the task. Your tracking scripts (like Google Analytics, Google Adwords, etc.) will still continue working as they always did unless you import my GTM recipe and then reconfigure all of your tracking tags. Yup, there’s a lot of manual work waiting ahead.
  
  cookie consent
5. TylerRick 03 Mar 2020
  
  in Public
  
  if you are using some tools/scripts on your website that are used to identify individuals and their data is processed by you or 3rd parties), that can be done only when a person gives consent
  
  GDPR cookie consent
Visit annotations in context

Tags

GDPR

tracking (Internet)

cookie consent

Annotators

TylerRick

URL

analyticsmania.com/post/gdpr-cookie-consent-notification-with-google-tag-manager/
en.wikipedia.org en.wikipedia.org

New South Wales Citizens Bushmen - Wikipedia

1
1. katelyn 09 Mar 2020
  
  in Public
  
  ron and Headq
  
  hello - edit
  
  tag tag2
Visit annotations in context

Tags

tag

tag2

Annotators

katelyn

URL

en.wikipedia.org/wiki/New_South_Wales_Citizens_Bushmen
en.wikipedia.org en.wikipedia.org

Outlaw: The Saga of Gisli - Wikipedia

1
1. katelyn 09 Mar 2020
  
  in Public
  
  hello
  
  tag
Visit annotations in context

Tags

tag

Annotators

katelyn

URL

en.wikipedia.org/wiki/Outlaw:_The_Saga_of_Gisli
hypothes.is hypothes.is

Hypothesis

1
1. marvinanha 09 Mar 2020
  
  in Public
  
  started
  
  tag
Visit annotations in context

Tags

tag

Annotators

marvinanha

URL

hypothes.is/users/marvinanha
www.vialogues.com www.vialogues.com

Waiting for You

1
1. tellio 09 Mar 2020
  
  in Public
  
  Where will we tag this?remind me.Where will we tag this?And with what?remind me
  
  https://youtu.be/7Exi0cCFAfU
Visit annotations in context

Annotators

tellio

URL

vialogues.com/vialogues/play/55282
dev.to dev.to

Guide for the daily JavaScripter - DEV Community 👩‍💻👨‍💻

1
1. pyxelr 08 Mar 2020
  
  in Public
  
  Use JSDOC https://jsdoc.app/about-getting-started.html format.
  
  Standardise your JavaScript comments:
  
  Use block comment
  /** This is a description of the foo function. */ function foo() { }
  
  Use JSDOC tag to describe a function: ```javascript /**
  
  Represents a book.
  
  @constructor
  
  @param {string} title - The title of the book.
  
  @param {string} author - The author of the book. */ function Book(title, author) { } ```
  
  JavaScript
Visit annotations in context

Tags

JavaScript

Annotators

pyxelr

URL

dev.to/damxipo/guide-for-the-daily-javascripter-87o
donohuehumn100.blogs.bucknell.edu donohuehumn100.blogs.bucknell.edu

Reflection Essay #4

1
1. gtorres4128 03 Mar 2020
  
  in Public
  
  As explained by Flanders, “An element has three core constituent parts: the start tag, the end tag, and the content, which is whatever lies between the start tag and the end tag” (Flanders 106).
  
  This is a great quote fro Julia Flanders, I came upon this text as well and really helped me understand the reason behind encoding.
Visit annotations in context

Annotators

gtorres4128

URL

donohuehumn100.blogs.bucknell.edu/2020/03/02/reflection-essay-4/
support.google.com support.google.com

About Tags - Tag Manager Help

1
1. TylerRick 03 Mar 2020
  
  in Public
  
  Tags in the digital marketing and analytics context are similar to, but distinct from, the standard HTML tags that developers will use to code web pages. The analytics version of the word "tag" is derived from the fact that the tags provided by vendors are often encapsulated by HTML <script> or <img> tags. When we speak of tags in an HTML context, we refer to tags such as <body>, <p>, <li>, <blockquote>, and so on. When we refer to tags used in the analytics and marketing industry, we refer to code that an organization provides to install the desired product or functionality on your website or mobile app.
  
  definition
Visit annotations in context

Tags

definition

Annotators

TylerRick

URL

support.google.com/tagmanager/answer/3281060
engineering.instawork.com engineering.instawork.com

When XML beats JSON: UI layouts

1
1. pyxelr 02 Mar 2020
  
  in Public
  
  The advantages of XML over JSON for trees becomes more pronounced when we introduce different node types. Assume we wanted to introduce departments into the org chart above. In XML, we can just use an element with a new tag name
  
  XML JSON
Visit annotations in context

Tags

JSON

XML

Annotators

pyxelr

URL

engineering.instawork.com/when-xml-beats-json-ui-layouts-53c7f1d3fdb7
blog.axosoft.com blog.axosoft.com

Git Dos and Don’ts

1
1. pyxelr 02 Mar 2020
  
  in Public
  
  Create a Git repository for every new project. Learn more about what a Git repo is in this beginner Learning Git with GitKraken tutorial. Always create a new branch for every new feature and bug. Regularly commit and push changes to the remote branch to avoid loss of work. Include a gitignore file in your project to avoid unwanted files being committed. Always commit changes with a concise and useful commit message. Utilize git-submodule for large projects. Keep your branch up to date with development branches. Follow a workflow like Gitflow. There are many workflows available, so choose the one that best suits your needs. Always create a pull request for merging changes from one branch to another. Learn more about what a pull request is and how to create them in this intermediate Learning Git with GitKraken tutorial. Always create one pull request addressing one issue. Always review your code once by yourself before creating a pull request. Have more than one person review a pull request. It’s not necessary, but is a best practice. Enforce standards by using pull request templates and adding continuous integrations. Learn more about enhancing the pull request process with templates. Merge changes from the release branch to master after each release. Tag the master sources after every release. Delete branches if a feature or bug fix is merged to its intended branches and the branch is no longer required. Automate general workflow checks using Git hooks. Learn more about how to trigger Git hooks in this intermediate Learning Git with GitKraken tutorial. Include read/write permission access control to repositories to prevent unauthorized access. Add protection for special branches like master and development to safeguard against accidental deletion.
  
  Git Dos
  
  Git
Visit annotations in context

Tags

Git

Annotators

pyxelr

URL

blog.axosoft.com/git-dos-donts/
github.com github.com

Implement X-Robots-tag headers (avoid dev/stage websites "leaking" into SERPs) · Issue #804 · h5bp/html5-boilerplate

1
1. TylerRick 02 Mar 2020
  
  in Public
  
  SetEnvIfNoCase Host "^www" crawlable Header set X-Robots-Tag "noindex, nofollow" env=!crawlable
  
  good example
Visit annotations in context

Tags

good example

Annotators

TylerRick

URL

github.com/h5bp/html5-boilerplate/issues/804
sutherlandhumn100.blogs.bucknell.edu sutherlandhumn100.blogs.bucknell.edu

Reflection Essay 4

2
1. jzm001 02 Mar 2020
  
  in Public
  
  Throughout the process, there were situations where I was torn between different ways to tag certain words or phrases.
  
  From class discussion and personal experience, this problem was common for the entirety of the assignment. In regards to your encounter with the dog, by identifying if the dog contains human traits would allow you to label him as a person versus an object. In this case, the dog is compared to a person, ensuring why the dog should be tagged as a person.
2. jzm001 02 Mar 2020
  
  in Public
  
  I found this quote to be very true because although it was harder to do the semantic encoding because I had to make certain decisions about what to tag, I found it more interesting. The structural encoding was very tedious because I had to go line by line giving the same tags over and over again.
  
  I agree. The structural encoding was repetitive, tedious, and boring. Whereas the semantic encoding differed in regards to tagging objects, people, places, and concepts. However, both are required to acquire a PDF document comprised with color coded words in relationship to their specific tagging.
Visit annotations in context

Annotators

jzm001

URL

sutherlandhumn100.blogs.bucknell.edu/2020/02/29/reflection-essay-4/
developers.google.com developers.google.com

Robots meta tag, data-nosnippet, and X-Robots-Tag specifications

1
1. TylerRick 02 Mar 2020
  
  in Public
  
  The <meta name="robots" content="noindex" /> tag or directive applies to search engine crawlers. To block non-search crawlers, such as AdsBot-Google, you might need to add directives targeted to the specific crawler
  
  deceptive unintuitive confusing Google Search
Visit annotations in context

Tags

Google Search

unintuitive

confusing

deceptive

Annotators

TylerRick

URL

developers.google.com/search/reference/robots_meta_tag
orzellhumn100.blogs.bucknell.edu orzellhumn100.blogs.bucknell.edu

Reflection Essay #4

2
1. cushing_j 02 Mar 2020
  
  in Public
  
  Learning will not happen quickly and it will be a good journey to further educate myself in the space.
  
  I think you did a great job, but maybe for the module assignment talk about your experience collaborating with the class about how to tag specific words.
2. cushing_j 02 Mar 2020
  
  in Public
  
  . I felt that finishing this before the rest of the encoding made things simpler because once we started encoding different parts of the songs,
  
  I followed the same process to my encoding as you because I felt that it also made it easier to tag specific words after encoding the lines, speakers, speech, and title.
Visit annotations in context

Annotators

cushing_j

URL

orzellhumn100.blogs.bucknell.edu/2020/03/02/reflection-essay-4/
bp387lpw.myraidbox.de bp387lpw.myraidbox.de

Rapid Application Development: groß, schnell, einfach mit yuuvis® RAD

1
1. Sven.Hoffmann.OS 02 Mar 2020
  
  in Public
  
  <img loading="lazy" class='avia_image' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/illu-innovate-01.svg' alt='Die Abbildung zeigt einen Mann, der vor einem Desktop sitzt' title='illu-innovate-01' height="500" width="458" itemprop="thumbnailUrl" />yuuvis® RAD – die optimale Anwendung, um Ihren Content zu managen: komplett neu entwickelter Technologie-Stack perfekt in andere Systeme integrierbar API-First entwickelt auf Microservices-Architektur basierend Low-Code-Plattform hoch skalierbar und äußerst fehlertolerant Was ist yuuvis® RAD? Umfassendes Dokumentenmanagement dank schneller Anwendungsentwicklung yuuvis® RAD ist ein umfassendes Framework für schnelle Anwendungsentwicklung. Mit ihm angepasste Applikationen helfen, das gesamte Wissen eines Unternehmens zu schützen, zu verwalten und zu organisieren. Es lässt sich flexibel erweitern und an individuelle Anforderungen anpassen. Applikationen können clientseitig neu entwickelt oder einfach modifiziert werden. Durch eigene „Custom“-Microservices wird die Geschäftslogik nach Bedarf erweitert. <img loading="lazy" class='avia_image' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/illu-creative-process-tuerkis-01.svg' alt='Die Abbildung zeigt drei Personen, die um eine Glühbirne herumstehen' title='illu-creative-process-tuerkis-01' height="500" width="449" itemprop="thumbnailUrl" />Mehr informationen zu yuuvis® RAD Sie möchten mehr zu yuuvis® RAD erfahren? <img loading="lazy" class='avia_image ' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/yuuvis-whitepaper-rad-preview-de.png' alt='yuuvis whitepaper rad vorschau' title='yuuvis-whitepaper-rad-preview-de' itemprop="thumbnailUrl" /> Offene Standards Architektur der Microservices Rahmenkonzept Übersicht der Dienstleistungen Modellierungs- und Anpassungswerkzeuge Microsoft-Integration Laden Sie unser Whitepaper herunter! Hinweis: Für diesen Inhalt ist JavaScript erforderlich. Felder mit einem * sind Pflichtfelder. Bitte Anrede auswählen: * Sehr geehrte Frau Sehr geehrter Herr Guten Tag Vorname Nachname * Email * Ihre Nachricht an uns Divider HTML Erleben Sie yuuvis® live! Die Einführung der Software-Lösung ist geplant. Ich bin an einer Software-Präsentation interessiert. Divider HTML Felder mit * sind Pflichtfelder. Falls du ein Mensch bist und dieses Feld siehst, dann lasse es bitte leer. var formDisplay=1;var nfForms=nfForms||[];var form=[];form.id='3';form.settings={"objectType":"Form Setting","editActive":true,"title":"yuuvis-Downloadformular RAD DE","created_at":"2020-01-30 16:02:46","default_label_pos":"above","show_title":0,"clear_complete":"1","hide_complete":"1","logged_in":"","wrapper_class":"","element_class":"","key":"yuuvis-downloadformular-en","add_submit":"0","changeEmailErrorMsg":"Please enter a valid e-mail address!","changeDateErrorMsg":"Bitte gib ein g\u00fcltiges Datum ein!","confirmFieldErrorMsg":"Diese Felder m\u00fcssen \u00fcbereinstimmen!","fieldNumberNumMinError":"Die eingegebene Zahl ist zu klein.","fieldNumberNumMaxError":"Die eingegebene Zahl ist zu gross.","fieldNumberIncrementBy":"Bitte erh\u00f6hen in Schritten von","formErrorsCorrectErrors":"Bitte korrigieren Sie die angezeigten Fehler, bevor Sie das Formular absenden.","validateRequiredField":"Dies ist ein Pflichtfeld.","honeypotHoneypotError":"Es ist ein Fehler aufgetreten.","fieldsMarkedRequired":"Felder mit einem * sind Pflichtfelder.","currency":"","unique_field_error":"A form with this value has already been submitted.","not_logged_in_msg":"","sub_limit_msg":"The form has reached its submission limit.","calculations":[],"container_styles_show_advanced_css":"0","title_styles_show_advanced_css":"0","row_styles_show_advanced_css":"0","row-odd_styles_show_advanced_css":"0","success-msg_styles_show_advanced_css":"0","error_msg_styles_show_advanced_css":"0","conditions":[],"mp_breadcrumb":0,"mp_progress_bar":"1","mp_display_titles":0,"breadcrumb_container_styles_show_advanced_css":"0","breadcrumb_buttons_styles_show_advanced_css":"0","breadcrumb_button_hover_styles_show_advanced_css":"0","breadcrumb_active_button_styles_show_advanced_css":"0","progress_bar_container_styles_show_advanced_css":"0","progress_bar_fill_styles_show_advanced_css":"0","part_titles_styles_show_advanced_css":"0","navigation_container_styles_show_advanced_css":"0","previous_button_styles_show_advanced_css":"0","next_button_styles_show_advanced_css":"0","navigation_hover_styles_show_advanced_css":"0","formContentData":[{"formContentData":[{"order":1,"cells":[{"order":0,"fields":["bitte_anrede_auswaehlen_1580396600390"],"width":"100"}]},{"order":2,"cells":[{"order":0,"fields":["vorname_1580396717767"],"width":50},{"order":1,"fields":["nachname_1580820256875"],"width":50}]},{"order":4,"cells":[{"order":0,"fields":["email_1580391190571"],"width":"100"}]},{"order":5,"cells":[{"order":0,"fields":["ihre_nachricht_an_uns_1581506628019"],"width":"100"}]},{"order":6,"cells":[{"order":0,"fields":["hr_1580820679276"],"width":"100"}]},{"order":7,"cells":[{"order":0,"fields":["html_1581506435696"],"width":"100"}]},{"order":8,"cells":[{"order":0,"fields":["softwaresolution_1581506481772"],"width":"100"}]},{"order":9,"cells":[{"order":0,"fields":["softwarepresentation_1581506440960"],"width":"100"}]},{"order":10,"cells":[{"order":0,"fields":["hr_1581506444943"],"width":"100"}]},{"order":11,"cells":[{"order":0,"fields":["html_1581506446831"],"width":"100"}]}],"order":0,"type":"part","clean":false,"title":"1","key":"hgorwjor","editActive":true,"drawerDisabled":false},{"formContentData":[{"order":1,"cells":[{"order":2,"fields":["telefon_1582895904537","position_1580820607732"],"width":50},{"order":3,"fields":["plz_1580820645250","unternehmen_1581582782634"],"width":50}]},{"order":2,"cells":[{"order":0,"fields":["hr_1582556164156"],"width":"100"}]},{"order":3,"cells":[{"order":0,"fields":["html_1580391371746"],"width":"100"}]},{"order":4,"cells":[{"order":0,"fields":["newsletter_checkbox_1580391409240"],"width":"100"}]},{"order":5,"cells":[{"order":0,"fields":["datenschutz_1580392331180"],"width":"100"}]},{"order":6,"cells":[{"order":0,"fields":["hr_1582556170397"],"width":"100"}]},{"order":7,"cells":[{"order":0,"fields":["html_1581506553720"],"width":"100"}]},{"order":8,"cells":[{"order":0,"fields":["hidden_optinherkunft_1580392579819"],"width":"100"}]},{"order":9,"cells":[{"order":0,"fields":["hidden_sprache_1580392763492"],"width":"100"}]},{"order":11,"cells":[{"order":0,"fields":["page-url_1580393093116"],"width":"100"}]},{"order":12,"cells":[{"order":0,"fields":["yuuvis-produkt_1581512743513"],"width":"100"}]},{"order":13,"cells":[{"order":0,"fields":["jetzt_whitepaper_anfordern_1582641368769"],"width":"100"}]}],"order":1,"type":"part","clean":false,"title":"2","key":"glmhxs","editActive":true,"drawerDisabled":false}],"drawerDisabled":false,"manual_key":"1","allow_public_link":0,"embed_form":"","container_styles_border":"","container_styles_height":"","container_styles_margin":"","container_styles_padding":"","container_styles_float":"","title_styles_border":"","title_styles_height":"","title_styles_width":"","title_styles_font-size":"","title_styles_margin":"","title_styles_padding":"","title_styles_float":"","row_styles_border":"","row_styles_width":"","row_styles_font-size":"","row_styles_margin":"","row_styles_padding":"","row-odd_styles_border":"","row-odd_styles_height":"","row-odd_styles_width":"","row-odd_styles_font-size":"","row-odd_styles_margin":"","row-odd_styles_padding":"","success-msg_styles_border":"","success-msg_styles_height":"","success-msg_styles_width":"","success-msg_styles_font-size":"","success-msg_styles_margin":"","success-msg_styles_padding":"","error_msg_styles_border":"","error_msg_styles_width":"","error_msg_styles_font-size":"","error_msg_styles_margin":"","error_msg_styles_padding":"","mp_validate":1,"mp_prev_label":"Zur\u00fcck","mp_next_label":"Weiter","breadcrumb_container_styles_border":"","breadcrumb_container_styles_height":"","breadcrumb_container_styles_width":"","breadcrumb_container_styles_margin":"","breadcrumb_container_styles_padding":"","breadcrumb_container_styles_float":"","breadcrumb_buttons_styles_border":"","breadcrumb_buttons_styles_height":"","breadcrumb_buttons_styles_width":"","breadcrumb_buttons_styles_font-size":"","breadcrumb_buttons_styles_margin":"","breadcrumb_buttons_styles_padding":"","breadcrumb_buttons_styles_float":"","breadcrumb_button_hover_styles_border":"","breadcrumb_button_hover_styles_height":"","breadcrumb_button_hover_styles_width":"","breadcrumb_button_hover_styles_font-size":"","breadcrumb_button_hover_styles_margin":"","breadcrumb_button_hover_styles_padding":"","breadcrumb_button_hover_styles_float":"","breadcrumb_active_button_styles_border":"","breadcrumb_active_button_styles_height":"","breadcrumb_active_button_styles_width":"","breadcrumb_active_button_styles_font-size":"","breadcrumb_active_button_styles_margin":"","breadcrumb_active_button_styles_padding":"","breadcrumb_active_button_styles_float":"","progress_bar_container_styles_border":"","progress_bar_container_styles_height":"","progress_bar_container_styles_width":"","progress_bar_container_styles_margin":"","progress_bar_container_styles_padding":"","progress_bar_container_styles_float":"","progress_bar_fill_styles_border":"","progress_bar_fill_styles_height":"","progress_bar_fill_styles_margin":"","progress_bar_fill_styles_padding":"","part_titles_styles_border":"","part_titles_styles_height":"","part_titles_styles_width":"","part_titles_styles_font-size":"","part_titles_styles_margin":"","part_titles_styles_padding":"","part_titles_styles_float":"","navigation_container_styles_border":"","navigation_container_styles_height":"","navigation_container_styles_width":"","navigation_container_styles_margin":"","navigation_container_styles_padding":"","navigation_container_styles_float":"","previous_button_styles_border":"","previous_button_styles_height":"","previous_button_styles_width":"","previous_button_styles_font-size":"","previous_button_styles_margin":"","previous_button_styles_padding":"","next_button_styles_border":"","next_button_styles_height":"","next_button_styles_width":"","next_button_styles_font-size":"","next_button_styles_margin":"","next_button_styles_padding":"","navigation_hover_styles_border":"","navigation_hover_styles_height":"","navigation_hover_styles_width":"","navigation_hover_styles_font-size":"","navigation_hover_styles_margin":"","navigation_hover_styles_padding":"","ninjaForms":"Ninja Forms","fieldTextareaRTEInsertLink":"Link einf\u00fcgen","fieldTextareaRTEInsertMedia":"Medien einf\u00fcgen","fieldTextareaRTESelectAFile":"Eine Datei ausw\u00e4hlen","formHoneypot":"Falls du ein Mensch bist und dieses Feld siehst, dann lasse es bitte leer.","fileUploadOldCodeFileUploadInProgress":"Datei-Upload l\u00e4uft.","fileUploadOldCodeFileUpload":"DATEI-UPLOAD","currencySymbol":"€","thousands_sep":".","decimal_point":",","siteLocale":"de_DE","dateFormat":"m\/d\/Y","startOfWeek":"1","of":"von","previousMonth":"Vorheriger Monat","nextMonth":"N\u00e4chster Monat","months":["Januar","Februar","M\u00e4rz","April","Mai","Juni","Juli","August","September","Oktober","November","Dezember"],"monthsShort":["Jan","Feb","Mrz","Apr","Mai","Jun","Jul","Aug","Sep","Okt","Nov","Dez"],"weekdays":["Sonntag","Montag","Dienstag","Mittwoch","Donnerstag","Freitag","Samstag"],"weekdaysShort":["So","Mo","Die","Mi","Do","Fr","Sa"],"weekdaysMin":["So","Mo","Di","Mi","Do","Fr","Sa"],"currency_symbol":"","beforeForm":"","beforeFields":"","afterFields":"","afterForm":""};form.fields=[{"objectType":"Field","objectDomain":"fields","editActive":false,"order":1,"label":"Bitte Anrede ausw\u00e4hlen:","type":"listradio","key":"bitte_anrede_auswaehlen_1580396600390","label_pos":"above","required":1,"container_class":"","element_class":"","admin_label":"","help_text":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"list_item_row_styles_border":"","list_item_row_styles_height":"","list_item_row_styles_width":"","list_item_row_styles_font-size":"","list_item_row_styles_margin":"","list_item_row_styles_padding":"","list_item_row_styles_float":"","list_item_row_styles_show_advanced_css":0,"list_item_label_styles_border":"","list_item_label_styles_height":"","list_item_label_styles_width":"","list_item_label_styles_font-size":"","list_item_label_styles_margin":"","list_item_label_styles_padding":"","list_item_label_styles_float":"","list_item_label_styles_show_advanced_css":0,"list_item_element_styles_border":"","list_item_element_styles_height":"","list_item_element_styles_width":"","list_item_element_styles_font-size":"","list_item_element_styles_margin":"","list_item_element_styles_padding":"","list_item_element_styles_float":"","list_item_element_styles_show_advanced_css":0,"cellcid":"c3776","drawerDisabled":false,"options":[{"errors":[],"max_options":0,"label":"Sehr geehrte Frau","value":"1","calc":"1","selected":0,"order":0,"settingModel":{"settings":false,"hide_merge_tags":false,"error":false,"name":"options","type":"option-repeater","label":"Options <a href=\"#\" class=\"nf-add-new\">Add New<\/a> <a href=\"#\" class=\"extra nf-open-import-tooltip\"><i class=\"fa fa-sign-in\" aria-hidden=\"true\"><\/i> Import<\/a>","width":"full","group":"","value":[{"label":"One","value":"one","calc":"","selected":0,"order":0},{"label":"Two","value":"two","calc":"","selected":0,"order":1},{"label":"Three","value":"three","calc":"","selected":0,"order":2}],"columns":{"label":{"header":"Label","default":""},"value":{"header":"Value","default":""},"calc":{"header":"Calc Value","default":""},"selected":{"header":"<span class=\"dashicons dashicons-yes\"><\/span>","default":0}}},"manual_value":true},{"errors":[],"max_options":0,"label":"Sehr geehrter Herr","value":"2","calc":"2","selected":0,"order":1,"settingModel":{"settings":false,"hide_merge_tags":false,"error":false,"name":"options","type":"option-repeater","label":"Options <a href=\"#\" class=\"nf-add-new\">Add New<\/a> <a href=\"#\" class=\"extra nf-open-import-tooltip\"><i class=\"fa fa-sign-in\" aria-hidden=\"true\"><\/i> Import<\/a>","width":"full","group":"","value":[{"label":"One","value":"one","calc":"","selected":0,"order":0},{"label":"Two","value":"two","calc":"","selected":0,"order":1},{"label":"Three","value":"three","calc":"","selected":0,"order":2}],"columns":{"label":{"header":"Label","default":""},"value":{"header":"Value","default":""},"calc":{"header":"Calc Value","default":""},"selected":{"header":"<span class=\"dashicons dashicons-yes\"><\/span>","default":0}}},"manual_value":true},{"errors":[],"max_options":0,"label":"Guten Tag","value":"3","calc":"3","selected":0,"order":2,"settingModel":{"settings":false,"hide_merge_tags":false,"error":false,"name":"options","type":"option-repeater","label":"Options <a href=\"#\" class=\"nf-add-new\">Add New<\/a> <a href=\"#\" class=\"extra nf-open-import-tooltip\"><i class=\"fa fa-sign-in\" aria-hidden=\"true\"><\/i> Import<\/a>","width":"full","group":"","value":[{"label":"One","value":"one","calc":"","selected":0,"order":0},{"label":"Two","value":"two","calc":"","selected":0,"order":1},{"label":"Three","value":"three","calc":"","selected":0,"order":2}],"columns":{"label":{"header":"Label","default":""},"value":{"header":"Value","default":""},"calc":{"header":"Calc Value","default":""},"selected":{"header":"<span class=\"dashicons dashicons-yes\"><\/span>","default":0}}},"manual_value":true}],"field_label":"Please choose salutation:","field_key":"please_choose_salutation_1580391018648","id":18,"beforeField":"","afterField":"","value":"","parentType":"list","element_templates":["listradio","input"],"old_classname":"list-radio","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":2,"label":"Vorname","type":"firstname","key":"vorname_1580396717767","label_pos":"hidden","required":"","default":"","placeholder":"Vorname","container_class":"","element_class":"","admin_label":"","help_text":"","custom_name_attribute":"fname","personally_identifiable":1,"wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3780","value":"","drawerDisabled":false,"field_label":"First Name","field_key":"firstname_1580391082629","id":19,"beforeField":"","afterField":"","parentType":"firstname","element_templates":["firstname","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":3,"label":"Nachname","type":"lastname","key":"nachname_1580820256875","label_pos":"hidden","required":1,"default":"","placeholder":"Nachname*","container_class":"","element_class":"","admin_label":"","help_text":"","custom_name_attribute":"lname","personally_identifiable":1,"wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3782","value":"","drawerDisabled":false,"field_label":"Last Name","field_key":"lastname_1580391137379","id":20,"beforeField":"","afterField":"","parentType":"lastname","element_templates":["lastname","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":4,"type":"email","label":"Email","key":"email_1580391190571","label_pos":"hidden","required":1,"default":"","placeholder":"E-Mail-Adresse*","container_class":"","element_class":"","admin_label":"","help_text":"","custom_name_attribute":"email","personally_identifiable":1,"wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3785","value":"","drawerDisabled":false,"field_label":"Email","field_key":"email_1580391190571","id":21,"beforeField":"","afterField":"","parentType":"email","element_templates":["email","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":5,"label":"Ihre Nachricht an uns","type":"textarea","key":"ihre_nachricht_an_uns_1581506628019","label_pos":"hidden","required":false,"default":"","placeholder":"Ihre Nachricht an uns","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":false,"admin_label":"","help_text":"","textarea_rte":"","disable_rte_mobile":"","textarea_media":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3788","value":"","drawerDisabled":false,"id":333,"beforeField":"","afterField":"","parentType":"textarea","element_templates":["textarea","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":8,"label":"Divider","type":"hr","container_class":"","element_class":"","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"key":"hr_1580820679276","cellcid":"c3791","oldCellcid":"c3774","id":235,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":999,"type":"html","label":"HTML","default":"<p><b>Erleben Sie yuuvis\u00ae live!<\/b><\/p>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1581506435696","cellcid":"c3794","drawerDisabled":false,"oldCellcid":"c3777","id":328,"beforeField":"","afterField":"","value":"<p><b>Erleben Sie yuuvis\u00ae live!<\/b><\/p>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":999,"type":"checkbox","label":"Die Einf\u00fchrung der Software-L\u00f6sung ist geplant.","key":"softwaresolution_1581506481772","label_pos":"right","required":false,"container_class":"","element_class":"","manual_key":true,"admin_label":"","help_text":"","default_value":"unchecked","checked_value":1,"unchecked_value":2,"checked_calc_value":"","unchecked_calc_value":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3797","drawerDisabled":false,"oldCellcid":"c3780","id":329,"beforeField":"","afterField":"","value":"","parentType":"checkbox","element_templates":["checkbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":999,"type":"checkbox","label":"Ich bin an einer Software-Pr\u00e4sentation interessiert.","key":"softwarepresentation_1581506440960","label_pos":"right","required":false,"container_class":"","element_class":"","manual_key":true,"admin_label":"","help_text":"","default_value":"unchecked","checked_value":1,"unchecked_value":2,"checked_calc_value":"","unchecked_calc_value":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3800","drawerDisabled":false,"oldCellcid":"c3783","id":330,"beforeField":"","afterField":"","value":"","parentType":"checkbox","element_templates":["checkbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":999,"type":"hr","label":"Divider","container_class":"","element_class":"","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"key":"hr_1581506444943","cellcid":"c3803","oldCellcid":"c3786","id":331,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":999,"type":"html","label":"HTML","default":"<small>Felder mit * sind Pflichtfelder.<\/small>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1581506446831","cellcid":"c3806","drawerDisabled":false,"oldCellcid":"c3789","id":332,"beforeField":"","afterField":"","value":"<small>Felder mit * sind Pflichtfelder.<\/small>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":1,"label":"Telefon","type":"phone","key":"telefon_1582895904537","label_pos":"hidden","required":false,"default":"","placeholder":"Telefon (Durchwahl)","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":false,"admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"phone","personally_identifiable":1,"wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3812","value":"","cellOrder":1,"drawerDisabled":false,"id":232,"beforeField":"","afterField":"","parentType":"textbox","element_templates":["tel","textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":2,"label":"Position","type":"textbox","cellcid":"c3812","key":"position_1580820607732","label_pos":"hidden","required":1,"default":"","placeholder":"Position*","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":false,"admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"","personally_identifiable":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"value":"","cellOrder":3,"drawerDisabled":false,"id":233,"beforeField":"","afterField":"","parentType":"textbox","element_templates":["textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":3,"label":"PLZ","type":"zip","cellcid":"c3814","key":"plz_1580820645250","label_pos":"hidden","required":false,"default":"","placeholder":"PLZ","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":false,"admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"zip","personally_identifiable":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"value":"","cellOrder":1,"drawerDisabled":false,"id":234,"beforeField":"","afterField":"","parentType":"zip","element_templates":["zip","textbox","input","textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":4,"label":"Unternehmen","type":"textbox","key":"unternehmen_1581582782634","label_pos":"hidden","required":1,"default":"","placeholder":"Ihr Unternehmen*","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":false,"admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"company","personally_identifiable":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3814","value":"","drawerDisabled":false,"oldCellcid":"c3782","cellOrder":2,"id":226,"beforeField":"","afterField":"","parentType":"textbox","element_templates":["textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":5,"label":"Divider","type":"hr","container_class":"","element_class":"","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"key":"hr_1582556164156","cellcid":"c3817","id":504,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":6,"type":"html","label":"HTML","default":"<p>Bitte best\u00e4tigen Sie Ihr Einverst\u00e4ndnis zum Erhalt weiterer Informationen sowie unsere Datenschutzbestimmungen.<br><\/p>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1580391371746","cellcid":"c3820","drawerDisabled":false,"field_label":"HTML","field_key":"html_1580391371746","oldCellcid":"c3785","id":22,"beforeField":"","afterField":"","value":"<p>Bitte best\u00e4tigen Sie Ihr Einverst\u00e4ndnis zum Erhalt weiterer Informationen sowie unsere Datenschutzbestimmungen.<br><\/p>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":7,"type":"checkbox","label":"Ja, ich m\u00f6chte relevante Produktinformationen und wichtige News erhalten.","key":"newsletter_checkbox_1580391409240","label_pos":"right","required":1,"container_class":"","element_class":"","manual_key":true,"admin_label":"","help_text":"","default_value":"unchecked","checked_value":1,"unchecked_value":2,"checked_calc_value":"","unchecked_calc_value":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3823","drawerDisabled":false,"field_label":"Yes, I would like to receive relevant product information and important news.","field_key":"newsletter_checkbox_1580391409240","oldCellcid":"c3788","id":23,"beforeField":"","afterField":"","value":"","parentType":"checkbox","element_templates":["checkbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":8,"type":"checkbox","label":"Ja, ich habe die <a href=\"\/de\/impressum-und-datenschutz\" target=\"_blank\" rel=\"noopener\">Datenschutzbestimmungen<\/a> gelesen und stimme ihnen zu.","key":"datenschutz_1580392331180","label_pos":"right","required":1,"container_class":"","element_class":"","manual_key":1,"admin_label":"","help_text":"","default_value":"unchecked","checked_value":1,"unchecked_value":2,"checked_calc_value":"","unchecked_calc_value":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3826","drawerDisabled":false,"field_label":"Yes, I have read and agree to the <a href=\"https:\/\/bp387lpw.myraidbox.de\/privacy-policy\/#data-privacy\" target=\"_blank\" rel=\"noopener\">privacy policy<\/a>.","field_key":"datenschutz_1580392331180","oldCellcid":"c3791","id":24,"beforeField":"","afterField":"","value":"","parentType":"checkbox","element_templates":["checkbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":9,"label":"Divider","type":"hr","container_class":"","element_class":"","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"key":"hr_1582556170397","cellcid":"c3829","id":505,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":10,"label":"HTML","type":"html","default":"<small>Felder mit * sind Pflichtfelder.<\/small>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1581506553720","cellcid":"c3832","drawerDisabled":false,"oldCellcid":"c3804","id":327,"beforeField":"","afterField":"","value":"<small>Felder mit * sind Pflichtfelder.<\/small>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":11,"type":"hidden","label":"Optin-Herkunft","key":"hidden_optinherkunft_1580392579819","default":"{wp:site_url}","admin_label":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3835","drawerDisabled":"","manual_key":1,"field_label":"Optin-Herkunft","field_key":"hidden_optinherkunft_1580392579819","oldCellcid":"c3794","id":25,"beforeField":"","afterField":"","value":"https:\/\/bp387lpw.myraidbox.de\/de\/","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":12,"type":"hidden","label":"SPRACHE","key":"hidden_sprache_1580392763492","default":"DE","admin_label":"sprache","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3838","drawerDisabled":false,"manual_key":1,"field_label":"Sprache","field_key":"hidden_sprache_1580392763492","oldCellcid":"c3797","id":26,"beforeField":"","afterField":"","value":"DE","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":14,"type":"hidden","label":"PAGE-URL","key":"page-url_1580393093116","default":"{wp:post_url}","admin_label":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3841","drawerDisabled":"","manual_key":1,"field_label":"PAGE-URL","field_key":"page-url_1580393093116","oldCellcid":"c3803","id":28,"beforeField":"","afterField":"","value":"https:\/\/bp387lpw.myraidbox.de\/de\/rapid-application-development\/","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":15,"label":"YUUVIS-PRODUKT","type":"hidden","key":"yuuvis-produkt_1581512743513","default":"RAD","admin_label":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3844","drawerDisabled":false,"oldCellcid":"c3819","id":354,"beforeField":"","afterField":"","value":"RAD","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":16,"type":"submit","label":"Jetzt Whitepaper anfordern","processing_label":"Daten werden \u00fcbertragen ...","container_class":"","element_class":"","key":"jetzt_whitepaper_anfordern_1582641368769","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"submit_element_hover_styles_border":"","submit_element_hover_styles_width":"","submit_element_hover_styles_font-size":"","submit_element_hover_styles_margin":"","submit_element_hover_styles_padding":"","submit_element_hover_styles_float":"","submit_element_hover_styles_show_advanced_css":0,"cellcid":"c3847","drawerDisabled":false,"field_label":"Register now","field_key":"register_now_1580393213788","oldCellcid":"c3806","id":29,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"textbox","element_templates":["submit","button","input"],"old_classname":"","wrap_template":"wrap-no-label"}];nfForms.push(form); <div class="nf-mp-header"></div> <div class="nf-mp-body"></div> <div class="nf-mp-footer"></div> {{{ data.renderProgressBar() }}} {{{ data.renderBreadcrumbs() }}} {{{ data.renderPartTitle() }}} <h3> {{{ data.title }}} </h3> {{{ data.renderNextPrevious() }}} <ul class="nf-next-previous"> <# if ( data.showPrevious ) { #> <li class="nf-previous-item"> <input type="button" class="nf-previous" value="{{{ data.prevLabel }}}" /> </li> <# } #> <# if ( data.showNext ) { #> <li class="nf-next-item"> <input type="button" class="nf-next" value="{{{ data.nextLabel }}}" /> </li> <# } #> </ul> <ul class="nf-breadcrumbs"> <# _.each( data.parts, function( part, index ) { #> <li class="{{{ ( data.currentIndex == index ) ? 'active' : '' }}} {{{ ( part.errors ) ? 'errors' : '' }}}"> <a href="#" class="nf-breadcrumb" data-index="{{{ index }}}">{{{ ( part.errors ) ? '' : '' }}} {{{ part.title }}}</a> </li> <# } ); #> </ul> <div class="nf-progress-container"> <div class="nf-progress" style="width: {{{ data.percent }}}%;"></div> </div> <nf-fields></nf-fields> <nf-cells></nf-cells> Formular schließen Wer profitiert von yuuvis® RAD? Rapid Application Development – für starke Anwendungen <img loading="lazy" class='avia_image' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/illu-startup-01.svg' alt='Die Abbildung zeigt einen Mann, der vor einem Desktop steht' title='illu-startup-01' height="500" width="489" itemprop="thumbnailUrl" /> Systemintegratoren, die Tools in einem Bruchteil der Zeit und mit geringeren Kosten integrieren <img loading="lazy" class='avia_image' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/illu-data_analysis-01.svg' alt='Die Abbildung zeigt drei Personen, die um einen Tisch herum stehen.' title='illustration-drei-personen-arbeiten-erfolgreich-an-tisch' height="500" width="458" itemprop="thumbnailUrl" /> Anwendungsdesigner, die leistungsstarke, inhaltsbasierte Applikationen erstellen <img loading="lazy" class='avia_image' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/illu-web-hosting-01.svg' alt='Die Abbildung zeigt einen Mann, der vor 4 Servern steht mit grauem Schatten' title='illu-web-hosting-01' height="428" width="500" itemprop="thumbnailUrl" /> IT-Abteilungen, die kundenspezifische Anwendungen auf webbasierter IT-Ebene betreiben Das yuuvis® RAD Framework ist die ideale Lösung für kleinere bis mittelständische Unternehmen, die on-premise hoch performante Plattformen betreiben wollen. Außerdem ist es optimal, um bestehende Anwendungen um ein Archiv zu erweitern. Die mit yuuvis® RAD implementierten Applikationen skalieren dank des modernen Microservices-Stack und des responsiven Web-Clients (Angular/HTML5) auch in mittelgroßen, weltweit verteilten Firmennetzen. Finden Sie heraus, wer von yuuvis® RAD profitieren kann Warum yuuvis® RAD? Schnelle Anwendungsentwicklung – die Vorteile auf einen Blick State of the ArtHigh-End-ECM auf dem neuesten Stand der Technik. Perfekt für content-zentrische Anwendungen. Offene ArchitekturREST-API gewährleistet einfache Integration in bestehende Systemlandschaften. Low CodeWeniger Code bedeutet weniger potenzielle Fehle, einfachere Wartung und geringere Kosten. ZukunftssicherHoher Investitionsschutz durch ein nachhaltiges Konzept gegen zukünftige disruptive Technologien. Einfaches HandlingGemeinsame Benutzerverwaltung mit sehr gut integriertem Workflow und Datenmanagement. Custom MicroservicesKomplexe Anwendungsszenarien können mit eigenen passenden Erweiterungen im Client ergänzt werden. Erfahren Sie mehr über die Vorzüge von yuuvis® RAD Sie wollen mit yuuvis® RAD arbeiten? Kontaktieren Sie uns! Wie funktioniert yuuvis® RAD? Die Technik hinter unserer Rapid Application Development-Anwendung Offene Technologie yuuvis® RAD basiert auf offenen Standardtechnologien. Alle serverseitigen Komponenten sind java-basiert und implementieren eine saubere, skalierbare und leichtgewichtige Microservices-Architektur. Diese wird von Spring Cloud Netflix orchestriert. Kommunikation und Protokoll basieren auf HTTP(S). Der Webclient ist eine hundertprozentige HTML5 Angular (Typeskript) Anwendung. Die serverseitige API ist rein RESTbasiert. Neueste Standards Die strikte Anwendung neuester Standards ermöglicht den sofortigen Zugriff auf das größte Netzwerk von Software-Experten der Welt. Proprietär ist ein Fremdwort in yuuvis® RAD. Es bietet damit die schnellste und sicherste Architektur, die heute verfügbar ist. Rechenzentrumsfreundlich Dank des Webclients ist der Rollout auf Endbenutzerarbeitsplätzen einfach und unkompliziert, ohne lokalen Installationsaufwand. Das HTTP(S)/REST-Protokoll zwischen dem Webclient und dem Server entspricht vollständig den Standard-Firewall-Technologien. Microservices-basiert Der Microservices-Stack bietet eine umfassende Integration von Wartung, Lastausgleich, Failover und Health-Check in die Management- und Überwachungswerkzeuge von Rechenzentren. yuuvis® RAD bedient sowohl die S3-Schnittstelle, den De-facto-Standard der Datenspeicherung als auch OAuth 2.0 zur standardisierten, sicheren API-Autorisierung für Desktop-, Web- und Mobile-Anwendungen. <img loading="lazy" class='avia_image ' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/yuuvis-rad-framework-infografik.jpg' alt='Infografik zeigt das yuuvis® RAD Framework' title='yuuvis-rad-framework-Konzept' itemprop="thumbnailUrl" /> Das Rahmenkonzept der yuuvis® RAD Plattform Lesen Sie mehr über die Technologie von yuuvis® RAD Wer nutzt yuuvis® RAD? Diese Unternehmen vertrauen auf schnelle Anwendungsentwicklung Sie wollen mehr erfahren? Wir sind für Sie da! <img loading="lazy" class='avia_image' src='https://bp387lpw.myraidbox.de/wp-content/uploads/2020/02/yuuvis-photo-frank-dembach.jpg' alt='Foto Senior Key Account Manager yuuvis®' title='yuuvis-foto-frank-dembach-quer' height="694" width="1000" itemprop="thumbnailUrl" /> Frank Dembach Team Leader Sales – yuuvis® +49 (0)30 – 895 70 80 info@yuuvis.com Hinweis: Für diesen Inhalt ist JavaScript erforderlich. Felder mit einem * sind Pflichtfelder. Bitte Anrede auswählen: * Sehr geehrte Frau Sehr geehrter Herr Guten Tag Vorname Nachname * Email * Ihre Nachricht an uns 160 von 160 Zeichen übrig Divider HTML Erleben Sie yuuvis® live! Die Einführung der Software-Lösung ist geplant. Ich bin an einer Software-Präsentation interessiert. Trennung HTML Felder mit * sind Pflichtfelder. Falls du ein Mensch bist und dieses Feld siehst, dann lasse es bitte leer. var formDisplay=1;var nfForms=nfForms||[];var form=[];form.id='30';form.settings={"objectType":"Form Setting","editActive":true,"title":"Allgemeines Kontaktformular-DE - RAD","created_at":"2020-02-26 15:53:54","default_label_pos":"above","show_title":"0","clear_complete":"1","hide_complete":"1","logged_in":"0","wrapper_class":"","element_class":"","key":"yuuvis-kontakt-allgemein-de","add_submit":"0","changeEmailErrorMsg":"Bitte gib eine g\u00fcltige E-Mail-Adresse ein.","changeDateErrorMsg":"Bitte gib ein g\u00fcltiges Datum ein!","confirmFieldErrorMsg":"Diese Felder m\u00fcssen \u00fcbereinstimmen!","fieldNumberNumMinError":"Die eingegebene Zahl ist zu klein.","fieldNumberNumMaxError":"Die eingegebene Zahl ist zu gross.","fieldNumberIncrementBy":"Bitte erh\u00f6hen in Schritten von","formErrorsCorrectErrors":"Bitte korrigieren die angezeigten Fehler, bevor Sie das Formular absenden.","validateRequiredField":"Dies ist ein Pflichtfeld.","honeypotHoneypotError":"Es ist ein Fehler aufgetreten.","fieldsMarkedRequired":"Felder mit einem * sind Pflichtfelder.","currency":"","unique_field_error":"Ein Formular mit diesem Wert ist bereits eingereicht worden.","not_logged_in_msg":"","sub_limit_msg":"Das Formular hat seine Zeichengrenze erreicht.","calculations":[],"formContentData":[{"formContentData":[{"order":1,"cells":[{"order":0,"fields":["bitte_anrede_auswaehlen_1580465134673"],"width":"100"}]},{"order":2,"cells":[{"order":0,"fields":["firstname_1573210589193"],"width":"50"},{"order":1,"fields":["lastname_1573210591744"],"width":"50"}]},{"order":4,"cells":[{"order":0,"fields":["email_1582725914409"],"width":"100"}]},{"order":6,"cells":[{"order":0,"fields":["ihre_nachricht_an_uns_1573565913431"],"width":"100"}]},{"order":7,"cells":[{"order":0,"fields":["hr_1581506117361"],"width":"100"}]},{"order":8,"cells":[{"order":0,"fields":["html_1581506146896"],"width":"100"}]},{"order":9,"cells":[{"order":0,"fields":["softwaresolution_1581506167859"],"width":"100"}]},{"order":10,"cells":[{"order":0,"fields":["softwarepresentation_1581506142599"],"width":"100"}]},{"order":11,"cells":[{"order":0,"fields":["hr_1573210614298"],"width":"100"}]},{"order":12,"cells":[{"order":0,"fields":["html_1574181495044"],"width":"100"}]}],"order":0,"type":"part","clean":"","title":"1","key":"tyqqgqty","editActive":true,"drawerDisabled":false},{"formContentData":[{"order":1,"cells":[{"order":0,"fields":["telefon_1582726279559","position_1580818819203"],"width":50},{"order":1,"fields":["zip_code_1582726310139","unternehmen_1573565543483"],"width":50}]},{"order":2,"cells":[{"order":0,"fields":["hr_1582557416361"],"width":"100"}]},{"order":3,"cells":[{"order":0,"fields":["html_1573643938225"],"width":"100"}]},{"order":4,"cells":[{"order":0,"fields":["datenschutz_1573564502005"],"width":"100"}]},{"order":5,"cells":[{"order":0,"fields":["hr_1582557452605"],"width":"100"}]},{"order":6,"cells":[{"order":0,"fields":["html_1581505991324"],"width":"100"}]},{"order":7,"cells":[{"order":0,"fields":["hidden_sprache_1573570858706"],"width":"100"}]},{"order":8,"cells":[{"order":0,"fields":["hidden_optinherkunft_1573569958584"],"width":"100"}]},{"order":9,"cells":[{"order":0,"fields":["yuuvis-produkt_1582728684947"],"width":"100"}]},{"order":10,"cells":[{"order":0,"fields":["page-url_1574859765696"],"width":"100"}]},{"order":11,"cells":[{"order":0,"fields":["jetzt_kontakt_aufnehmen_1574694425581"],"width":"100"}]}],"order":2,"type":"part","clean":false,"title":"2","key":"rghcnnwx","editActive":true,"drawerDisabled":false}],"drawerDisabled":false,"manual_key":"1","container_styles_border":"","container_styles_height":"","container_styles_margin":"","container_styles_padding":"","container_styles_float":"","container_styles_show_advanced_css":"0","title_styles_border":"","title_styles_height":"","title_styles_width":"","title_styles_font-size":"","title_styles_margin":"","title_styles_padding":"","title_styles_float":"","title_styles_show_advanced_css":"0","row_styles_border":"","row_styles_width":"","row_styles_font-size":"","row_styles_margin":"","row_styles_padding":"","row_styles_show_advanced_css":"0","row-odd_styles_border":"","row-odd_styles_height":"","row-odd_styles_width":"","row-odd_styles_font-size":"","row-odd_styles_margin":"","row-odd_styles_padding":"","row-odd_styles_show_advanced_css":"0","success-msg_styles_border":"","success-msg_styles_height":"","success-msg_styles_width":"","success-msg_styles_font-size":"","success-msg_styles_margin":"","success-msg_styles_padding":"","success-msg_styles_show_advanced_css":"0","error_msg_styles_border":"","error_msg_styles_width":"","error_msg_styles_font-size":"","error_msg_styles_margin":"","error_msg_styles_padding":"","error_msg_styles_show_advanced_css":"0","conditions":[],"mp_breadcrumb":"0","mp_progress_bar":"1","mp_display_titles":"0","mp_prev_label":"Zur\u00fcck","mp_next_label":"Weiter","breadcrumb_container_styles_border":"","breadcrumb_container_styles_height":"","breadcrumb_container_styles_width":"","breadcrumb_container_styles_margin":"","breadcrumb_container_styles_padding":"","breadcrumb_container_styles_float":"","breadcrumb_container_styles_show_advanced_css":"0","breadcrumb_buttons_styles_border":"","breadcrumb_buttons_styles_height":"","breadcrumb_buttons_styles_width":"","breadcrumb_buttons_styles_font-size":"","breadcrumb_buttons_styles_margin":"","breadcrumb_buttons_styles_padding":"","breadcrumb_buttons_styles_float":"","breadcrumb_buttons_styles_show_advanced_css":"0","breadcrumb_button_hover_styles_border":"","breadcrumb_button_hover_styles_height":"","breadcrumb_button_hover_styles_width":"","breadcrumb_button_hover_styles_font-size":"","breadcrumb_button_hover_styles_margin":"","breadcrumb_button_hover_styles_padding":"","breadcrumb_button_hover_styles_float":"","breadcrumb_button_hover_styles_show_advanced_css":"0","breadcrumb_active_button_styles_border":"","breadcrumb_active_button_styles_height":"","breadcrumb_active_button_styles_width":"","breadcrumb_active_button_styles_font-size":"","breadcrumb_active_button_styles_margin":"","breadcrumb_active_button_styles_padding":"","breadcrumb_active_button_styles_float":"","breadcrumb_active_button_styles_show_advanced_css":"0","progress_bar_container_styles_border":"","progress_bar_container_styles_height":"","progress_bar_container_styles_width":"","progress_bar_container_styles_margin":"","progress_bar_container_styles_padding":"","progress_bar_container_styles_float":"","progress_bar_container_styles_show_advanced_css":"0","progress_bar_fill_styles_border":"","progress_bar_fill_styles_height":"","progress_bar_fill_styles_margin":"","progress_bar_fill_styles_padding":"","progress_bar_fill_styles_show_advanced_css":"0","part_titles_styles_border":"","part_titles_styles_height":"","part_titles_styles_width":"","part_titles_styles_font-size":"","part_titles_styles_margin":"","part_titles_styles_padding":"","part_titles_styles_float":"","part_titles_styles_show_advanced_css":"0","navigation_container_styles_border":"","navigation_container_styles_height":"","navigation_container_styles_width":"","navigation_container_styles_margin":"","navigation_container_styles_padding":"","navigation_container_styles_float":"","navigation_container_styles_show_advanced_css":"0","previous_button_styles_border":"","previous_button_styles_height":"","previous_button_styles_width":"","previous_button_styles_font-size":"","previous_button_styles_margin":"","previous_button_styles_padding":"","previous_button_styles_show_advanced_css":"0","next_button_styles_border":"","next_button_styles_height":"","next_button_styles_width":"","next_button_styles_font-size":"","next_button_styles_margin":"","next_button_styles_padding":"","next_button_styles_show_advanced_css":"0","navigation_hover_styles_border":"","navigation_hover_styles_height":"","navigation_hover_styles_width":"","navigation_hover_styles_font-size":"","navigation_hover_styles_margin":"","navigation_hover_styles_padding":"","navigation_hover_styles_show_advanced_css":"0","mp_validate":"1","allow_public_link":0,"embed_form":"","ninjaForms":"Ninja Forms","fieldTextareaRTEInsertLink":"Link einf\u00fcgen","fieldTextareaRTEInsertMedia":"Medien einf\u00fcgen","fieldTextareaRTESelectAFile":"Eine Datei ausw\u00e4hlen","formHoneypot":"Falls du ein Mensch bist und dieses Feld siehst, dann lasse es bitte leer.","fileUploadOldCodeFileUploadInProgress":"Datei-Upload l\u00e4uft.","fileUploadOldCodeFileUpload":"DATEI-UPLOAD","currencySymbol":"€","thousands_sep":".","decimal_point":",","siteLocale":"de_DE","dateFormat":"m\/d\/Y","startOfWeek":"1","of":"von","previousMonth":"Vorheriger Monat","nextMonth":"N\u00e4chster Monat","months":["Januar","Februar","M\u00e4rz","April","Mai","Juni","Juli","August","September","Oktober","November","Dezember"],"monthsShort":["Jan","Feb","Mrz","Apr","Mai","Jun","Jul","Aug","Sep","Okt","Nov","Dez"],"weekdays":["Sonntag","Montag","Dienstag","Mittwoch","Donnerstag","Freitag","Samstag"],"weekdaysShort":["So","Mo","Die","Mi","Do","Fr","Sa"],"weekdaysMin":["So","Mo","Di","Mi","Do","Fr","Sa"],"currency_symbol":"","beforeForm":"","beforeFields":"","afterFields":"","afterForm":""};form.fields=[{"objectType":"Field","objectDomain":"fields","editActive":false,"order":1,"label":"Bitte Anrede ausw\u00e4hlen:","type":"listradio","key":"bitte_anrede_auswaehlen_1580465134673","label_pos":"above","required":1,"container_class":"","element_class":"","admin_label":"","help_text":"","drawerDisabled":"","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"list_item_row_styles_show_advanced_css":0,"list_item_label_styles_show_advanced_css":0,"list_item_element_styles_show_advanced_css":0,"cellcid":"c3751","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","list_item_row_styles_border":"","list_item_row_styles_height":"","list_item_row_styles_width":"","list_item_row_styles_font-size":"","list_item_row_styles_margin":"","list_item_row_styles_padding":"","list_item_row_styles_float":"","list_item_label_styles_border":"","list_item_label_styles_height":"","list_item_label_styles_width":"","list_item_label_styles_font-size":"","list_item_label_styles_margin":"","list_item_label_styles_padding":"","list_item_label_styles_float":"","list_item_element_styles_border":"","list_item_element_styles_height":"","list_item_element_styles_width":"","list_item_element_styles_font-size":"","list_item_element_styles_margin":"","list_item_element_styles_padding":"","list_item_element_styles_float":"","field_label":"Bitte Anrede ausw\u00e4hlen:","field_key":"bitte_anrede_auswaehlen_1580465134673","options":[{"errors":[],"max_options":"0","label":"Sehr geehrte Frau","value":"1","calc":"1","selected":"0","order":"0","settingModel":{"settings":false,"hide_merge_tags":false,"error":false,"name":"options","type":"option-repeater","label":"Options <a href=\"#\" class=\"nf-add-new\">Add New<\/a> <a href=\"#\" class=\"extra nf-open-import-tooltip\"><i class=\"fa fa-sign-in\" aria-hidden=\"true\"><\/i> Import<\/a>","width":"full","group":"","value":[{"label":"One","value":"one","calc":"","selected":0,"order":0},{"label":"Two","value":"two","calc":"","selected":0,"order":1},{"label":"Three","value":"three","calc":"","selected":0,"order":2}],"columns":{"label":{"header":"Label","default":""},"value":{"header":"Value","default":""},"calc":{"header":"Calc Value","default":""},"selected":{"header":"<span class=\"dashicons dashicons-yes\"><\/span>","default":0}}},"manual_value":"1"},{"errors":[],"max_options":"0","label":"Sehr geehrter Herr","value":"2","calc":"2","selected":"0","order":"1","settingModel":{"settings":false,"hide_merge_tags":false,"error":false,"name":"options","type":"option-repeater","label":"Options <a href=\"#\" class=\"nf-add-new\">Add New<\/a> <a href=\"#\" class=\"extra nf-open-import-tooltip\"><i class=\"fa fa-sign-in\" aria-hidden=\"true\"><\/i> Import<\/a>","width":"full","group":"","value":[{"label":"One","value":"one","calc":"","selected":0,"order":0},{"label":"Two","value":"two","calc":"","selected":0,"order":1},{"label":"Three","value":"three","calc":"","selected":0,"order":2}],"columns":{"label":{"header":"Label","default":""},"value":{"header":"Value","default":""},"calc":{"header":"Calc Value","default":""},"selected":{"header":"<span class=\"dashicons dashicons-yes\"><\/span>","default":0}}},"manual_value":"1"},{"errors":[],"max_options":"0","label":"Guten Tag","value":"3","calc":"3","selected":"0","order":"2","settingModel":{"settings":false,"hide_merge_tags":false,"error":false,"name":"options","type":"option-repeater","label":"Options <a href=\"#\" class=\"nf-add-new\">Add New<\/a> <a href=\"#\" class=\"extra nf-open-import-tooltip\"><i class=\"fa fa-sign-in\" aria-hidden=\"true\"><\/i> Import<\/a>","width":"full","group":"","value":[{"label":"One","value":"one","calc":"","selected":0,"order":0},{"label":"Two","value":"two","calc":"","selected":0,"order":1},{"label":"Three","value":"three","calc":"","selected":0,"order":2}],"columns":{"label":{"header":"Label","default":""},"value":{"header":"Value","default":""},"calc":{"header":"Calc Value","default":""},"selected":{"header":"<span class=\"dashicons dashicons-yes\"><\/span>","default":0}}},"manual_value":"1"}],"id":747,"beforeField":"","afterField":"","value":"","parentType":"list","element_templates":["listradio","input"],"old_classname":"list-radio","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":2,"label":"Vorname","type":"firstname","key":"firstname_1573210589193","label_pos":"hidden","required":0,"default":"","placeholder":"Vorname","container_class":"","element_class":"","admin_label":"","help_text":"","custom_name_attribute":"fname","personally_identifiable":1,"value":"","drawerDisabled":"","field_label":"Vorname","field_key":"firstname_1573210589193","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3755","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","cellOrder":1,"id":748,"beforeField":"","afterField":"","parentType":"firstname","element_templates":["firstname","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":4,"label":"Nachname","type":"lastname","key":"lastname_1573210591744","label_pos":"hidden","required":1,"default":"","placeholder":"Nachname*","container_class":"","element_class":"","admin_label":"","help_text":"","custom_name_attribute":"lname","personally_identifiable":1,"value":"","drawerDisabled":"","field_label":"Nachname","field_key":"lastname_1573210591744","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3757","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","cellOrder":1,"id":749,"beforeField":"","afterField":"","parentType":"lastname","element_templates":["lastname","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":8,"label":"Email","type":"email","key":"email_1582725914409","label_pos":"hidden","required":1,"default":"","placeholder":"E-Mail-Adresse*","container_class":"","element_class":"","admin_label":"","help_text":"<p>Die E-Mailadresse ist ein Pflichtfeld.<br><\/p>","custom_name_attribute":"email","personally_identifiable":1,"value":"","drawerDisabled":"","field_label":"Email","field_key":"email_1582725914409","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3760","oldCellcid":"c4390","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","id":750,"beforeField":"","afterField":"","parentType":"email","element_templates":["email","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":9,"label":"Ihre Nachricht an uns","type":"textarea","key":"ihre_nachricht_an_uns_1573565913431","label_pos":"hidden","required":"","default":"","placeholder":"Ihre Nachricht an uns","container_class":"","element_class":"","input_limit":160,"input_limit_type":"characters","input_limit_msg":"Zeichen \u00fcbrig","manual_key":"","admin_label":"","help_text":"","textarea_rte":"","disable_rte_mobile":"","textarea_media":"","value":"","drawerDisabled":"","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3763","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","field_label":"Ihre Nachricht an uns","field_key":"ihre_nachricht_an_uns_1573565913431","id":751,"beforeField":"","afterField":"","parentType":"textarea","element_templates":["textarea","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":5,"label":"Divider","type":"hr","container_class":"","element_class":"","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"key":"hr_1581506117361","cellcid":"c3766","oldCellcid":"c3772","field_label":"Divider","field_key":"hr_1581506117361","id":764,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":6,"label":"HTML","type":"html","default":"<p><b>Erleben Sie yuuvis\u00ae live!<\/b><\/p>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1581506146896","cellcid":"c3769","drawerDisabled":"","oldCellcid":"c3775","field_label":"HTML","field_key":"html_1581506146896","id":768,"beforeField":"","afterField":"","value":"<p><b>Erleben Sie yuuvis\u00ae live!<\/b><\/p>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":7,"label":"Die Einf\u00fchrung der Software-L\u00f6sung ist geplant.","type":"checkbox","key":"softwaresolution_1581506167859","label_pos":"right","required":"","container_class":"","element_class":"","manual_key":1,"admin_label":"","help_text":"","default_value":"unchecked","checked_value":1,"unchecked_value":2,"checked_calc_value":"","unchecked_calc_value":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3772","drawerDisabled":"","oldCellcid":"c3778","field_label":"Die Einf\u00fchrung der Software-L\u00f6sung ist geplant.","field_key":"softwaresolution_1581506167859","id":765,"beforeField":"","afterField":"","value":"","parentType":"checkbox","element_templates":["checkbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":8,"label":"Ich bin an einer Software-Pr\u00e4sentation interessiert.","type":"checkbox","key":"softwarepresentation_1581506142599","label_pos":"right","required":"","container_class":"","element_class":"","manual_key":1,"admin_label":"","help_text":"","default_value":"unchecked","checked_value":1,"unchecked_value":2,"checked_calc_value":"","unchecked_calc_value":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3775","drawerDisabled":"","oldCellcid":"c3781","field_label":"Ich bin an einer Software-Pr\u00e4sentation interessiert.","field_key":"softwarepresentation_1581506142599","id":766,"beforeField":"","afterField":"","value":"","parentType":"checkbox","element_templates":["checkbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":9,"label":"Trennung","type":"hr","container_class":"","element_class":"","key":"hr_1573210614298","field_label":"Trennung","field_key":"hr_1573210614298","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3778","oldCellcid":"c4424","id":755,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":10,"label":"HTML","type":"html","default":"<small>Felder mit * sind Pflichtfelder.<\/small>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1574181495044","cellcid":"c3781","drawerDisabled":"","field_label":"HTML","field_key":"html_1574181495044","oldCellcid":"c3787","id":756,"beforeField":"","afterField":"","value":"<small>Felder mit * sind Pflichtfelder.<\/small>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":1,"label":"Telefon","type":"phone","cellcid":"c3787","key":"telefon_1582726279559","label_pos":"hidden","required":"","default":"","placeholder":"Telefon (Durchwahl)","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":"","admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"phone","personally_identifiable":1,"wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"value":"","cellOrder":1,"drawerDisabled":"","oldCellcid":"c3800","field_label":"Telefon","field_key":"telefon_1582726279559","id":761,"beforeField":"","afterField":"","parentType":"textbox","element_templates":["tel","textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":2,"label":"Position","type":"textbox","cellcid":"c3787","key":"position_1580818819203","label_pos":"hidden","required":1,"default":"","placeholder":"Position*","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":"","admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"","personally_identifiable":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"value":"","cellOrder":2,"drawerDisabled":"","oldCellcid":"c6784","field_label":"Position","field_key":"position_1580818819203","id":763,"beforeField":"","afterField":"","parentType":"textbox","element_templates":["textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":3,"label":"ZIP Code","type":"zip","cellcid":"c3789","key":"zip_code_1582726310139","label_pos":"hidden","required":"","default":"","placeholder":"PLZ","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Character(s) left","manual_key":"","admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"zip","personally_identifiable":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"value":"","cellOrder":1,"drawerDisabled":"","oldCellcid":"c3802","field_label":"ZIP Code","field_key":"zip_code_1582726310139","id":762,"beforeField":"","afterField":"","parentType":"zip","element_templates":["zip","textbox","input","textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":4,"label":"Unternehmen","type":"address","key":"unternehmen_1573565543483","label_pos":"hidden","required":1,"default":"","placeholder":"Unternehmen*","container_class":"","element_class":"","input_limit":"","input_limit_type":"characters","input_limit_msg":"Zeichen \u00c3\u00bcbrig","manual_key":"","admin_label":"","help_text":"","mask":"","custom_mask":"","custom_name_attribute":"company","personally_identifiable":1,"value":"","drawerDisabled":"","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3789","oldCellcid":"c4400","cellOrder":2,"field_label":"Unternehmen","field_key":"unternehmen_1573565543483","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","id":752,"beforeField":"","afterField":"","parentType":"address","element_templates":["address","textbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":5,"type":"hr","label":"Divider","container_class":"","element_class":"","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"key":"hr_1582557416361","cellcid":"c3792","field_label":"Divider","field_key":"hr_1582557416361","id":769,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":6,"label":"HTML","type":"html","default":"<p>Bitte lesen und best\u00e4tigen Sie unsere Datenschutzbestimmungen.<\/p>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1573643938225","cellcid":"c3795","drawerDisabled":"","field_label":"HTML","field_key":"html_1573643938225","oldCellcid":"c3828","id":753,"beforeField":"","afterField":"","value":"<p>Bitte lesen und best\u00e4tigen Sie unsere Datenschutzbestimmungen.<\/p>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":7,"label":"Ja, ich habe die <a href=\"\/de\/impressum-und-datenschutz\" target=\"_blank\" rel=\"noopener\">Datenschutzbestimmungen<\/a> gelesen und bin damit einverstanden.","type":"checkbox","key":"datenschutz_1573564502005","label_pos":"right","required":1,"container_class":"","element_class":"","manual_key":1,"admin_label":"DATENSCHUTZ","help_text":"<p><br><\/p>","default_value":"unchecked","checked_value":1,"unchecked_value":2,"checked_calc_value":"","unchecked_calc_value":"","drawerDisabled":"","field_label":"Ja, ich habe die <a href=\"\/de\/impressum-und-datenschutz\" target=\"_blank\" rel=\"noopener\">Datenschutzbestimmungen<\/a> gelesen und bin damit einverstanden.","field_key":"datenschutz_1573564502005","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3798","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","oldCellcid":"c4421","default":"unchecked","value":"unchecked","id":754,"beforeField":"","afterField":"","parentType":"checkbox","element_templates":["checkbox","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":8,"label":"Divider","type":"hr","container_class":"","element_class":"","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"key":"hr_1582557452605","cellcid":"c3801","field_label":"Divider","field_key":"hr_1582557452605","id":770,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"hr","element_templates":["hr","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":9,"label":"HTML Copy","type":"html","default":"<small>Felder mit * sind Pflichtfelder.<\/small>","container_class":"","element_class":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"key":"html_1581505991324","cellcid":"c3804","drawerDisabled":"","field_label":"HTML Copy","field_key":"html_1581505991324","oldCellcid":"c4735","id":767,"beforeField":"","afterField":"","value":"<small>Felder mit * sind Pflichtfelder.<\/small>","label_pos":"above","parentType":"html","element_templates":["html","input"],"old_classname":"","wrap_template":"wrap"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":10,"label":"SPRACHE","type":"hidden","key":"hidden_sprache_1573570858706","default":"DE","admin_label":"sprache","manual_key":1,"drawerDisabled":"","wrap_styles_show_advanced_css":0,"label_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"cellcid":"c3807","oldCellcid":"c4430","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","field_label":"SPRACHE","field_key":"hidden_sprache_1573570858706","id":757,"beforeField":"","afterField":"","value":"DE","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":11,"label":"Optin-Herkunft","type":"hidden","key":"hidden_optinherkunft_1573569958584","default":"{wp:site_url}","admin_label":"optin-herkunft","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3810","drawerDisabled":"","manual_key":1,"field_label":"Optin-Herkunft","field_key":"hidden_optinherkunft_1573569958584","oldCellcid":"c3843","id":758,"beforeField":"","afterField":"","value":"https:\/\/bp387lpw.myraidbox.de\/de\/","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":12,"label":"YUUVIS-PRODUKT","type":"hidden","key":"yuuvis-produkt_1582728684947","default":"RAD","admin_label":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3813","drawerDisabled":false,"field_label":"YUUVIS-PRODUKT","field_key":"yuuvis-produkt_1582728684947","id":771,"beforeField":"","afterField":"","value":"RAD","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":13,"label":"PAGE-URL","type":"hidden","key":"page-url_1574859765696","default":"{wp:post_url}","admin_label":"","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","wrap_styles_show_advanced_css":0,"label_styles_border":"","label_styles_width":"","label_styles_font-size":"","label_styles_margin":"","label_styles_padding":"","label_styles_float":"","label_styles_show_advanced_css":0,"element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","element_styles_show_advanced_css":0,"cellcid":"c3816","drawerDisabled":"","field_label":"PAGE-URL","field_key":"page-url_1574859765696","oldCellcid":"c3849","id":759,"beforeField":"","afterField":"","value":"https:\/\/bp387lpw.myraidbox.de\/de\/rapid-application-development\/","label_pos":"above","parentType":"hidden","element_templates":["hidden","input"],"old_classname":"","wrap_template":"wrap-no-label"},{"objectType":"Field","objectDomain":"fields","editActive":false,"order":14,"label":"Jetzt Kontakt aufnehmen","type":"submit","processing_label":"Daten werden \u00fcbermittelt ...","container_class":"","element_class":"form-send-button","key":"jetzt_kontakt_aufnehmen_1574694425581","drawerDisabled":"","field_label":"Jetzt Kontakt aufnehmen","field_key":"jetzt_kontakt_aufnehmen_1574694425581","wrap_styles_show_advanced_css":0,"element_styles_show_advanced_css":0,"submit_element_hover_styles_show_advanced_css":0,"cellcid":"c3819","wrap_styles_border":"","wrap_styles_width":"","wrap_styles_margin":"","wrap_styles_padding":"","wrap_styles_float":"","element_styles_border":"","element_styles_width":"","element_styles_font-size":"","element_styles_margin":"","element_styles_padding":"","element_styles_float":"","submit_element_hover_styles_border":"","submit_element_hover_styles_width":"","submit_element_hover_styles_font-size":"","submit_element_hover_styles_margin":"","submit_element_hover_styles_padding":"","submit_element_hover_styles_float":"","oldCellcid":"c4433","element_styles_background-color":"","element_styles_border-color":"","element_styles_color":"","submit_element_hover_styles_background-color":"","submit_element_hover_styles_border-color":"","submit_element_hover_styles_color":"","id":760,"beforeField":"","afterField":"","value":"","label_pos":"above","parentType":"textbox","element_templates":["submit","button","input"],"old_classname":"","wrap_template":"wrap-no-label"}];nfForms.push(form); <div class="nf-mp-header"></div> <div class="nf-mp-body"></div> <div class="nf-mp-footer"></div> {{{ data.renderProgressBar() }}} {{{ data.renderBreadcrumbs() }}} {{{ data.renderPartTitle() }}} <h3> {{{ data.title }}} </h3> {{{ data.renderNextPrevious() }}} <ul class="nf-next-previous"> <# if ( data.showPrevious ) { #> <li class="nf-previous-item"> <input type="button" class="nf-previous" value="{{{ data.prevLabel }}}" /> </li> <# } #> <# if ( data.showNext ) { #> <li class="nf-next-item"> <input type="button" class="nf-next" value="{{{ data.nextLabel }}}" /> </li> <# } #> </ul> <ul class="nf-breadcrumbs"> <# _.each( data.parts, function( part, index ) { #> <li class="{{{ ( data.currentIndex == index ) ? 'active' : '' }}} {{{ ( part.errors ) ? 'errors' : '' }}}"> <a href="#" class="nf-breadcrumb" data-index="{{{ index }}}">{{{ ( part.errors ) ? '' : '' }}} {{{ part.title }}}</a> </li> <# } ); #> </ul> <div class="nf-progress-container"> <div class="nf-progress" style="width: {{{ data.percent }}}%;"></div> </div> <nf-fields></nf-fields> <nf-cells></nf-cells> Q&A
  
  Hier wird die Q&A Seite direkt geladen und nicht in einem neuen TAB
Visit annotations in context

Annotators

Sven.Hoffmann.OS

URL

bp387lpw.myraidbox.de/de/rapid-application-development/
www.theguardian.com www.theguardian.com

Ho99o9 review – power and panic from pogoing punk-rap firestarters

1
1. MilesButters 02 Mar 2020
  
  in Public
  
  Tag-teaming vocalists Eaddy and theOGM don’t emerge in wedding dresses, smashing all equipment in sight like they did on 2015’s Warped Tour, causing their removal.
  
  This is a prepostion its the how it tell how the arrived in 2015
Visit annotations in context

Annotators

MilesButters

URL

theguardian.com/music/2019/apr/30/ho99o9-review-the-garage-london
Feb 2020
www.makeuseof.com www.makeuseof.com

How (and Why) to Disable Algorithmic Feeds on Twitter, Instagram, and Facebook

1
1. chrisaldrich 26 Feb 2020
  
  in Public
  
  The biggest drawback of algorithmic feeds is that you might be looking at irrelevant content. When you see something on your timeline and want to comment, you will have to check the timestamp to see if your comment is still relevant or not.
  
  context collapse algorithmic feeds
Visit annotations in context

Tags

algorithmic feeds

context collapse

Annotators

chrisaldrich

URL

makeuseof.com/tag/how-and-why-to-disable-algorithmic-feeds-on-twitter-instagram-and-facebook/
clas3304.wordpress.com clas3304.wordpress.com

Augustan Rome

1
1. markbassner 25 Feb 2020
  
  in Public
  
  Use google image search or the website site used last class to find illustrations to pair with a THREE quotes from the book. Briefly explain your choices. Categorize: Reading Response; tag your post: class10
  
  I totally forgot how to use this
Visit annotations in context

Annotators

markbassner

URL

clas3304.wordpress.com/readings/unit-two/
www.scienceintheclassroom.org www.scienceintheclassroom.org

Drivers of Daily Routines in an Ectothermic Marine Predator: Hunt Warm, Rest Warmer? | Science in the Classroom

1
1. hy323 24 Feb 2020
  
  in Public
  
  V16, V13 and V9 types
  
  The numbers refer to the diameter of the tag, with different sizes being better suited to a range of animal sizes and data types.
  
  Glossary
Visit annotations in context

Tags

Glossary

Annotators

hy323

URL

scienceintheclassroom.org/research-papers/drivers-daily-routines-ectothermic-marine-predator-hunt-warm-rest-warmer
www.rabbitmq.com www.rabbitmq.com

RabbitMQ - Consumer Acknowledgements and Publisher Confirms

1
1. NathanielHolden 20 Feb 2020
  
  in Public
  
  When a consumer (subscription) is registered, messages will be delivered (pushed) by RabbitMQ using the basic.deliver method. The method carries a delivery tag, which uniquely identifies the delivery on a channel.
  
  Messages are delivered with Unique Identifiers as delivery tag
Visit annotations in context

Annotators

NathanielHolden

URL

rabbitmq.com/confirms.html
www.nxp.com www.nxp.com

NXP UCODE 8

1
1. bernadette.konkel 18 Feb 2020
  
  in Public
  
  From Rain RFID Tag IC point of view it is therefore of highest importance to ensure correctness of these data.
  
  **Delete
Visit annotations in context

Annotators

bernadette.konkel

URL

nxp.com/docs/en/fact-sheet/UCODE8LF.pdf
www.businessnewsdaily.com www.businessnewsdaily.com

Entrepreneurship Defined: What It Means to Be an Entrepreneur

1
1. Mackiwg 17 Feb 2020
  
  in Public
  
  Entrepreneurship Defined
  
  TIP for OERu LEARNERS: Remember to add the course code tag in the tag field when annotating with Hypothes.is. Eg BMAN111 or IENT101. (Hit the enter key after typing the tag.)
  
  bman111 IENT101
Visit annotations in context

Tags

bman111

IENT101

Annotators

Mackiwg

URL

businessnewsdaily.com/7275-entrepreneurship-defined.html
www.4cid.org www.4cid.org

4CID.org - About 4C/ID

1
1. choppa1890 13 Feb 2020
  
  in Public
  
  four-component instructional design (4C/ID)
  
  4C/ID tag might be one of the models I want to look into as I might use it for the Art Modules Class
  
  4C/ID Reading Check 3
Visit annotations in context

Tags

Reading Check 3

4C/ID

Annotators

choppa1890

URL

4cid.org/about-4cid
www.klusster.com www.klusster.com

Top 10 Important Online Marketing Tools for Entrepreneurs | Shiv

1
1. everydesigns 13 Feb 2020
  
  in Public
  
  Top 10 Important Online Marketing Tools for Entrepreneurs 4.0 Thank you for your rating! Why not leave a message? Any additional feedback or questions are welcome below. Your Name Your Email Address Phone Number Message the Author Cancel By Shiv | Contact Author less than a minute ago Marketing tools are like a helping hand for many entrepreneurs. As time is money using online marketing tools make your job easy and fast and also saves money. Marketing tools are like a helping hand that makes your job so easy. These tools are designed in such a way that it saves you time, effort and money. Apart from finding customers, one of the most difficult problems faced by entrepreneurs is to save time and manage cash flow. Using tools for marketing your business ensures you meet your goal on time. Here are the top 10 most important online marketing tools for entrepreneurs- Hubspot- Inbound Marketing Tool If you want to buy one tool to grow your business exponentially, this is the tool you should buy. Hubspot offers an inbound marketing tool that promises you with attracting leads, converting them and closing the deal. This tool acts as a funnel for your business that narrows downs your audience into your customers and finally generates revenue out of them. You can get started using the Hubspot tool with just $113 per month. You can also get online tutorials on how to use Hubspot tools online or you can always call or email their support team to help you out. Freelancing Graphic Design Platform by EveryDesigns Marketing is impossible without good designs. In marketing, we have a lot of promotional content that uses heavy design work to attract customers. We, humans, are visual animals and it is easier to influence us visually rather than any other means. You can use the EveryDesigns platform to get designs for your business. It is a one-stop destination for your business to get any graphic design services. The innovation that they offer in their services is by organizing a contest for your designs where multiple designers work from around the world. You choose the best design submitted by designers and pay for only that design. You get multiple design options to choose from, along with a 100% money-back guarantee. MailChimp- Email Marketing MailChimp is the best tool when it comes to email marketing. As an entrepreneur, you know how important email marketing is. If you strategize your email marketing outreach, you can attract very valuable customers to your business website. The best part is that it provides some of its features for free, which is enough for a startup or small business to get started and if you want to buy the paid version, then it is as low as $10 per month. Zoom- Conference Meeting Tool Zoom is a web as well as an app-based conference meeting organization tool that helps you in connecting with different team members throughout the world. You can also use this tool to organize a webinar and post it on youtube by recording it. You can pretty much do anything, it is a better version of skype where you can whiteboard as well. The basic version can be used for free and the paid version with pro features starets at $20 for each member. Klusster- Publication Community Kluster is a tool used by digital marketers to spread content and attract customers to their website. Kluster offers a platform to publish content written by you and promote business. As an entrepreneur, you should understand the importance of the backlink for your website. Klusster is one such platform providing quality publications for businesses. The best part it's free. Grammarly-Content Optimization Tool As a marketer or entrepreneur, you know the power of content marketing. It is the most powerful way of branding your business. Creating content can be tough but, the most important thing is creating grammatically correct content. That’s where Grammarly comes and saves the day. It automatically suggests your corrections for every mistake you made. The basic version is free and the paid version with advanced features is $30. You can really see the difference in your writing after using Grammarly. Zoho Social-Social Media Marketing Tool Social media is a platform that has numerous potential and if you are not using social media then you are missing out on a huge opportunity. There are around 4.3 billion social media users and the primary rule of business is where there are people, there is business. Zoho Social is a social media management tool that helps you in promoting your business on multiple social media channels. You can use different channels like Facebook, Twitter Instagram, etc. and post your content, images using one app. UberSuggest- Digital Marketing Tool UberSuggest is a free digital marketing tool provided by Neil Patel Digital. This tool is best for a digital marketer who does not want to spend a big amount of money on buying digital marketing tools. This tool is very easy to use and offers many features that are enough for you to get started with your website’s SEO. Yoast SEO- WordPress Plugin for SEO Yoast SEO is a plugin that you can install on your WordPress site. This tool is a well-researched tool that recommends you SEO corrections that you should make in order to rank your website for specific keywords. It is the best on-page SEO management tool and comes with a one-time price tag of $89 for one website. You can also subscribe to their service starting from $19 per month where you get all the premium features with multiple plugins in one. If you do not want to buy Yoast SEO Plugin then you can also use the free version of the plugin but it is recommended that you should use the premium version if you want to stay ahead off of your competitors. Google- Search Engine & Other Tools You heard it right. Google is a tool that everyone uses and earlier in this post, I’ve said where there are people there is business, therefore, you should also use it. Google's market share when it comes to search engines is 80-90%. As an entrepreneur, you should aim for ranking your website higher in the high volume search keywords. To do that, you would need your website to be optimized according to the Google guidelines and recommendations. Google Offers multiple tools like search console, analytics, tag manager, etc to manage and optimize your website. You should use these tools to make your website better. Conclusion You can use these tools to have leverage over your competitors and quickly increase sales for your business. You are an entrepreneur and time is money for you, therefore, you should use these tools to save time and money. Comment below, what tool are you using to manage your business along with pricing and if you are using any of the above-mentioned tools, tell us your experience. Tags marketing tools / List of best marketing tools
  
  Marketing tools are like a helping hand for many entrepreneurs. As time is money using online marketing tools make your job easy and fast and also saves money. Top 10 Important Online Marketing Tools for Entrepreneurs
  
  MARKETING TOOLS / LIST OF BEST MARKETING TOOLS
Visit annotations in context

Tags

MARKETING TOOLS / LIST OF BEST MARKETING TOOLS

Annotators

everydesigns

URL

klusster.com/portfolios/shiv/contents/16672
omeka.org omeka.org

Items - Omeka Classic User Manual

1
1. cservin 13 Feb 2020
  
  in Public
  
  Add Tags” button
  
  Important in order to add the tag.
Visit annotations in context

Annotators

cservin

URL

omeka.org/classic/docs/Content/Items/
www.zeit.de www.zeit.de

Google Maps: Google Maps mit 99 Smartphones hacken – wirklich?

1
1. martinlindner 12 Feb 2020
  
  in Public
  
  dieses Mal führte uns Google Maps über einen anderen Weg. Zu Beginn bemerkten wir es gar nicht, aber hier und dort fuhren wir plötzlich Landstraße, wo wir bisher immer auf die Autobahn gelenkt worden waren. Da es keine größeren Umwege waren, dachte ich mir nichts dabei und folgte stur den Anweisungen des Gerätes. Kurz vor dem Ziel wollten wir noch mal ein Stück Autobahn fahren, wichen von der vorgeschlagenen Route ab und steckten plötzlich fest – eine offenbar eben erst errichtete Straßenblockade einiger Bauern versperrte uns den Weg. Google Maps hielt kurz inne und schlug schnell eine Umfahrung vor. Dann hörte ich im Radio, dass französische Bauern schon den ganzen Tag über landesweit für höhere Lebensmittelpreise streikten, viele Auffahrten und Kreuzungen seien von den wütenden Bauern blockiert worden. Da erst wurde mir klar, dass Google uns den ganzen Tag um die Blockaden herumgelenkt hatte
  
  Staus waren Demos, Google ersparte den Kontakt mit einer schwierigen und unbequemen Wirklichkeit. Was zeigt das Beispiel wirklich? Wo ist hier das Problem? Was ist anders als sonst in der urban-technischen "realen Welt"?
Visit annotations in context

Annotators

martinlindner

URL

zeit.de/digital/internet/2020-02/google-maps-jubilaeum-digitalisierung-berliner-kuenstler/seite-2
mitpress.mit.edu mitpress.mit.edu

Synesthesia

1
1. heatherstaines 07 Feb 2020
  
  in Public
  
  In this volume in the MIT Press Essential Knowledge series, Richard Cytowic, the expert who returned synesthesia to mainstream science after decades of oblivion, offers a concise, accessible primer on this fascinating human experience.
  
  This would be a great gift! I add a tag for the event and a tag for the person the gift might be great for. Sometimes, when a site is dynamic (so I might not be able to get all the way back) I include an image and enough info to identify the gift.! Back to the 100K blog for another adventure?
  
  Xmas2019 Mom
Visit annotations in context

Tags

Mom

Xmas2019

Annotators

heatherstaines

URL

mitpress.mit.edu/books/synesthesia
sites.nd.edu sites.nd.edu

Wonders of the East – Medieval Studies Research Blog: Meet us at the Crossroads of Everything

2
1. carlybenusa 05 Feb 2020
  
  in Public
  
  uld argue the text is most guilty of exoticism as it reflects a sense of wonder about different groups of people without any reference to respective racial superiority or inferi
  
  guilty of exoticism - wonder, "otherness"
2. carlybenusa 05 Feb 2020
  
  in Public
  
  “the Anglo-Saxon readers and viewers of these texts probably considered them true”
  
  early english readers probably considered the wonders real
Visit annotations in context

Annotators

carlybenusa

URL

sites.nd.edu/manuscript-studies/tag/wonders-of-the-east/
www.salesforce.com www.salesforce.com

Untitled document

1
1. wangwen123 01 Feb 2020
  
  in Public
  
  CRM means greater efficiency for multiple teams. Automatically stored communication allows you to view emails, calendar, and phone call details in one easily accessible place. Add that to the ability for multiple teams to access the same information, and the amount of achievable progress simply skyrockets. Sales, marketing, and customer service teams can share valuable information about clients to continue to funnel them down the pipeline to get the desired result of closing a sale, knowledge of new products, or excellent customer service. Every department can now tag team to get the right information to the right individual. With this new found ease, teams can seamlessly work together to improve the bottom line.
  
  Good CRM can help people to focus on high value activities. And it is very important to avoid duplicating tasks among different groups.
Visit annotations in context

Annotators

wangwen123

URL

salesforce.com/hub/crm/benefits-of-crm/
Jan 2020
cgi.tu-harburg.de cgi.tu-harburg.de

Geo-Engineering - wirksamer Klimaschutz oder Größenwahn?

1
1. DolusMaximus 24 Jan 2020
  
  in Public
  
  m Bereich der Theorie.
  
  Das stimmt nicht ganz, alljährlich herrscht in Moskau bei der Parade über Nazi-Deutschland Sonnenschein, nur möglich mit sogenannter "Wolkenimpfung":
  
  Einen Tag vor der großen Militärparade regnet es kräftig in Moskau. Auch für Dienstag ist Schneeregen angekündigt. Doch voraussichtlich wird die Sonne scheinen. Wie geht das?
  
  https://www.dw.com/de/wolkenimpfung-sch%C3%B6nes-wetter-auf-bestellung/a-38751024
Visit annotations in context

Annotators

DolusMaximus

URL

cgi.tu-harburg.de/~pbbad/shared/Ginzky_et_al_2011_Geo-Engineering.pdf
www.newadvent.org www.newadvent.org

CHURCH FATHERS: De Principiis, Book I (Origen)

1
1. sheridantyson 23 Jan 2020
  
  in Public
  
  For whatever be the knowledge which we are able to obtain of God, either by perception or reflection, we must of necessity believe that He is by many degrees far better than what we perceive Him to be.
  
  tag: interesting
Visit annotations in context

Annotators

sheridantyson

URL

newadvent.org/fathers/04121.htm
www.freecodecamp.org www.freecodecamp.org

The Ultimate Linux Command Line Guide - Full Bash Tutorial

1
1. daaronr 23 Jan 2020
  
  in Public
  
  It’s worth noting that first line of the script starts with #!. It is a special directive which Unix treats differently.
  
  Term hash tag at top of bash scripts are NOT comments... they are important
  
  bash
Visit annotations in context

Tags

bash

Annotators

daaronr

URL

freecodecamp.org/news/linux-command-line-bash-tutorial/
www.blaetter.de www.blaetter.de

Soziale Kontrolle 4.0? Chinas Social Credit Systems | Blätter für deutsche und internationale Politik

1
1. Jim_Morrison 19 Jan 2020
  
  in Public
  
  spielerischer Überwachung keineswegs nur in Fernost, sondern auch im Westen angewandt werden, treffen Berichte über das chinesische SCS hierzulande meist auf großes Befremden.
  
  In der spielerischen Überwachung liegt eine der großen Gefahren für die westlich sozialisierte Welt. Da China durch seine autoritäre Regierungsform und das zensierte Internet eine Sonderrolle einnimmt, die für Europäer weit entfernt von ihrem Alltag erscheint, werden SCS für ebenfalls befremdlich und genauso weit entfernt gehalten.
  
  Dabei schleichen sich diese immer weiter ein. Neben den seit langem geläufigen Bonitätsprüfungen für Kreditnehmer versuchen viele Teile der digitalen Welt Nutzerverhalten durch Belohnungen zu steuern.
  
  Dies beginnt bei in der Freizeit verwendeten Diensten und Videospielen, die für tägliches Einloggen und Nutzen bzw. das Erledigen kleiner banaler Aufgaben einen Bonus vergeben - z.T. sogar für kumulierte Zeiträume, die dann die Notwendigkeit erzeugen, keinen Tag verpassen zu dürfen.
  
  Andererseits eröffnen sich ähnliche Systeme auch schon außerhalb der rein hobbymäßigen Tätigkeiten. Die meisten der Krankenkassen in Deutschland arbeiten bereits mit Scoring-Systemen (Bonus-Programmen), die einem bei ausreichender Teilnahme an Sportveranstaltungen und gesundheitlicher Vorsorge Prämien auszahlen.
  
  Die genannten Beispiele finden natürlich alle auf freiwilliger Basis statt und sind bisher ohne Negativ-Folgen bei Missachtung , produzieren trotzdem gleichzeit wieder Daten über das Nutzerverhalten. Sollten solche Systeme in Zukunft spontan in Pflichtprogramme verwandelt werden, kann sich am Beispiel der Krankenversicherung eine freiwillige Zusatzprämie schnell in eine nicht genehmigte Kassenleistung verwandeln, da die erforderlichen Punkte nicht erbracht wurden.
  
  Dass bei einigen Krankenversicherungen beispielsweise eine Zahnreinigung im Jahr kostenlos ist, bei anderen wiederum erst Punkte dafür gesammelt werden müssen, ist bereits jetzt schon Realität.
Visit annotations in context

Annotators

Jim_Morrison

URL

blaetter.de/ausgabe/2018/juli/soziale-kontrolle-40-chinas-social-credit-systems
Local file Local file

Untitled document

3
1. choppa1890 19 Jan 2020
  
  in Public
  
  Equivalency Theory
  
  another tag ?
2. choppa1890 19 Jan 2020
  
  in Public
  
  Theory of Interaction and Communication
  
  tag
  
  theory of interaction and communication
3. choppa1890 19 Jan 2020
  
  in Public
  
  Theory of Industrialization of Teaching
  
  another tag.
  
  theory of industrialization of teaching
Tags

theory of industrialization of teaching

theory of interaction and communication

Annotators

choppa1890
codewords.recurse.com codewords.recurse.com

Git from the inside out

1
1. gola520xxx 17 Jan 2020
  
  in Public
  
  tag of annotation type
  
  header
  
  must know
  
  confusing
  
  技術文章
Visit annotations in context

Tags

技術文章

Annotators

gola520xxx

URL

codewords.recurse.com/issues/two/git-from-the-inside-out
stackoverflow.com stackoverflow.com

`ffmpeg` can not dump RTSP streaming but `ffplay` can play it

1
1. TylerRick 16 Jan 2020
  
  in Public
  
  [mp4 @ 0x7fe98400dc00] track 1: could not find tag, codec not currently supported in container This means that pcm_mulaw is not allowed in the MP4 output container format. Your choices are: Use a different output container format ffmpeg -i input -codec copy output.mkv Or re-encode the audio ffmpeg -i input -c:v copy -c:a aac output.mp4
  
  ffmpeg recording IP camera recording stream
Visit annotations in context

Tags

recording IP camera

ffmpeg

recording stream

Annotators

TylerRick

URL

stackoverflow.com/questions/22345560/ffmpeg-can-not-dump-rtsp-streaming-but-ffplay-can-play-it
stackoverflow.com stackoverflow.com

Could not find tag for codec pcm_alaw in stream #1, codec not currently supported in container when concatenating 2 files using ffmpeg

1
1. TylerRick 16 Jan 2020
  
  in Public
  
  Encode the audio to a supported format, such as AAC by replacing -acodec copy with -c:a aac, or use a more flexible output container format that does support pcm_alaw, such as MOV or MKV.
  
  ffmpeg
Visit annotations in context

Tags

ffmpeg

Annotators

TylerRick

URL

stackoverflow.com/questions/47495713/could-not-find-tag-for-codec-pcm-alaw-in-stream-1-codec-not-currently-supporte
en.wikipedia.org en.wikipedia.org

Wikipedia:Articles for deletion/Katie Bouman - Wikipedia

1
1. hiperterminal 10 Jan 2020
  
  in Public
  
  it's an embarrassment to Wikipedia to have the AfD tag on top
  
  Article for Deletion
  
  Grounded theory
Visit annotations in context

Tags

Grounded theory

Annotators

hiperterminal

URL

en.wikipedia.org/wiki/Wikipedia:Articles_for_deletion/Katie_Bouman
www.econometrics-with-r.org www.econometrics-with-r.org

3.3 Hypothesis Tests Concerning the Population Mean | Introduction to Econometrics with R

1
1. Gfuschini 02 Jan 2020
  
  in Public
  
  p-value=PH0[|¯¯¯¯Y−μY,0|>|¯¯¯¯Yact−μY,0|](3.2)(3.2)p-value=PH0[|Y¯−μY,0|>|Y¯act−μY,0|]\begin{equation} p \text{-value} = P_{H_0}\left[ \lvert \overline{Y} - \mu_{Y,0} \rvert > \lvert \overline{Y}^{act} - \mu_{Y,0} \rvert \right] \tag{3.2} \end{equation} In (3.2), ¯¯¯¯YactY¯act\overline{Y}^{act} is the mean of the sample actually computed. Consequently, in order to compute the ppp-value as in (3.2), knowledge about the sampling distribution of ¯¯¯¯YY¯\overline{Y} when the null hypothesis is true is required. However in most cases the sampling distribution of ¯¯¯¯YY¯\overline{Y} is unknown. Fortunately, as stated by the CLT (see Key Concept 2.7), the large-sample approximation ¯¯¯¯Y≈N(μY,0,σ2¯¯¯¯Y) , σ2¯¯¯¯Y=σ2YnY¯≈N(μY,0,σY¯2) , σY¯2=σY2n \overline{Y} \approx \mathcal{N}(\mu_{Y,0}, \, \sigma^2_{\overline{Y}}) \ \ , \ \ \sigma^2_{\overline{Y}} = \frac{\sigma_Y^2}{n} can be made under the null. Thus, ¯¯¯¯Y−μY,0σY/√n∼N(0,1).Y¯−μY,0σY/n∼N(0,1). \frac{\overline{Y} - \mu_{Y,0}}{\sigma_Y/\sqrt{n}} \sim \mathcal{N}(0,1). So in large samples, the ppp-value can be computed without knowledge of the exact sampling distribution of ¯¯¯¯YY¯\overline{Y}. Calculating the p-Value when the Standard Deviation is Known For now, let us assume that σ¯¯¯¯YσY¯\sigma_{\overline{Y}} is known. Then, we can rewrite (3.2) as p-value=PH0⎡⎢⎣∣∣ ∣∣¯¯¯¯Y−μY,0σ¯¯¯¯Y∣∣ ∣∣>∣∣ ∣∣¯¯¯¯Yact−μY,0σ¯¯¯¯Y∣∣ ∣∣⎤⎥⎦=2⋅Φ⎡⎢⎣−∣∣ ∣∣¯¯¯¯Yact−μY,0σ¯¯¯¯Y∣∣ ∣∣⎤⎥⎦.(3.3)p-value=PH0[|Y¯−μY,0σY¯|>|Y¯act−μY,0σY¯|](3.3)=2⋅Φ[−|Y¯act−μY,0σY¯|].\begin{align} p \text{-value} =& \, P_{H_0}\left[ \left\lvert \frac{\overline{Y} - \mu_{Y,0}}{\sigma_{\overline{Y}}} \right\rvert > \left\lvert \frac{\overline{Y}^{act} - \mu_{Y,0}}{\sigma_{\overline{Y}}} \right\rvert \right] \\ =& \, 2 \cdot \Phi \left[ - \left\lvert \frac{\overline{Y}^{act} - \mu_{Y,0}}{\sigma_{\overline{Y}}} \right\rvert\right]. \tag{3.3} \end{align} The ppp-value can be seen as the area in the tails of the N(0,1)N(0,1)\mathcal{N}(0,1) distribution that lies beyond ±∣∣ ∣∣¯¯¯¯Yact−μY,0σ¯¯¯¯Y∣∣ ∣∣
  
  This part is very confusing
Visit annotations in context

Annotators

Gfuschini

URL

econometrics-with-r.org/3-3-hypothesis-tests-concerning-the-population-mean.html
Dec 2019
zapier.com zapier.com

Why Google Sheets Should Be Your To-Do List

1
1. TylerRick 30 Dec 2019
  
  in Public
  
  Alternatively, you can use the Explore function to ask Google Sheets any number of questions about your to do list. It'll give you suggestions like "Most frequent Task" or "What percentage of Tag is [tag name]" and so on. This can give you insight into what you're working on the most and how you're spending your time, which can help you plan your workdays more productively.
  
  Google Sheets features to try
Visit annotations in context

Tags

to try

Google Sheets

features

Annotators

TylerRick

URL

zapier.com/blog/google-sheets-to-do-list/
Local file Local file

Autophagy and Noroviruses

1
1. jessicaoros123 25 Dec 2019
  
  in Public
  
  As neither microautophagy, which involves direct uptake ofbulk cargo by lysosomes [10], nor CMA, which degrades a small subset of proteins via recognitionof a specific protein tag [11], have been implicated in host-pathogen defense against noroviruses,macroautophagy will primarily be discussed here
  
  Microautophagy: direct bulk uptake by lysosomes Chaperone-mediated autophagy: requires protein tag recognition to degrade small subset of proteins Macroautophagy: involved in host-pathogen interactions
Annotators

jessicaoros123
www.spokesman.com www.spokesman.com

‘We need each other more than we pretend’: Data contradicts sense of geographic imbalance in state taxes, services

1
1. dsiddy 24 Dec 2019
  
  in Public
  
  I may need a new (data-driven) Hypothesis tag.
  
  (action items)
Visit annotations in context

Tags

(action items)

Annotators

dsiddy

URL

spokesman.com/stories/2019/dec/21/we-need-each-other-more-than-we-pretend-data-contr/
edtechfactotum.com edtechfactotum.com

In the year of our blog 2019

2
1. mrkrndvs 21 Dec 2019
  
  in Public
  
  What is telling is the list of blogs that have refereed traffic to my blog are all written by men. I am not sure what – or if – there is anything to make out of that fact, but anytime I see a list of people that is exclusively male it does make me pause and go hmmmm.
  
  As another white male sending traffic back to your blog, I am left thinking.
2. chrisaldrich 19 Dec 2019
  
  in Public
  
  And I am planning on cutting back on my personal use of social media (easier said than done) and want to try to return to using my blog more than Twitter for sharing.
  
  certainly a laudable goal!
  
  It helped me a lot to simply delete most of the social media apps off of my phone. I scribbled a bit about the beginning of the process back in November and there's a link there to a post by Ben doing the same thing on his own website.
  
  More people are leaving social feeds for RSS feeds lately. I've recently started following Jeremy Felt who is taking this same sort of journey himself. See: https://jeremyfelt.com/tag/people-still-blog/
  
  Kudos as well to making the jump here:
  
  Taking a bit of a Twitter break. I'm going to try to stay off until the new year, but likely lack the willpower to stay off for more than a few hours. Wish me luck!<br><br>....but silently. Not via reply to to this tweet. Cause that'll just suck me back into the vortext.
  — Clint Lalonde (he/him) (@edtechfactotum) December 19, 2019
  <script async src="https://platform.twitter.com/widgets.js" charset="utf-8"></script>
  
  In part, it's what prompted me to visit your site to write a comment. (Sorry for upping your cis-gendered white male count, but 2019 was a bad year, and hopefully we can all make 2020 better as you've indicated.)
  
  A Domain of One's Own RSS feed readers social media
Visit annotations in context

Tags

RSS

social media

A Domain of One's Own

feed readers

Annotators

mrkrndvs

chrisaldrich

URL

edtechfactotum.com/in-the-year-of-our-blog-2019/
vitalirosati.net vitalirosati.net

Metaontology. Toward a prehuman thinking

1
1. marviro 19 Dec 2019
  
  in Public
  
  inscription
  
  See annotations on inscription
Visit annotations in context

Annotators

marviro

URL

vitalirosati.net/metaontology/proposal.html
infokaistatic.s3.amazonaws.com infokaistatic.s3.amazonaws.com

PHP 7 Zend Certification Study Guide_ Ace the ZCE 2017-PHP Exam ( PDFDrive.com ).pdf

1
1. _Taha 19 Dec 2019
  
  in Public
  
  Classes encapsulate code into instantiable units. Namespaces group functions, classes, and constants into spaces where their name is unique.The namespace declaration must occur straight after the opening <?php tag and no other statements may precede it.Namespaces affect constants, but you must declare them with the const keyword and not with define()
  
  Rule of using Namespaces
Visit annotations in context

Annotators

_Taha

URL

infokaistatic.s3.amazonaws.com/books/php/_pdf/PHP 7 Zend Certification Study Guide_ Ace the ZCE 2017-PHP Exam ( PDFDrive.com ).pdf
serverfault.com serverfault.com

Can I support my product on this site? - Help Center

1
1. TylerRick 19 Dec 2019
  
  in Public
  
  Types of questions and where to ask: How do I? -- ask on Server Fault (tell them what tags to use -- your product tag at minimum) I got this error, why? -- ask on Server Fault I got this error and I'm sure it's a bug -- report it on your own site I have an idea/request -- report it on your own site Why do you? -- ask in your own community (support forum, etc) When will you? -- ask in your own community
  
  types of questions and where to ask them
Visit annotations in context

Tags

types of questions and where to ask them

Annotators

TylerRick

URL

serverfault.com/help/product-support
en.wikipedia.org en.wikipedia.org

Windsor Castle - Wikipedia

1
1. dwhly 12 Dec 2019
  
  in Public
  
  Since the time of Henry I, it has been used by the reigning monarch and is the longest-occupied palace in Europe
  
  Make an note.
  
  'this is a tag'
Visit annotations in context

Tags

'this is a tag'

Annotators

dwhly

URL

en.wikipedia.org/wiki/Windsor_Castle
web.hypothes.is web.hypothes.is

Annotating the law | Hypothes.is

1
1. J.FAURE 09 Dec 2019
  
  in Public
  
  Archiving and downloading annotations Annotation viewing and export, from Hypothesis Labs Link: https://jonudell.info/h/facet Screencast: https://jonudell.net/h/facet.mp4 Description: View annotations by user, group, URL, or tag. Export results to HTML, CSV, text, or Markdown.
  
  Did anyone tried to use this to feed mind/concept-mapping tools like Tinderbox or Cmap?
  
  workflow questions
Visit annotations in context

Tags

questions

workflow

Annotators

J.FAURE

URL

web.hypothes.is/blog/introducing-math/
www.grc.com www.grc.com

Security Now! Transcript of Episode #373

1
1. stevelw 09 Dec 2019
  
  in Public
  
  Leo: Stephen in Glasgow, Scotland shares his recent NFC experience: I think I know of a problem with NFC. When I first got my Galaxy S3 it would quite often beep for no apparent reason. Every time I put it in my jacket pocket or on my table, it would beep. Then I noticed it was when I put it on my table resting on my wallet that it was beeping. I felt like an idiot for not figuring it out. Some of my newer credit cards have RFID chips inside for the new contactless payment systems. We don't have these in the U.S. yet; or, if we do, they're in very limited areas.Steve: Yes.Leo: One of the problems I had, when you go to Europe it's hard to buy gas because our credit cards are dumb, and you need a smart credit card to buy gas. And the Galaxy S3's reader was shouting out, "Hey, I found a tag." And sure enough, when I downloaded an NFC app from the Android store, the beep would then be accompanied by the card info displayed on the screen when I put my S3 near my wallet. If these phones are going to go crazy when we put them near a wallet with RFID cards, no wonder Apple is holding back. As far as I can see, there is no way to tell Android to ignore a tag. And even if you could, would that use battery as the RFID tag in your wallet was constantly shouting out, "Hey, hey, I'm here," and your phone listened to the details before ignoring it again? Love the show. That's a great question.
  
  Highlight
Visit annotations in context

Annotators

stevelw

URL

grc.com/sn/sn-373.htm
debuggable.com debuggable.com

Private npm modules » Debuggable - Node.js Consulting

1
1. TylerRick 05 Dec 2019
  
  in Public
  
  The url also contains an optional refspec (#v0.0.1) that tells npm which branch, commit, or in this case tag you want to have checked out.
  
  installing npm package from git repository
Visit annotations in context

Tags

installing npm package from git repository

Annotators

TylerRick

URL

debuggable.com/posts/private-npm-modules:4e68cc7d-1ac4-42d9-995a-343dcbdd56cb
myclass.dau.edu myclass.dau.edu

Practicum 1 Situation and applicable artifacts v3.pdf

1
1. mattschneider 05 Dec 2019
  
  in Public
  
  We passed Milestone A!
  
  Annotation (or tag) required to make this visible to Public/group. But you can still "Post to Only Me".
Visit annotations in context

Annotators

mattschneider

URL

myclass.dau.edu/bbcswebdav/institution/Courses/Deployed/ISA/ISA201/Student_Materials/LSN12 Practicum 1/Practicum 1 Situation and applicable artifacts v3.pdf
collect.readwriterespond.com collect.readwriterespond.com

📑 Secrets of the Edu-Twitter Influencers | Read Write Collect

1
1. chrisaldrich 04 Dec 2019
  
  in Public
  
  Alexander Samuel reflects on tagging and its origins as a backbone to the social web. Along with RSS, tags allowed users to connect and collate content using such tools as feed readers. This all changed with the advent of social media and the algorithmically curated news feed.
  
  Tags were used for discovery of specific types of content. Who needs that now that our new overlords of artificial intelligence and algorithmic feeds can tell us what we want to see?!
  
  Of course we still need tags!!! How are you going to know serendipitously that you need more poetry in your life until you run into the tag on a service like IndieWeb.xyz? An algorithmic feed is unlikely to notice--or at least in my decade of living with them I've yet to run into poetry in one.
  
  tags algorithms algorithmic feeds artificial intelligence social media
Visit annotations in context

Tags

artificial intelligence

social media

algorithmic feeds

algorithms

tags

Annotators

chrisaldrich

URL

collect.readwriterespond.com/📺-videodrome/
github.com github.com

makerdao/dss

3
1. vam 03 Dec 2019
  
  in MakerDAO
  
  function flow(bytes32 ilk) external note { require(debt != 0, "End/debt-zero"); require(fix[ilk] == 0, "End/fix-ilk-already-defined"); (, uint rate,,,) = vat.ilks(ilk); uint256 wad = rmul(rmul(Art[ilk], rate), tag[ilk]); fix[ilk] = rdiv(mul(sub(wad, gap[ilk]), RAY), debt); }
  
  Important notes
  
  fix[ilk] is the exchange ratio between collateral and Dai. This is where the haircut could be applied to dai holders. The gap[ilk] is the aggregation of insolvency across all under water CDPs
2. vam 03 Dec 2019
  
  in MakerDAO
  
  function skim(bytes32 ilk, address urn) external note { require(tag[ilk] != 0, "End/tag-ilk-not-defined"); (, uint rate,,,) = vat.ilks(ilk); (uint ink, uint art) = vat.urns(ilk, urn); uint owe = rmul(rmul(art, rate), tag[ilk]); uint wad = min(ink, owe); gap[ilk] = add(gap[ilk], sub(owe, wad)); require(wad <= 2**255 && art <= 2**255, "End/overflow"); vat.grab(ilk, urn, address(this), address(vow), -int(wad), -int(art)); }
  
  Important notes to consider
  
  Vow is empty → dai holders
  
  Surplus in the vow → then it will eventually be canceled (or "burned") with the incoming sin. And since the every collateral's art (and thus DCS debt) is unaltered, then this dai burning is distributing the surplus to dai holders in the form of greater share of the pie
  
  e.g. 10 MM Dai claims 1 MM eth. If 1 MM Dai is in the form of surplus and is subsequently burned, then each Dai now has a great claim on eth when only 9 MM Dai is remaining)
  
  Bad debt in the vow → increases the ending balance of Sin within the Vow. This bad debt would otherwise be covered by MKR holders, but now, it is actually covered by Dai holders
  
  Undercollateralized CDPs → more sin is accrued in the vow than ink accrued in the End
  
  These are present when CDPs have not yet been bitten when ES is initiated. They are unfortunately not penalized if they're not bitten.
  
  Overcollateralized CDPs → exact amount of sin is accrued in the vow as ink value in End
3. vam 03 Dec 2019
  
  in MakerDAO
  
  function skip(bytes32 ilk, uint256 id) external note { require(tag[ilk] != 0, "End/tag-ilk-not-defined"); (address flipV,,) = cat.ilks(ilk); Flippy flip = Flippy(flipV); (, uint rate,,,) = vat.ilks(ilk); (uint bid, uint lot,,,, address usr,, uint tab) = flip.bids(id); vat.suck(address(vow), address(vow), tab); vat.suck(address(vow), address(this), bid); vat.hope(address(flip)); flip.yank(id); uint art = tab / rate; Art[ilk] = add(Art[ilk], art); require(int(lot) >= 0 && int(art) >= 0, "End/overflow"); vat.grab(ilk, usr, address(this), address(vow), int(lot), int(art)); }
  
  Important notes
  
  First vow.suck used to accrue (TODO - still not entirely sure, but I think it has something to do with refunding the last bidder with their Dai bid)
  
  Second vow.suck accrues the amount of dai in End from the last bid to be returned to the bid.usr(which is the bitten urn) in the form of art
  
  The final vat.grab returns dai and collateral from End to bid.usr. This same process decreases the amount of sin that was accrued in the second vow. Now the CDP is an over/undercollateralized and can be skim'ed
  
  Skip doesn't have a material impact on the auction or vow accounting. For more gas, it simply stops the tend auctions, returns the dai bid to the last bidder and transfers the collateral from the flip auction to End and finally back to the bid.usr. In other words, it pulls a CDP back into a state that can be skim'ed
Visit annotations in context

Annotators

vam

URL

github.com/makerdao/dss/blob/master/src/end.sol
github.com github.com

insin/nwb

1
1. TylerRick 02 Dec 2019
  
  in Public
  
  A UMD build will let people use your module via a global variable by dropping it into a <script> tag - this makes it easier to try without any build tooling in an HTML file, and in tools like JS Bin and CodePen.
  
  excellent: explanation UMD
Visit annotations in context

Tags

excellent: explanation

UMD

Annotators

TylerRick

URL

github.com/insin/nwb/blob/master/docs/guides/ReactComponents.md
Nov 2019
scalar.usc.edu scalar.usc.edu

Scalar Class Project: Loss: The Landscape

1
1. Knk 26 Nov 2019
  
  in Public
  
  ”,
  
  needs a tag (p#).<br> The quotation needs more integration to show how "industry norms" fit in with the complex layers you're pointing to in "Dawn"
Visit annotations in context

Annotators

Knk

URL

scalar.usc.edu/works/scalar-class-project-/the-landscape
diglib.hab.de diglib.hab.de

Untitled document

1
1. KatharinaLuiseFoerg 26 Nov 2019
  
  in Public
  
  c.
  
  bitte tag erweitern: "de simpt. c." c. 3
Visit annotations in context

Annotators

KatharinaLuiseFoerg

URL

diglib.hab.de/edoc/ed000261/alpha/index.html
scalar.usc.edu scalar.usc.edu

Being Wronged: The Brain Drawing the Bullet

1
1. Knk 24 Nov 2019
  
  in Public
  
  The narrative is a hypertext story that changes every time the reader interacts with it, to create the sense of an unreliable narrator and stand as an allegory for the process of memory.
  
  you are closely summarizing the editor's and author's introductions to the work on the ELC3. You even use many of the same words.
  
  Without a citation, this is plagiarism. Please revise to include citations either with the lead in format or all in the tag (cite)
Visit annotations in context

Annotators

Knk

URL

scalar.usc.edu/works/essay-3---raghav-kris-sara-dom/the-brain-drawing-the-bullet
scalar.usc.edu scalar.usc.edu

RUSI DIGITAL EXHIBIT: RUSI Digital Exhibit [3]

1
1. schuster 24 Nov 2019
  
  in Public
  
  For this option to work we will need to: * work through a way to tag the landing page in a consistent way
  
  Add titles and captions to tagged images that help with navigation
  
  provide introductory and instructional text to help users understand how to navigate content
  
  Questions:
  
  Do we want just one structured media gallery or a structured media gallery for each seminar path?
Visit annotations in context

Annotators

schuster

URL

scalar.usc.edu/works/rusi-digital-exhibit/rusi-landing-page-3
garingtaylor1.multiscreensite.com garingtaylor1.multiscreensite.com

Garing, Taylor & Associates, INC.

1
1. rreillycp3 22 Nov 2019
  
  in Public
  
  Let's ditch the Civil Engineers, Surveyors and Planners part for this logo. It makes our tag line repetitive.
Visit annotations in context

Annotators

rreillycp3

URL

garingtaylor1.multiscreensite.com/
dis.lib.usf.edu dis.lib.usf.edu

Cities, Heterotopias & Science Fiction (F19): Short Story Draft 2

2
1. crwestley 19 Nov 2019
  
  in Public
  
  Jacqueline knew better than to interrupt her sister when she was this heated; you don’t get to be the richest woman in the tech industry without a ruthless reputation.
  
  Maybe add another dialogue tag for Lizzie; she should probably be mentioned since these are her words.
2. crwestley 19 Nov 2019
  
  in Public
  
  Jacqueline fired back at her sister.
  
  MY BAD, there is a tag here lol; possibly shift it up earlier in the dialogue though.
Visit annotations in context

Annotators

crwestley

URL

dis.lib.usf.edu/scalar/cities-heterotopias--science-fiction-f19/short-story-draft-2

Tags

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Tags

Annotators

URL

Annotators

URL

Annotators

URL

Reply to the reviewers

Tags

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Tags

Annotators

URL

Annotators

URL

Reply to the reviewers

1. Summary of main points by the reviewers, and authors’ responses:

2. Detailed revision plan for the preprint:

3. Point-by-point responses to the reviewers

Referee #2

Evidence, reproducibility and clarity

Significance

Tags

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Annotators

URL

Tags

Annotators

URL

Tags

Annotators

URL

Annotators

URL

Annotators

URL

Tags

Annotators

URL

Annotators

URL

Tags

Annotators

URL

Annotators

URL

Tags

Annotators

URL