Immunohistochemistry Sections (4μm) from formalin fixed paraffin embedded pancreas tissues were deparaffinized and rehydrated with serial passage through changes of xylene and graded ethanol. All slides were subjected to heat induced antigen retrieval in Target Retrieval Solution (Dako). The tissue sections were stained for insulin as a part of routine collection protocol for nPOD tissues (previously described (Campbell-Thompson et al., 2016; Campbell-Thompson et al., 2012b)) or double stained for insulin (polyclonal guinea pig anti-insulin,1:1000 dilution, Dako (Santa Clara, CA)) and glucagon (monoclonal mouse anti-glucagon, 1:5000 dilution, Abcam, Cambridge, MA) by immunohistochemistry (IHC), scanned using an Aperio CS Scanscope (Leica/Aperio, Vista, CA), and stored in the nPOD online digital pathology database (eSLIDE version 12, Leica/Aperio). For donors indicated in Table S1, scanned images of insulin-stained slides available from the block(s) nearest to the tissues used for total protein extraction (described above) were evaluated for fractional insulin area using Indica Labs, Inc image analysis software (Corrales, NM). For insulin and glucagon double stained slides, antigen-antibody binding was visualized using the EnVision G/2 Doublestain System (Dako). For control (n=5) and T1D subjects (n=11), glucagon + insulin positive islets, glucagon positive islets, insulin positive single cells, and clusters of two to five insulin positive cells were annotated by hand using the Aperio viewing platform and analyzed using Image Scope, Leica Biosystems analysis software (Version 12.1.0.5029, Buffalo Grove, IL).

Proinsulin, Insulin, C-peptide, Islet Amyloid Polypeptide and Glucagon by ELISA and Luminex Commercially available kits from ALPCO (Salem, NH) were utilized to measure total proinsulin, insulin and C-peptide from pancreas protein extracts as indicated in Table S1. The proinsulin, insulin and C-peptide assays are specific and do not cross react with each other. Glucagon was measured with a commercial ELISA assay provided by Mercodia (Winston Salem, NC). Islet amyloid polypeptide was measured by an assay from Millipore (Billerica, MA) using magnetic bead technology (Luminex). A total protein Bradford assay from Thermo-Fisher (Waltham, MA) was performed on each supernatant and used to normalize mass and extraction efficiency.