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  1. May 2020
    1. RNA analysesFrozen tissue (optimal cutting temperature [OCT]) sections were obtained from the nPOD [18] and DiViD tissue collections. Tissue slides were fixed and laser-capture of islets conducted as previously described [19]. All islets in two to five sections of tissue from each donor were captured and pooled and RNA extracted using the Arcturus PicoPure RNA Isolation Kit (Applied Biosystems, Grand Island, NY, USA). Quality and quantity of RNA was determined on a Bioanalyzer 2100 instrument (Agilent Technologies, Santa Clara, CA, USA). Samples with sufficient quantity and quality of RNA were then subjected to gene expression analysis using Affymetrix expression arrays (GeneChip Human Gene 2.0 ST) and scaled normalised gene expression values produced as previously described [20]. The normalised expression data for 70 genes of interest were then subjected to analysis as described below.

      RNA Analysis

      (GeneChip HUman Gene 2.0 ST)