GSM9032694: IGRP-CD8 T cells from HEL tolerized graft [HEL_graftt_GEX]; Mus musculus; RNA-Seq
Single cell suspension were made from islet graft by digesting for 15 minutes in 2U/ml dispase II in HBSS. IGRPCD8 T cells were sorted by FACS from day 8 islet grafts (lymphocytes, live, CD45+, lineage-(CD11c, CD11b, CD19, Ly6G, CD4), IGRP tetramer+). After sorting, cells from individual animals were barcoded with hashtag antibodies biolegend type C (information on hastags in "feature_README.txt") to deconvolute after sequencing. Three animals were hashed in the 2.5HIP sample and 4 animals hashed in the HEL sample. Cells were resuspended in .1% FBS in PBS for capture. Single cell gene expression profile, cell surface proteomics profile, and T cell receptor profile were obtained using the Chromium Single-Cell 5' NextGEM Library and Gel Bead Kit from 10X Genomics. Sorted and hashed cells were partitioned on the 10X Genomics Chromium X instrument for single cell capture, followed by amplification of cDNA and library preparation to recover 10,000 - 12,000 cells per sample. Individual libraries corresponding to gene expression (GEX), cell surface protein (CSP), and T cell receptor (TCR) were quantified, pooled, and sequenced on the Illumina NovaSeq 6000 sequencer at a depth of 50,000 reads/cell for gene expression, 5,000 reads/cell for cell surface protein, and 5,000 reads/cell for T cell receptor, respectively.