GSM9686031: CD4⁺ T-cells, PCS High,GIR08; Homo sapiens; RNA-Seq
Frozen PBMCs were thawed, and total CD4+T-cells were enriched using the EasySep™ Human CD4+T-Cell Enrichment Kit (StemCell Technologies, Vancouver, CA), following the manufacturer's instructions. Cells were washed with PBS containing BSA 0.04%, and viability was assessed by staining the cells with Trypan blue. Cells were resuspended at the desired concentration to target 10,000 cells per sample. Single-cell suspensions were prepared using the Chromium Next GEM Single Cell 5' Reagent Kits v2 (10x Genomics, Pleasanton, CA) and partitioned into Gel Beads-in-emulsion (GEMs) using the 10x Chromium Controller and Next GEM Chip K. GEMs were visually inspected, and only samples with an opaque and uniform aspect were used for library preparation. Gene Expression (GEX) libraries were generated from the cDNA. Representative traces and quantitation of libraries were determined using Bioanalyzer High Sensitivity DNA Analysis (Agilent, Santa Clara, CA). GEX libraries were sequenced on Illumina NovaSeq S4 system, using 10x's recommended parameters. The 10x Barcodes in the library were used to associate individual reads back to the individual partitions, and thereby, to each single cell.