GSM9230247: 1920B007 day 20, BAL, GEX, biol rep 1; Homo sapiens; RNA-Seq
Non-naïve CD8+ T cells were FACS sorted from cryopreserved PBMC using a FACSAriaII (BD Biosciences) cell sorter. The following antibodies were employed: CD3 FITC (clone HIT3a, BioLegend), CD8 PE (clone RPA-T8, BioLegend), CD45RO Pacific Blue (clone UCHL1, BioLegend), CCR7 APC-R700 (clone 3D12, BD Biosciences), and live-dead discrimination with Zombie Aqua Fixable Viability Kit (BioLegend). We sorted all live CD3+CD8+ PBMC that were not CD45RO-CCR7+ for inclusion in the single cell RNAseq. Total cryopreserved BALF immune cells were thawed and further purified using the EasySep Dead Cell Removal (Annexin V) Kit (STEMCELL Technologies) prior to single cell RNAseq. Following purification, non-naïve CD8+ T cells or BALF cells were re-suspended in PBS supplemented with 0.05% bovine serum albumin. 10x Genomics Chromium Single Cell 5' v2 Gene Expression Dual Index libraries and Chromium Single Cell V(D)J Dual Index libraries were prepared according to the manufacturer's instructions without modifications.