GSM6215769
Seven days after sort, 10X 3'v3.1 with Feature Barcoding was performed. 60,000-120,000 cells were harvested and resuspended in 50 uL of PBS + 1% BSA. Cells were then blocked with 5 uL of TruStain FX for 10 minutes. Next, cells were stained with 0.5 uL of each TotalSeq-B antibody (CD34, CD38, CD45RA, CD90, CD49f, CD35, CD11a, CD59, CD117) for 30 minutes. Following 4 washes with PBS + 1% BSA, 10,000 cells were loaded onto a Chromium Next GEM Chip G GEM generation & barcoding, post GEM–RT cleanup & cDNA amplification, 3' gene expression library construction, and cell surface protein library construction were performed as described in CG000317_ChromiumNextGEMSingleCell3'v3.1_CellSurfaceProtein_RevC (https://support.10xgenomics.com/single-cell-gene-expression/index/doc/user-guide-chromium-single-cell-3-reagent-kits-user-guide-v31-chemistry-dual-index-with-feature-barcoding-technology-for-cell-surface-protein). Gene expression and cell surface protein libraries were pooled together at a ratio of 4:1 and sequenced on an Illumina NovaSeq.