GSM9669093: recurrent GBM tumors; Homo sapiens; RNA-Seq
CD45+ cells and EGFP+ cells from tumor tissues were extracted using a cell sorter (ARIA I). CD45+ cells in PB were purified using EasySep Mouse/Human Chimera Isolation Kit (STEMCELL). Cell viability was assessed using trypan blue exclusion; only samples with >90% viability were used. To minimize technical batch variability, samples from humanized and naive mice were harvested on the same day and processed in parallel. Surface staining was performed using TotalSeq™-B anti-human hashtag antibodies described in the BioLegend protocol (TotalSeq™-b or -c with 10x feature barcoding technology. Following the manufacturer's protocol, single-cell RNA libraries were prepared using the Chromium Single Cell 3' Reagent Kits. Briefly, cells were loaded onto the 10x Genomics Chromium to capture approximately 20,000 cells. Reverse transcription and cDNA amplification were performed according to the kit protocol. Libraries were sequenced on Illumina NovaSeq 6000 with paired-end 150 bp reads to a depth of approximately 5000 reads per cell.