3199A_2
Single-cell RNA sequencing (sc-RNA seq) was performed using the 10x genomics chromium single cell controller. Briefly, tumor cell single cell suspensions were prepared as indicated above. Cells were resuspended in freezing media containing 90% AB serum and 10% DMSO and stored in liquid nitrogen until analysis. For sc-RNA seq analysis cells were thawed, washed and sorted for viable CD45+ cells using the BD FACSAria. Next, cells were droplet separated using Chromium Single Cell 3 v2 Reagent Kit with the 10x genomics microfluidic system creating cDNA library with individual barcodes for individual cells. Barcoded cDNA transcripts from GBM patients were pooled and sequenced using the Ilumina HiSeq 4000 Sequencing System.