GSM4861160: PBS; Mus musculus; RNA-Seq
Tumors were excised, minced with scissors, and incubated in RPMI with tumor digestion enzymes (Miltenyi tumor dissociation kit, Cat. no. 130-096-730) for 30 minutes at 37°C. Tumors were filtered through a 40µM strainer and cells were resuspended in FACS buffer (PBS with 2% FBS). Cells were counted with a hemocytometer and equal portions of cells from each mouse in a sample group were pooled before staining with anti-CD45-FITC (Biolegend clone 30-F11) and ZombieNIR (Biolegend #432105). Cells were sorted for ZombieNIR- CD45+ events into collection buffer (RPMI 1640, 25mM HEPES, 10% FBS) on a BD FACS Aria II at the Dana-Farber Flow Sorting Core. Sorted cells were washed one time with 5 mL of PBS + 0.05% ultrapure BSA, counted via hemocytometer, and resuspended to a concentration of 1,000 cells per microliter. Library preparation was performed with the Chromium Single Cell 5' Library and Gel Bead Kit (Part Number 1000006)