GSM9169279: Ctrl, bio rep 2, TCR; Mus musculus; OTHER
Tumors were minced with razor blade and then digested with dissociation media (RPMI media supplemented with collagenase, dispase II, DNase and Calcium chloride (5mM))for 30 minutes in 37oC using a rotating platform. Digested tumor cells were filtered with 40μm filter and counted using Countess 3 (Invitrogen). Resuspend the cells pass through filter to 10x106 cells/ml in 1% BSA/PBS; followed by staining with Live/Dead Far-Red (ThermoFisher) and FITC conjugated anti-mouse CD45 antibody (Biolegend) on ice for 20 minutes. CD45 positive live cells were sorted with Sony MA900 multiapplication cell sorter at the Duke Cancer Institute Flow Cytometry core. The sorted cells were resuspended in 45ul of 0.04% BSA/PBS, stained with trypan blue and counted with hemocytometer. Cells (10,000) were used for GEM generation and barcoding, followed by the construction of V(D)J library and 5' gene expression (GEX) libraries using the Chromium Next GEM single cells 5' V2 (Dual Index; 10X Genomics) according to manufacturer's protocol.