GSM6268589: EMT-6 tumor, anti-PD1, replicate 1, scRNAseq; Mus musculus; RNA-Seq
Tumors were isolated from the mouse mammary pad and placed in a Petri dish with C10 media (RPMI 1640, HEPES 1%, Penicillin/Streptomycin 1X, fetal calf serum 10%, sodium pyruvate 1 mM, non-essential amino acids 1X, and beta-mercaptoethanol 0.45%) and minced with sterile razor blades. The tissue slurry was digested with Collagenase XI 2 mg/mL, Hyaluronidase 0.5 mg/mL, and DNase 0.1 mg/mL prepared in C10 medium. Cells were incubated in a shaker at 255 rpm for 30 minutes at 37 C°, then digestion was stopped with ice cold media.  The slurry was passed through a 100 µm mesh strainer into a clean 50 mL conical tube. Cells were pelleted by centrifugation for 5 minutes at 200 x g at 4 C°and mixed with RBC lysis buffer and passed through a 40 µm mesh strainer and washed with PBS. Live cells stained with Thy1.2 and CD45 were sorted into tumor and non-tumor cells groups and resuspended to enrich non-tumor cells. Library was constructed according to manufacturer's protocol (Chromium single cell 3' v2 protocol, 10X genomics)