GSM6996162: KID90, gd T cells, biol rep 1; Homo sapiens; RNA-Seq
Surgical specimens were obtained fresh from the operating field, and dissected in surgical pathology, where tumors were isolated, minced, and transported at room temperature immersed in L15 media with 15 mM HEPES and 600 mg/mL glucose. Resected tumor kidney tissues were further minced using surgical scissors in GentleMACs C Tubes (Miltenyi Biotec) containing RPMI-1640 media with 0.1 mg/mL of Liberase TL and 0.2 mg/mL of DNAse I. Samples were then simultaneously mechanically and enzymatically digested for 1 hour at 37C using a GentleMACS dissociator (Miltenyi Biotec), according to the manufacturer TDK-1 tumor protocol. The digestion reaction was quenched with FACS-buffer (PBS buffer supplemented with 2% heat-inactivated FCS and 2 mM EDTA) and filtered through a 100 μm cell strainer. Samples were resuspended in a 155 mM NH4Cl solution to lyse all erythrocytes and washed in FACS-buffer. Finally, the samples were either frozen for storage or resuspended for flow cytometry staining or RNAseq. RNA libraries for RNA-seq were prepared according the Smart-seq2 protocol. ( https://www.nature.com/articles/nprot.2014.006 )