GSM9686913: JR19_1207_D12 | JR191207_D12_008 | NKG7- CD8+; Homo sapiens; OTHER
Mouse splenic and thymic CD8+ T cells were isolated using CD8a+ T cell isolation kit (Miltenyi Biotec, Cat#130-104-075). Mouse splenic naïve CD8+ T cells were isolated using Naive CD8a+ T Cell Isolation Kit, mouse (Miltenyi Biotec, Cat#130-096-543). For isolating CD8+ tumor infiltrating lymphocytes (TILs), tumors were first cut into small pieces of 2-4mm and digested using tumor dissociation kit at 37°C for 45 minutes with gentle agitation (Miltenyi Biotec, Cat#130-096-730). Dead cells were further removed using Dead cell Removal kit (Miltenyi Biotec, Cat# 130-090-101). CD8+ TILs were then isolated using CD8 (TIL) microbeads, mouse (Miltenyi Biotec, Cat#130-116-478). ROIs were selected based on fluorescence imaging using the GeoMx instrument. The selected ROIs were then hybridized with barcoded RNA detection probes (GeoMx DSP RNA assays). Following hybridization, ultraviolet (UV) light was used to release ROI-specific oligonucleotide barcodes. Released barcodes were collected, amplified by PCR. Library quality and quantity were assessed using an Agilent 2100 Bioanalyzer and qPCR. Sequencing was performed on an Illumina NovaSeq S4 platform, generating paired end reads.