GSM5533977: all_cells_tcr; Mus musculus; RNA-Seq
Murine tissue samples (quadriceps femoris muscle and inguinal lymph nodes) were obtained for single cell flow cytometry and/or fluorescence activated cell-sorting. Quad tissue samples were finely diced and digested with 1.67 Wünsch U/mL Liberase TL (Roche Diagnostics) and deoxyribonuclease I (0.2 mg/ml; Roche Diagnostics) in RPMI-1640 medium (Gibco) for 45 min at 37°C. The digested quad tissues were ground through 70-µm cell strainers (Thermo Fisher Scientific), rinsed with RPMI-1640 + 5% fetal bovine serum. Percoll (GE Healthcare) density gradient centrifugation was used to remove debris from quad digestions and enrich the leukocyte fraction. Inguinal lymph node samples were ground through 70-µm cell strainers with excess RPMI-1640 + 10 mM HEPES buffer solution. The enriched single-cell suspensions were washed and stained following antibody panels, respective to the intended application. No differences in single-cell isolation from different material environments were observed. According to 10X protocol All samples were pooled into one after hashing. The library was split appropriately into sequencing lanes as needed for sequencing depth.