GSM8902865: TILs from CT26 tumors 6 days after 12 Gy treatment 3; Mus musculus; RNA-Seq
Tumors from treated mouse models were harvested on day 6 after radiotherapy. To obtain single-cell suspensions, tumors were processed using the Tumor Dissociation Kit, mouse (Miltenyi Biotec, Bergisch Gladbach, Germany) according to the manufacturer's instructions. Following dissociation, cells were resuspended in autoMACS® Rinsing Solution with MACS® BSA Stock Solution diluted 1:20 and the cell suspensions were passed through a 70-µm cell strainer. Immune cells were isolated from cell suspensions using CD45 (TIL) MicroBeads, mouse (Miltenyi Biotec, Cat. No. 130-110-618) following manufacturer's instructions. Isolated cells were counted using Cellaca MX Cell Counter (Nexcelom Bioscience, Lawrence, MA). Cells were processed for scRNAseq using the 10x Genomics chromium v3 3' chemistry pipeline according to the manufacturer's instructions. Cells were resuspended in 0.04% BSA in PBS at a concentration of 1 × 10⁶ cells/ml. Cells were loaded onto a Chromium Single Cell B Chip (10x Genomics, Pleasanton, CA) to aim for target cell recovery of 8 × 10³ cells. Library construction was performed according to Chromium Single Cell 3ʹ GEM, Library & Gel Bead Kit v3 protocol (10x Genomics). Libraries were sequenced using the NovoSeq 6000 platform (Illumina; San Diego, CA) aiming for a minimum of 200 × 10⁶ reads per library (25,000 reads per cell).