GSM5941798: T5_TCR; Mus musculus; RNA-Seq
Tumors were dissected, cleaned, digested with MA dissociation buffer for 30 min, passed through a 70 um cell strainer and centrifuge 5 min at 1500rpm. ACK lysis was performed for 3 min and centrifuge 5 min at 1500rpm and filter with 70um cell strainer. Cells were resupend in PBS 0.04% BSA and counted. Spleens were cleaned, smashed on 40um cell strainer and centrifuge 5 min at 1500rpm. ACK lysis was performed for 3 min, centrifuge 5 min at 1500rpm and filter with 40um cell strainer. Cells were resupend in PBS 0.04% BSA and counted. Cells were prepared for scRNAseq at 10 000 cells/ul (10.106 cells/mL) in PBS 0.04% BSA and 1.106 cells/50uL in PBS 2% BSA. The 10X sample processing was performed according to established protocols, aiming at 4000 cells/sample. Single-cell experiments were performed using the 10X Chromium platform using the 5' v2 kit (dual index, PN-1000263). TCR V(D)J amplification was performed using 10x Chromium Single Cell mouse TCR amplification kit (PN-1000254). The libraries were sequenced on an Illumina HiSeq4000 platform, with run conditions as per 10X recommendations, aiming at 50'000 reads/cell.