GSM8412806: LNP-M/mRNA-Spike, scRNA-seq-BCR (Spleen); Mus musculus; RNA-Seq
The single-cell suspension of spleens from mice immunized 3 times vaccine was prepared, and CD45+ cells were purified using the EasySep™ Mouse TIL (CD45) Positive Selection Kit following the manufacturer's instructions. Tumors were harvested 14 days following LNP-pR282W-mAb or LNP-Ctrl treatment, single cell suspensions were prepared. The cells of spleen or tumor samples were resuspended in RPMI1640 with 5% FBS. To create the scRNA-seq libraries, equivalent numbers of single cells from three mice per group were combined at a density of 1 × 103 cells/μl in a 10% FBS RPMI1640 medium. Each group used about 10,000 cells, which were converted to barcoded scRNA-seq libraries using the ChromiumTM Next GEM Single Cell 5′ Kit v2 (10× Genomics, Pleasanton, CA, USA; 1000263) according to the manufacturer's protocol. To prepare full-length T-cell Receptor (TCR) and B-cell Receptor (BCR)V(D)J libraries, Chromium Single Cell Mouse TCR Amplification Kit (10× Genomics, 1000254) and Chromium Single Cell Mouse BCR Amplification Kit (10× Genomics, 1000255) was used to enrich amplified cDNA from the 5′ libraries. Approximately 10,000 cells were allocated for the 10× Genomics 5′ v2 single-cell assay. TCR/BCR libraries were prepared using the Chromium™ single cell V(D)J enrichment kit (10× Genomics). Reverse transcription, cDNA amplification and library preparation were executed following the manufacturer's protocol.