scRNA-Seq of mus musculus after R-CNP@M vaccination: adult female lung
The female BALB/c mice (6-8-weeks old) were immunized with R-CNP or R-CNP@M (n=4 per group) as described above, and mice were sacrificed at 21 days post prime vaccination. The lungs were harvested and minced with scissors, following treated with collagenase I (1 mg/mL) and DNase I (0.05 mg/mL) and dissociated to a single-cell suspension. After treated with red blood cell lysing reagent, the single-cell suspension was passed through a 40 m strainers and resuspended in PBS. The single-cell suspension of four mice in each group were pooled. The CD45+ cells were isolated from single-cell suspension in each group by CD45 MicroBeads (130-052-301, Miltenyi Biotec), and then these CD45+ single-cell suspensions that containing about 10000 cells in each group were loaded on a 10X Genomics GemCode Single-cell instrument to generates single-cell Gel Bead-In-EMlusion (GEMs). Libraries were generated and sequenced from the cDNAs with Chromium Next GEM Single Cell 3 Reagent Kits v3.1. The Gene Denovo Biotechnology Co. (Guangzhou, China) were entrusted to perform GEMs generation, cDNA amplification, library preparation and sequencing in this study. Overall, 10694 cells from the PBS group, 9294 cells from the R-CNP group and 13033 cells from the R-CNP@M group passed the quality control threshold of more than 200 genes were identified in each cell.