GSM8510673: CD45+ cells, NCD; Mus musculus; RNA-Seq
After euthanizing the mouse, the liver was perfused with 10 mL of PBS through the heart, collected, and disrupted using a gentleMACS tissue dissociator. The resulting tissue was resuspended in RPMI-1640 media containing 10% FBS and further digested with collagenase. The suspension was centrifuged at 60 g for 1 minute, and the supernatant containing hepatic immune cells was collected and centrifuged again at 480 g for 8 minutes. The pellet was resuspended in 10 mL of 37.5% Percoll (Sigma) in PBS and centrifuged at 850 g for 30 minutes. The resulting pellet was washed, followed by red blood cell lysis using ammonium-chloride-potassium (ACK) lysis buffer for 2 minutes at room temperature. Total CD45+ cells were then purified magnetically according to the manufacturer's protocol. After labeling with hashtags, cells were pooled and loaded into the wells of a 10x Genomics chip using the single-cell 3' kit. Gene expression and CMO libraries were sequenced on a NovaSeq S4 chip (2x150 bp PE).