GSM9814814: IL-1a TNFa C1q EV+MAPS; Mus musculus; RNA-Seq
Brain tissues were immediately collected and washed twice with cold RNase-free PBS. Samples were minced and enzymatically dissociated in digestion buffer containing Collagenase IV, Papain, and DNase I for 20 min at 37 °C. Enzymatic digestion was terminated with PBS supplemented with 10% FBS, followed by mechanical trituration. Cell suspensions were filtered through 70 µm and 30 µm strainers. Red blood cells and dead cells were removed using erythrocyte lysis buffer and Dead Cell Removal MicroBeads (MACS), respectively. Live cells were washed and resuspended in PBS containing 0.04% BSA. Only cell suspensions with viability greater than 85% were used for downstream library preparation. Single-cell suspensions were loaded onto Chromium Single Cell 3′ Chips (10x Genomics, v3) targeting approximately 8,000 cells per sample. Single-cell barcoding, reverse transcription, cDNA amplification, and library construction were performed according to the manufacturer's instructions. Libraries were sequenced on an Illumina NovaSeq 6000 platform using paired-end 150 bp sequencing.