GSM8641188: Aged + Control + ICH, replicate 3, scRNA-seq; Mus musculus; RNA-Seq
The perihematomal brain tissues from aged ICH mice were dissociated enzymatically to obtain a single-cell suspension with the self-prepared hydrolysate or the hydrolysate kit (MACS Mouse Tumor Dissociation kit DS_130-096-730). Next, Hank's Balanced Salt Solution with calcium and magnesium (Gibco, Germany) was added to stop the enzymatic reaction. A 70 and 40 μm strainer was used to filter the resulting cell suspension, followed by centrifuging at 300 × g, 4 °C for 10 min. Then, myelin was removed by centrifugation on a 22% Percoll gradient. Briefly, cells were suspended in 25 mL Percoll solution [18.9 mL gradient buffer containing 5.65 mM NaH2PO4H2O, 20 mM Na2HPO42 (H2O), 135 mM NaCl, 5 mM KCl, 10 mM glucose, 7.4 pH; 5.5 mL Percoll (GE Healthcare, Germany); 0.6 mL 1.5 M NaCl)], overlayered with 5 mL PBS and centrifuged for 20 min at 950 × g and 4 °C, without acceleration and brakes. We prepared single cell suspensions from cultured cell lines and single cells were suspended in PBS containing 0.04% BSA. The cell suspension was loaded onto the BMKMANU C1000 Chip and ran the Droplet Generation Equipment to generate single-cell gel beads in the emulsion(GEMs) according to the manufacturer's recommendation.captured cells were lysed and the released RNA was barcoded through reverse transcription in individual GEMs. Barcoded, full-length cDNA was generated and libraries were constructed according to the performer's protocol. The quality of libraries was assessed by Qubit 4.0 and the Agilent 2100.