GSM8252352: Processed_donor; Mus musculus; RNA-Seq
Spleen tissues were processed with the flat end of a syringe in a 100 mm culture dish containing 5 ml cold FACS buffer (2% FBS in PBS), then passed through a 70 μm cell strainer into a 15 ml tube. Cells were centrifuged to remove the supernatant. Cell pellets were treated with 1ml ACK (Ammonium-Chloride-Potassium) Lysing Buffer to remove the red blood cells. After washing with 10 ml cold FACS buffer, the remaining cells were stained with 7AAD (Part 76332; Lot B226294 Biolegend) for 30 min at 4 °C, then washed and resuspended in cold FACS buffer for flow cytometric sorting using FACS Aria II Cell Sorter (BD Biosciences). Sorted YFP+7AAD- cells with a viability higher than 90% were used for 10X genomics scRNA-seq. Finally, libraries were sequenced on a NovaSeq 6000 system with NovaSeq 6000 S4 Reagent Kit (300 cycles). scRNA libraries were performed according to the manufacter's instructions (single cell 3' v2 protocol, 10x Genomics) and were prepared for sequencing using standard Illumina protocols.