GSM8502293: Lung CD31+ cells, HO, rep2; Mus musculus; RNA-Seq
Lung tissues were harvested and placed into 1.5 ml microcentrifuge tubes. Subsequently, 1 ml of digestion solution was added (50 μg/ml Liberase TM(Sigma) in RPMI1640 culture medium (Gibco)). The lung tissue was finely minced with scissors and incubated at 37°C on a rotary shaker at 300 rpm for 30 minutes, tissues were then transferred to C-tubes, and single-cell suspension was obtained using the GentleMACS Dissociator (Miltenyi). Cell pellet was collected by centrifugation at 300g for 5 minutes in FACS Buffer (1 mM EDTA, 25 mM HEPES, 0.5% BSA in DPBS). 100 μL of FcR (BioLegend, 101302, 1:200) blocking solution was then added, and incubated at 4°C for 45 minutes. Cells were incubated with anti-mouse CD31-micro beads (MiltenylBiotec, 130-097-418), at 4°C for 45 minutes. Magnetic columns were balanced with FACS Buffer, and the cell-bead suspension was applied to the column on a magnetic field to obtain CD31+ endothelial cells. RNA-seq library was prepared using the VAHTS Universal V8 RNA-seq Library Prep Kit for Illumina from Vazyme according to the manufacture's instruction.