GSM8808080: Blood, Immune cells, PLX5622, 14 dpi, rep1; Mus musculus; RNA-Seq
Mice were euthanized and subjected to ice-cold saline perfusion at either 5- or 14-days post MCAO. Ischemic hemispheres were collected, excluding meninges, the olfactory bulb, or cerebellum. For the scRNA-seq analysis of entire brain cells, brain homogenates were created using the Adult Brain Dissociation Kit (T) with a gentle MACS dissociator with heaters (Miltenyi Biotec), following the provided guidelines. In the case of scRNA-seq for CD45high immune cells, brain homogenates were generated using the Neural Tissue Dissociation Kit (T), and CD45high immune cells were isolated through FACS using the anti-CD45-PE-Cy5 antibody (Thermo Fisher Scientific, 15-0451-81). For PBMC specimens, peripheral blood was collected from the right atrium, followed by two rounds of red blood cell lysis (eBioscience, 1X RBC lysis buffer, 00-4333057). Libraries for 10X Genomics were prepared using the 10X Genomics Chromium Single Cell 3' v3 chemistry, in accordance with the manufacturer's instructions.Subsequently, the mixture containing captured and barcoded mRNAs was retrieved from the Chromium instrument, followed by reverse transcription. The cDNA samples were fragmented and amplified per the 10X protocol. The libraries were then purified, quantified, and sequenced utilizing the Illumina NovaSeq sequencer.