title: GSM8807038: TBI_Hypothermia_rep2; Mus musculus; RNA-Seq
library_construction_protocol: 1. At 24 hours post-injury (or sham), mice were deeply anesthetized (ketamine 100 mg/kg + xylazine 10 mg/kg).  2. Mice were transcardially perfused with ice-cold oxygenated artificial cerebrospinal fluid (aCSF) to remove blood.  3. The ipsilateral cortex was dissected, avoiding white matter tracts.  4. Cortical tissue was enzymatically and mechanically dissociated using the Miltenyi Neural Tissue Dissociation Kit-P, with modifications:  • 30-minute incubation in Enzyme P/Buffer X at 37°C, triturating every 5 minutes.  • Sequential trituration with decreasing bore pipettes to generate single-cell suspensions.  • Filtration through a 70µm strainer.  • Myelin removal using Myelin Removal Beads II and LS Magnetic Bead Columns (Miltenyi).  5. Cells were resuspended in MACS buffer, counted, and viability assessed (>75% required).  6. Approximately 10,000 cells per sample were loaded into the 10x Chromium platform. Single-cell 3' libraries were prepared using the 10x Genomics Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (Cat# PN-1000121) following the manufacturer's protocol.Libraries were sequenced on an Illumina NovaSeq 6000 with paired-end 150 bp reads.
identifiers.namespace: GEO
identifiers.id: GSM8807038_r1