GSM5223078: LM; Mus musculus; RNA-Seq
Mice were euthanized and adipose tissue was separated from lymph nodes and surrounding muscle. Isolated tissue was then minced and digested in DMEM (Gibco) containing 2.5% bovine serum albumin, 2 mM L-glutamine, 100 U/mL Penicillin/Streptomycin, 2 mg/mL Collagenase I (Thermo) and 2 mg/mL Collagenase II (Thermo) for 45 min at 37°C with gentle rotation. During the last 15 minutes of incubation 2 mM EDTA was added to the media. Finally, the cell suspension was filtered through a 100 µm strainer. Cells in stromal vascular fraction (SVF) were separated from the adipocyte layer by centrifugation. Single live cells were further purified via fluorescent activated cell sorting, excluding dead cells labelled with LIVE/DEAD Fixable Aqua Dead Cell Stain Kit and doublets based on side and forward light scatter. scRNA-seq of SVF cells was performed using a 10X Genomics Chromium Controller. Single cells were processed with GemCode Single Cell Platform using GemCode Gel Beads, Chip and Library Kits (v2) following the manufacturer's protocol. Libraries were sequenced on HiSeq 3000 (Illumina).
GEO
GSM5223078