GSM5525496: CD45Pos_eAT_SVF_Lean_HTO&ADT; Mus musculus; OTHER
Epididymal adipose tissue was collagenase digested using Collagenase Type 2 (Worthington) for 30 minutes at 37 degrees C. Red blood cells were lysed for 3 minutes using ACK lysis buffer. The stromal vascular fraction was collected after centrifugation at 400RCF for 8 minutes. Cells were immunolabeled with anti-CD45 magnetic beads (Miltenyi) and sorted on an AutoMACs (Miltenyi) using the `possel_s` setting. The positive magnetic fraction (CD45+ cells) were sorted for viability (DAPI-) on a BD FACSAria III. Libraries were prepared according to manufacturer's instructions for 10X Chromium Single Cell 5' Reagent Kit v2 with the 5' Feature Barcode Library Kit by the Vanderbilt Technologies for Advanced Genomics core targeting 20,000 cells/lane and sequenced on a NovaSeq 6000 targeting 50,000 reads per cell.