Fitc conjugated mouse anti cd150a pdgfra antibody

SVF isolation was performed as described elsewhere (Cho et al., 2014 (link)); briefly, mouse gonadal fat depots were washed with PBS on ice and then minced and incubated with collagenase type II (1 mg/ml) (Sigma-Aldrich) and incubated on a shaker water bath incubator at 37°C for 20 minutes with vigorous shaking by hand every 5 minutes. EDTA, pH 8.0 at a concentration of 10 mM was then added for an additional 5 minutes, after which wash media (DMEM+glutamax with 10% bovine calf serum and 1% penicillin-streptomycin) or FACS buffer (PBS with 1mM EDTA, 25 mM HEPES and 1% fetal bovine serum) was added. Cells were filtered through a 100 µm filter and washed, and if flow cytometry was to be performed, incubated with ACK lysis buffer for 5 minutes. SVF was then washed and filtered through 40 µm filter followed by plating on collagen-coated multiwell plates in growth media (DMEM+glutamax, 15% FBS, 1% penicillin-streptomycin) or suspended in FACS buffer for flow cytometry. For flow cytometry, cells were incubated in FITC-conjugated mouse anti-CD150a (PDGFRA) antibody (1:1000 dilution) (ThermoFisher 11–1401-82). Flow cytometric cell sorting and analysis was performed on a SORP FACSAria III (BD). Flow cytometry data was processed using FlowJo version 10.7.1 (BD).