Stromal vascular cells (SVCs) isolation and FACS analyses were performed as previously described61 ,62 . Briefly, epididymal fat pads were weighed, rinsed three times in PBS, and then minced in 1% BSA PBS. Tissue suspensions were digested with collagenase (1 mg ml−1, Sigma-Aldrich) for 30 minutes. After centrifugation at 500 g for 5 min, the pellet containing SVCs was incubated with red blood cell (RBC) lysis buffer (eBioscience) for 5 min followed by another centrifugation (300 g, 5 min) and resuspension in 1% BSA PBS. SVCs were incubated with Fc Block (BD Biosciences) for 20 min at 4 °C before staining with fluorescently labeled primary antibodies or control IgGs for 30 min at 4 °C. F4/80-APC FACS antibody was purchased from AbD Serotec (Raleigh, NC); FITC-Cd11b, PE-Cd11c and APC- Cy7-Ly6G antibodies were from BD Biosciences.
F4 80 apc facs antibody
Manufactured by Bio-Rad
The F4/80-APC FACS antibody is a fluorescently labeled antibody that binds to the F4/80 antigen, a cell surface marker expressed on macrophages. It is designed for use in flow cytometry applications to identify and quantify macrophage populations within a sample.
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2 protocols using f4 80 apc facs antibody
1
Isolation and FACS Analysis of Stromal Vascular Cells
2
Isolation and FACS Analysis of Stromal Vascular Cells
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Stromal vascular cells (SVCs) isolation and FACS analyses were performed as previously described61 ,62 . Briefly, epididymal fat pads were weighed, rinsed three times in PBS, and then minced in 1% BSA PBS. Tissue suspensions were digested with collagenase (1 mg ml−1, Sigma-Aldrich) for 30 minutes. After centrifugation at 500 g for 5 min, the pellet containing SVCs was incubated with red blood cell (RBC) lysis buffer (eBioscience) for 5 min followed by another centrifugation (300 g, 5 min) and resuspension in 1% BSA PBS. SVCs were incubated with Fc Block (BD Biosciences) for 20 min at 4 °C before staining with fluorescently labeled primary antibodies or control IgGs for 30 min at 4 °C. F4/80-APC FACS antibody was purchased from AbD Serotec (Raleigh, NC); FITC-Cd11b, PE-Cd11c and APC- Cy7-Ly6G antibodies were from BD Biosciences.
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