HMDMs were isolated, differentiated, and polarized as described above for 24 h. Polarized cells were treated with 50 nM thioA for 7 h. Cells were detached using accutase solution (Sigma-Aldrich #A6964), washed, and resuspended in FACS wash (PBS, 2.5% FCS, 0.1% sodium azide). Cells were blocked in human Fc Block (BD, #564220) for 15 min, and stained for 30 min on ice with anti-CD14-APC (BD #555399), anti-CD163-PE-CF594 (BD #562670), anti-CD80-BB515 (BD #565008), and anti-HLA-DR-PerCP-CY5.5 (BD #560652) antibodies. After washing, stained cells were resuspended in 1% paraformaldehyde in PBS prior to flow cytometric analysis on a BD LSRFortessa. Data were analyzed using BD FACSDiva software (BD Biosciences, San Jose, CA, USA). Median fluorescence intensity of singlet cells was used to quantify surface marker expression.
Anti cd80 bb515
Manufactured by BD
Sourced in United States
Anti-CD80-BB515 is a fluorescent-conjugated monoclonal antibody that binds to the CD80 cell surface protein. It is designed for use in flow cytometry applications.
Automatically generated - may contain errors
2 protocols using anti cd80 bb515
1
Phenotypic Analysis of Polarized HMDMs
2
Phenotypic Analysis of Polarized Macrophages
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Flow cytometric analyses were performed as previously described [36] . Briefly, MDMs were differentiated for five to six days and polarized as described above. 2.5-5 × 105 cells were resuspended in FACSwash (PBS, 2.5% FCS, 0.1% sodium azide), blocked in human Fc Block solution (#564220, BD Biosciences, San Jose, CA, USA) for 15 minutes, and stained for 30 minutes on ice with anti-CD14-APC (#555399, BD Biosciences), anti-CD163-PE-CF594 (#562670, BD Biosciences), anti-CD80-BB515 (#565008, BD Biosciences) or anti-CD86-BB515 (#564544, BD Biosciences), and anti-HLA-DR-PerCP-Cy5.5 (#560652, BD Biosciences) antibodies. After washing, stained cells were resuspended in 1% paraformaldehyde in PBS prior to flow cytometric analysis on a BD LSRFortessa. Data were analyzed using BD FACSDiva software (BD Biosciences). Median fluorescence intensities of singlet cells were used to quantify surface marker expression.
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