Leukocytes were suspended in cold PBS (1 × 107 cells/ml), incubated with anti-mouse CD 16/32 (eBioscience, San Diego, CA, 1 μl/ml), then flurochrome-conjugated antibodies were added (0.5 μg/106 cells/0.1 ml) at 4°C for 30 minutes. Flow antibodies used for these studies included anti-F4/80-FITC (clone BM8, eBioscience), anti-Ly6G-PE (clone 1A8, BD Biosciences), anti-Ly6C-PE Cy5.5 and appropriate isotype controls (eBioscience and BD Biosciences). Neutrophils are identified as Ly6G+F4/80− cells, macrophages are identified as Ly6C−F4/80+ cells, and monocytes are identified as Ly6C+F4/80+ cells. Samples were run on an Accuri C6 flow cytometer (BD Biosciences, San Diego, CA). Data were analyzed using Accuri C6 software.
Anti f4 80 fitc clone bm8
Manufactured by Thermo Fisher Scientific
Sourced in Belgium
Anti-F4/80-FITC (clone BM8) is a fluorescently-labeled monoclonal antibody that specifically binds to the F4/80 antigen, which is expressed on the surface of mouse macrophages. This antibody can be used for the identification and enumeration of mouse macrophages in flow cytometry applications.
Automatically generated - may contain errors
Lab products found in correlation
Anti ly6g pe clone 1a8, by BD (3 mentions)
Anti mouse cd16 32, by Thermo Fisher Scientific (3 mentions)
Accuri c6 flow cytometer, by BD (2 mentions)
Accuri c6, by BD (2 mentions)
Anti ly6c pe cy5, by Thermo Fisher Scientific (1 mentions)
Anti cd16 cd32, by BD (1 mentions)
Anti cd8 apc clone 53 6, by Thermo Fisher Scientific (1 mentions)
Anti cd3 pe clone 145 2c11, by Thermo Fisher Scientific (1 mentions)
Pe cy7 anti cd11b clone m1 70, by Thermo Fisher Scientific (1 mentions)
Facscanto flow cytometer, by BD (1 mentions)
Anti cd11b apc clone m1 70, by Thermo Fisher Scientific (1 mentions)
Anti ly6c, by Thermo Fisher Scientific (1 mentions)
Anti ly6g apc clone 1a8, by BioLegend (1 mentions)
6 protocols using anti f4 80 fitc clone bm8
1
Leukocyte Phenotyping by Flow Cytometry
2
Immune Cell Profiling by Flow Cytometry
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Spleen cell surface antigens were analyzed by use of antibodies: Anti-CD3-PE clone 145-2C11, anti-CD4-APC clone RM 4-5, anti-CD8-APC clone 53-6.7, anti-CD11b-PECy7 clone M1/70, anti-Ly6G(Gr-1)-APC clone RB6-8C5, and anti-F4/80-FITC clone BM8 (all eBioscience, San Diego, CA) after blocking of FC antibody-binding by anti-CD16/CD32 (BD Biosciences, Franklin Lakes, NJ) for 10 min. Cells were fixed in PBS with 2% methanol-free formaldehyde and analyzed within 1–3 days on a FACScanto flow cytometer (BD Biosciences). Data analyses and layouts were performed using FlowJo V10 (Tree Star, Ashland, OR).
3
Multicolor Flow Cytometry for Cell Identification
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Cell identification was performed according to a previous study with slight modifications [29] (link). In brief, cells were kept on ice, and all incubations were performed at 4 °C. Fc receptors were first blocked with anti-CD16/32 antibodies (clone 93, eBioscience). Cells (1 × 106) were then incubated with the following fluorophore-labeled antibodies specific for cell-surface components: anti-F4/80-FITC (clone BM8, eBioscience) and anti-CD11b-APC (clone M1/70, eBioscience) in flow buffer (1% bovine serum albumin and 0.09% sodium azide in PBS) for 30 min. After that, cells were washed with PBS and fixed with 4% paraformaldehyde in PBS for 30 min. Finally, cells were washed and resuspended in PBS and stored at 4 °C until analysis by flow cytometry.
4
Leukocyte Immunophenotyping by Flow Cytometry
5
Characterization of Peritoneal Leukocytes
6
Multiparametric Flow Cytometric Analysis
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For flow cytometric analysis, cells were first stained with LIVE/ DEAD V R Fixable Dead Cell Stain (Molecular Probes, Invitrogen, Carlsbad, California) followed by incubation with anti-CD16/ CD32 (clone 2.4G2) to block the FcR, stained with fluorescently labeled antibodies and stored in 1% paraformaldehyde until analysis. The following antibodies were used: anti-CD45.2 Pacific Blue (clone 104, Biolegend, Uithoorn, The Netherlands), anti-LY6G APC (clone 1A8, Biolegend), anti-F4/80 FITC (clone BM8, eBioscience, Halle-Zoersel, Belgium), anti-CD11b PE and FITC (clone M1/70, eBioscience), anti-Gr1 APC (clone RBG8C5, eBioscience) in fluorescence-activated cell sorting buffer (PBS containing 0.25% BSA, 0.05% NaN 3 , 0.5 mM EDTA) for 30 min at 4 C.
HepG2 cells were stained with Annexin V-FITC, 7-AAD, and TO-PRO3 in Annexin V-binding buffer for 10 min at room temperature and immediately placed on ice until flow cytometric analysis.
Data were acquired by means of FACS Canto II and analyzed using Weasel flow analysis package (The Walter and Eliza Hall Institute of Medical Research, Melbourne, Australia).
HepG2 cells were stained with Annexin V-FITC, 7-AAD, and TO-PRO3 in Annexin V-binding buffer for 10 min at room temperature and immediately placed on ice until flow cytometric analysis.
Data were acquired by means of FACS Canto II and analyzed using Weasel flow analysis package (The Walter and Eliza Hall Institute of Medical Research, Melbourne, Australia).
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