Anti f4 80 percp cy5

Single-cell suspensions were generated from spleen and liver of tumor-bearing mice. The following anti-mouse antibodies were used: anti-CD45-BUV395, anti-CD3E-PE/CF594, anti-CD44-PE, anti-CD4-APC/H7, anti-CD8-APC, anti-CD69-FITC, anti-CD86-FITC, anti-CD11c-PE/Cy7, anti-Gr1-PE, anti-CD11b-APC, anti-CD19-PE/Cy7, anti-NK1.1-PerCP/Cy5.5 (BD Bioscience, San Diego, CA), anti-Ly6C-BV421, anti-Ly6G-AF700, anti-CD4-FITC, anti-CD62L-PerCP/Cy5.5, anti-NKG2D-FITC, anti-F4/80-PerCP/Cy5.5, anti-IFN-γ-PE and anti-TNF-α-PE/Cy7 (Biolegend, San Diego, CA). Anti-mouse CD16/32 FcR block antibody was purchased from Biolegend (San Diego, CA). For intracellular cytokine staining, cells were stimulated with Cell Activation Cocktail (Biolegend, San Diego, CA) for 4 hours and stained with surface antibodies. Then cells were fixed, permeabilized and stained with intracellular cytokine antibodies. Cells were analyzed using a FACS Canto II flow cytometer (BD Biosciences, San Jose, CA).^{29 (link)} Data were analyzed using FlowJo software (Tree Star, Ashland, OR).