Pe conjugated anti mouse f4 80

Flow cytometric analysis of single-cell suspensions from the corneas,¹⁰ (link) spleen,⁵³ (link) bone marrow,⁵⁴ (link) and peripheral blood⁵⁵ (link) was performed as previously described. Single cells were blocked in flow cytometry staining buffer (cat. 00-4222, eBioscience, San Diego, CA, USA) containing anti-mouse CD16/32 antibody (cat. 14-0161-85, eBioscience) for 10 minutes at room temperature. The cells were then incubated for 30 minutes at room temperature with the following antibodies (diluted 1:100): APC-conjugated anti-mouse CD45 antibody (cat. 559864, BD Biosciences, San Jose, CA, USA), Brilliant Violet 421–conjugated anti-mouse CD64 (cat. 139309, BioLegend, San Diego, CA, USA), Brilliant Violet 421–conjugated anti-mouse Lineage Cocktail (cat. 139309, BioLegend), PE-conjugated anti-mouse F4/80 (cat. 565410, BD Biosciences), FITC-conjugated anti-mouse Ly-6G (cat. 551460, BD Biosciences), PerCP-conjugated anti-mouse CD11b (cat. 45-0112-82, Thermo, Waltham, MA, USA), and PE-CY7-conjugated anti-mouse Ly6C (cat. 25-5932-82, eBioscience). Cells labeled with fluorescent antibodies were run on a flow cytometer (BD Canto Plus, BD Bioscience, San Jose, CA, USA), and the resulting data were analyzed using FlowJo flow cytometry analysis software (version 10, Ashland, OR, USA).