Anti siglecf af647

For the identification of immune cell populations, mouse BALF and lung cells were stained in the presence of counting beads (15.45 μm DragonGreen, Bangs Laboratories Inc., FS07F) with LIVE/DEAD stain (Invitrogen, L23105A) and an antibody mixture of anti-CD45-AF700 (BioLegend, 103127), anti-CD11b-AF594 (BioLegend, 101254), anti-CD11c-Bv605 (BioLegend, 117334), anti-SiglecF-AF647 (BD, 562680), anti-MHCII-APC Cy7 (BioLegend, 107628), anti-Ly6C-Bv421 (BioLegend, 128032), and anti-Ly6G-PerCP-Cy5.5 (BioLegend #127616), each at a concentration of 1:200 in PBS, for 1 h at 4°C. After washing, the cells were stored in 2% paraformaldehyde (PFA, Electron Microscopy Sciences, 15714-S) until analysis on the BD LSRII (BD Biosciences) using FACSDiva v9. Flow cytometry was analyzed with FlowJo v10. Mouse BAL and lung cells were identified as follows:
alveolar macrophages: CD45⁺CD11b^+/−SiglecF⁺CD11c⁺;
monocytes: CD45⁺CD11b⁺SiglecF⁻MHCII⁻CD11c⁻Ly6G⁻Ly6C^+/−;
neutrophils: CD45⁺CD11b⁺SiglecF⁻ MHCII⁻CD11c⁻Ly6G⁺Ly6C^+/−.