Anti cd45 v450

Blood and BM samples from killed mice were blocked for 15 min at 4 °C in PBS containing 5% BSA and a 1:100 dilution of anti-CD16/CD32 (24g2, BD Pharmingen, 553142). Samples were then stained for 30 min at 4 °C with anti-CD11b 647 (eBiosciences, 51-0112-82) or biotinylated (BD Pharmingen, 51.01712 J), anti-CD45 V450 (eBiosciences, 48-0451-82), anti-Ly6C FITC (BD Bioscience, 553104) or APC (BD Pharmingen, 560595), anti-CCR5 PE (eBioscience, 12.1951-12), and anti-CCR2 (Biolegend, 150607) primary antibodies and with streptavidin PE (BD Bioscience, 554061) secondary reagent. Erythrocytes in blood samples were lysed with FACS Lysis Solution (BD Biosciences, 349202) for 7 min at room temperature. Before gating, granulocytes were excluded by FCS/SSC. Peritoneal macrophages were collected on the indicated days after TG administration and blocked with BSA/anti-CD16/CD32. Macrophages were then stained for 30 min at 4 °C with anti-CD11b 647 (eBiosciences, 51-0112-82), anti-CD45 V450 (eBiosciences, 48-0451-82), anti-F4/80 Pe-Cy7 (Biolegend, 123114), anti-AIM (GeneTex, GTX37448), and anti-CD36 (Cascade BioScience, ABM-5525) antibodies; for quantification of dead cells, Hoechst 33258 (Sigma, 861405) was added 5 min previous to flow cytometry analysis. Data were acquired in a FACSCanto III cytometer (BD) and analyzed using FlowJo software (Tree Star).