Cd11b pacific blue

Whole venous blood was stained with the following antibodies: CD14-APC, CD16-PE-Cy7, CD86-PE, CD42b-FITC, CD11b-Pacific Blue, CD66b-PE-Cy5, CD56-PE-Cy5, CD123-PE-Cy5 (all BD Pharmingen), and TLR2-FITC (all BD Pharmingen) or permeabilized and stained with Ki67-FITC (BD Pharmingen) for validation studies.

Anti-mouse CD11c-FITC, CD8-PE-Cy7, TCRβ-PE-Cy5.5, CD11b-Pacific Blue, B220-PE-Cy7, CD4-Pacific Blue, Foxp3-APC and CD86-PE antibodies were purchased from BD Biosciences. MHCII-AF700 and CD11c-Pacific Blue were purchased from Biolegend. 2×10⁶ cells were stained in PBS containing 5% fetal calf serum (FCS) and 0.1% w/v sodium azide at 4°C for 30 min, washed with the same buffer and fixed with 1% paraformaldehyde containing 2 mM EDTA. Data were acquired on 4-laser LSR II using FACSDiva software (Becton Dickinson) and analyzed using Flow Jo 9.0 software (Tree Star, OR). For splenic DC subset analyses, TCRβ- CD11c+ cells were gated on and the MHCII and CD86 expression levels evaluated in CD11c+CD8+ and CD11c+CD11b+ cells.