Apc cy7 anti cd8 mab clone 53 6

Lung leukocytes, prepared as previously described [38 (link)], were stained with APC-anti-CD3ε mAb (clone 145-2C11, Biolegend, San Diego, CA, USA), Pacific Blue-anti-CD4 mAb (clone GK1.5, Biolegend), APC/Cy7-anti-CD8 mAb (clone 53–6.7, Biolegend), PE/Cy7-anti-NK1.1 mAb (clone PK136, Biolegend) and PE-anti-TCRδ mAb (clone GL3, Biolegend). The cells were also stained with PE-anti-CD11b mAb (clone M1/70, Biolegend), PE/Cy7-anti-Gr-1 mAb (clone RB6-8C5, Biolegend) and Pacific Blue-anti-F4/80 mAb (clone BM8, Biolegend). After washing twice, the cells were incubated in the presence of cytofix/cytoperm (BD Biosciences), washed twice in BD perm/wash solution (BD Biosciences) and stained with FITC-anti-IFN-γ mAb (clone XMG1.2, Biolegend). Isotype-matched IgG was used for control staining. The stained cells were analyzed using a BD FACS Canto^TM II flow cytometer (BD Biosciences). Data were collected from 20,000 to 30,000 individual cells using forward-scatter and side-scatter parameters to set a gate on the lymphocyte or myeloid cell population.

The BALF cells were washed three times in PBS containing 1% fetal calf serum (FCS) and 0.1% sodium azide and then stained with APC-anti-CD3ε mAb (clone 145-2C11; BioLegend), FITC-anti-CD4 mAb (clone GK1.5; BioLegend), APC/Cy7-anti-CD8 mAb (clone 53–6.7; BioLegend), and PE/Cy7-anti-NK1.1 mAb (clone PK136; BioLegend), or FITC-anti-CD11c mAb (clone N418; BioLegend), PE-anti-CD11b mAb (clone M1/70; BioLegend), PE/Cy7-anti-Gr-1 mAb (clone RB6-8C5; BioLegend), and Pacific Blue-anti-F4/80 mAb (clone BM8; BioLegend). Isotype-matched IgG was used for control staining. The stained cells were analyzed using a BD FACS Canto II flow cytometer (BD Bioscience) as previously described^{11 (link),31 (link)–33 (link)}.