Percp cy5.5 antihuman ifn γ

For analysis of human intracellular cytokine production, PBMCs were stimulated with PMA (50 ng/ml), ionomycin (1 μg/ml), and brefeldin A solution (5 μg/ml) for 5 h. T cells and B cells were stimulated with 0.5 mg/ml purified plate-bound CD3 mAb for 3 days. For intracellular staining, cells were stained with combinations of FITC antihuman CD4 (5 μl/test), PerCP/Cy5.5 antihuman CD19 mAb (5 μl/test), PE antihuman CD24, and FITC antihuman CD38 (Biolegend, San Diego, USA). Cells were washed, fixed, permeabilized (Cytofix/Cytoperm, BD, USA), and stained for the detection of intracellular cytokines with PerCP/Cy5.5 antihuman IFN-γ (5 μl/test), PE antihuman TNF-α (5 μl/test), and APC antihuman IL-10 mAb (5 μl/test) (Biolegend, San Diego, USA). Appropriate isotype controls were used for gate setting for cytokine expression.

FITC mouse IgG1 κ isotype control (400110), PerCP-Cy5.5 mouse IgG1 κ isotype control (400149), PE mouse IgG1 κ isotype control (400114), PerCP-Cy5.5 anti-human CD3 (300430), FITC anti-human CD8a (300906), PE anti-human CD45 (368510), PerCP-Cy5.5 anti-human IFN-γ (506528), FITC Rat IgG2a κ isotype control (400506), PerCP-Cy5.5 Rat IgG1 κ isotype control (400426), FITC anti-mouse CD3 (100204), FITC anti-mouse CD8a (100705), PE anti-mouse CD45 (103106), and PerCP-Cy5.5 anti-mouse IFN-γ (505822) were purchased from Biolegend. PE anti-human PD-L1 (557924), FITC anti-human PD-L1 (558065), FITC anti-human HLA-A2 (343303), FITC Rat IgG2b κ isotype control (400605), PE Rat IgG2a λ isotype control (400635), and PE anti-mouse PD-L1 (558091) were obtained from BD Biosciences.