In vitro induction of B7-H6 cell-surface expression on monocytes. Flow cytometric analysis of CD45⁺CD14⁺CD19⁻CD3⁻ monocytes, gated from freshly isolated PBMCs, (A) left untreated or (B) after stimulation for 48 hours with TNFα, LPS, flagellin, or IL-1β. B7-H6 expression was analyzed by flow cytometry with directly conjugated 17B1.3 anti–B7-H6 mAbs (black histograms) or mIgG1 isotype control (gray histograms). Data are representative of at least 3 independent experiments. MFI stim represents the value of the MFI obtained with anti–B7-H6 mAbs minus the MFI obtained with mIgG1 isotype control. (C) PBMCs were left untreated or treated with Poly IC, TNFα, LPS, flagellin, or IL-1β at indicated time points. B7-H6 cell-surface expression was assessed by flow cytometry and fold change in B7-H6 expression was quantified by dividing the MFI of treated samples by that of untreated cells at each time points. Data correspond to a pool of at least 3 independent experiments. Statistical analyses were performed using the 2-way ANOVA test with Bonferroni correction. *P < .05; **P < .01; and ***P < .001. ANOVA, analysis of variance.