Characterizing virus-induced DCs. (A) Monocytes irradiated with 2000 rad of γ-radiation or nonirradiated cells were infected with live or UV-inactivated IAV. At 18 hours after infection, CD11c⁺CD123⁻ cDCs instead of CD123⁺ monocytes or pDCs among the CD14⁻HLA-DR⁺ DC population were assessed by flow cytometry. (B) Surface CD16 or CD83 expression on the cDCs in panel A from infected and uninfected samples at 18 hours after IAV infection. The results represent 1 of 3 separate experiments. (C) Quantitative real-time PCR analysis of monocyte/DC-related markers in monocytes after 6 hours after IAV infection at a MOI of 10 vs uninfected sample. The results represent the average of 5 subject samples. (D) Intracellular TNF production by IAV-infected PBMCs or monocytes at 6 hours after infection. (E) Levels of surface CD1c, CD141, CLEC4C, and CLEC9A, as well as intracellular CLEC9A, by IAV-infected monocytes at 18 hours after infection. Shaded and open histograms, respectively, represent isotype and specific antibody staining. (F) Monocytes infected for 6 hours with different doses of VSV were cocultured for 7 days with purified CD4⁺ T cells from the same donor at a monocyte/T-cell ratio of 1:5 in the presence of varied concentrations of UV-inactivated IAV antigen. IFN-γ levels in supernatants of the cocultures were determined by ELISA. Data in panels D and F are representative of 3 separate experiments. % Max indicates percent of maximum.