Fig. 3.

Loss of virus amplification within hematopoietic cells does not alter generation of the antigen-specific CD8 T-cell response. (A) Northern blot analysis for miR-142 (Upper) and U6 (Lower) of RNA from JAWS II, MDCK, and MDCK142. (B) JAWS II cells infected with NPctrl, NP142t, or UV-inactivated IAV at an MOI of 10 (Right) or pulsed with 1 μM peptide (Left), and 24 h later cells were irradiated and incubated with NP-specific CD8 T-cell hybridomas for 18 h and total number of β-gal⁺ cells enumerated per well. Data are representative of three independent experiments with three to four samples per group. P = 0.003 for NPctrl vs. NP142t. Mice were infected with IAV NPctrl or IAV NP142t and 9 dpi lung cells analyzed by FACS. (C) Representative FACS plots for CD3⁺CD8⁺ cells analyzed for NP₃₆₆ and PA₂₂₄ tetramer expression. (D) Frequency (Left) and total number (Right) of total IAV-specific NP₃₆₆ and PA₂₂₄ tetramer⁺ CD8 T cells. (E) Representative plots for CD3⁺CD8⁺ cells analyzed for intracellular IFN-γ expression following peptide restimulation. (F) Frequency (Left) and total number (Right) of total IFN-γ⁺ CD8 T cells after IAV NP₃₆₆ and PA₂₂₄ peptide restimulation. Data are representative of four independent experiments for tetramer analysis and two independent experiments for peptide restimulation with three to four mice per group.