Lack of influenza specific T_CD8+ responses in Batf3^o/o mice.

<p>(A) Pre-gating strategy to identify DC (B–C) DC from the inguinal and mediastinal lymph nodes of wildtype or Batf3^o/o mice on a B6 background were analyzed for their expression of either (B) CD8 and CD205 or (C) CD8 and CD103. Representative plots from 3-pooled mice from two independent experiments are shown. (D–E) B6 and Batf3^o/o mice were infected with PR8. On day 10, the absolute number of influenza specific T cells specific for defined peptide sequences were measured in the spleen (D) or BAL (E). The specific T cell response was elucidated following stimulation without peptides (Nil) or the peptides NP_366–374, PA_224–233, PB1-F2_62–70, or PB1_703–711. Shown is the absolute number of IFNγ⁺ CD8⁺ T cells, calculated using the following equation: cell count x%PI⁻ x%CD8⁺ x%IFNγ⁺. Average is taken from between 5–6 mice per group over two independent experiments and the error shows the SEM.</p

Antigen presentation by DC subsets after influenza infection.

<p>B6 mice were inoculated with PR8 and 3 days post infection the lung draining mediastinal lymph nodes were pooled and DC isolated. (A) Gating strategy for isolation of enriched DC subpopulations: CD8⁺ DC were purified on the basis of expression of CD11c and CD8 (upper left; right gate); CD11c⁺CD8⁻ cells (upper left; left gate) were segregated into CD103⁺CD11b⁻ (upper right; top gate) and CD103⁻CD11b⁺ (upper right; lower gate); and finally CD11c⁻ cells were isolated (upper left; bottom gate). (B) The antigen-specific T cell activation for the T cell line specific for the H-2D^b restricted influenza epitope NP_366–374 was assessed using B6 bone-marrow derived DCs pulsed with NP_366–374 peptide at indicated dilutions in a standard ICS assay for IFNγ. (C) Production of IFNγ by NP_366–374 T cells (5×10⁴) co-cultured for 6 hours with serially diluted DC subsets as identified in (A). Data are representative of two independent experiments, which showed a similar trend.</p

PA_224–33 T_CD8+ can be generated in Batf3^o/o mice.

<p>B6 or Batf3^o/o mice were inoculated intraperitoneally with 2.5×10⁶ LPS-treated, PA_224–233 peptide-pulsed B6 bone-marrow-derived DC. 7 days later, the number of PA_224–233 responding T_CD8+ present in the spleen was determined by ICS. Average is taken from 6 mice per group over two independent experiments and the error shows the SEM.</p