Analysis of the Gag-specific CD8⁺ T Cells in the spleen after NDV-Gag prime and heterologous boost.

<p>(A) Schedule of prime-boost vaccinations. Mice were primed intramuscularly on Day 0 and 6. 35 days later the mice were boosted. Mice mock-immunized with PBS were included as a negative control. 5 days after boost, mice were euthanized and spleens harvested for analysis of the cellular response. (B–D) Each point is representative of splenocytes from one mouse (n = 5 per group). (B) The quantity of activated HIV-1 Gag-specific CD8⁺ T cells in the spleen was analyzed by flow cytometry and the percentage of cells are shown. Activated cells were determined by gating on CD62L^lo cells and HIV-1 Gag-specific cells were determined by tetramer staining against the H2^d restricted AMQMKLETI epitope. (C–E) The functionality of the CD8⁺ T cells was measured by intracellular cytokine staining for (C) IFNγ⁺, (D) TNFα⁺, and (E) IFNγ⁺TNFα⁺ cells after stimulation of the cells with AMQMKLETI peptide. Statistical analysis was performed using One-WayANOVA. Results shown are presented as the mean. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.