Figure 3.

ZIKV-specific CD8+ T cell epitope identification in the acute and memory phases of infection. (A–D) C57BL/6J mice (n between 3 and 5) were injected IV with 10⁵ FFU of ZIKV bled or harvested at the timepoints indicated (A) The functional avidity of the CD8 T cell epitopes identified. On day 8 post infection splenocytes were harvested and stimulated with the indicated peptides (E₂₉₄, E₂₉₇, and NS2b₁₄₇₈) with the concentrations listed on the y-axis in the presence of brefeldin A. Cells were gated using a lymphocyte gate, CD19⁻, CD4⁻, CD8⁺, and were functionally analyzed by expression of IFNγ. Data is presented as the normalized maximal percent of CD8+ T cells that produced IFNγ in response to stimulation). (B) Representative flow plot comparing tetramer binding frequency to IFN-γ response for E₂₉₄, E₂₉₇, and NS2b₁₄₇₈. On day 8 post infection splenocytes were harvested and either stimulated with the E₂₉₄, and E₂₉₇ peptides or stained with peptide specific tetramers. (C) Expression hierarchy of E₂₉₄, NS2B₁₄₇₈, and E₂₉₇ during the acute infection. On day 8 post infection splenocytes were stimulated with E₂₉₄, NS2B₁₄₇₈, or E₂₉₇ peptides in the presence of BFA and the proportion of the CD8+ T cell response dedicated to each of epitope was determined. (D) Functional response to E₂₉₄ is preserved in memory. C57BL/6J mice were injected IV with 10⁵ FFU of ZIKV and on day 0, 5, 8 15, and 45 mice were bled and were stimulated and stained as described above. Data is presented as the percent of CD8+ T cells that produced IFNγ and TNFα in response to stimulation. Data is from a single experiment (n = 6). Asterisks indicate values that are statistically significant (^**p < 0.005) as determined by Mann-Whitney test.