Figure 4.

Adenovector-Expressed Antigen Is Effective at Recalling CD8⁺ T Cell Responses from Mice Immunized with Protective Regimens of Sporozoite Vaccines

Target A20 cells were infected at the indicated MOIs with AdPyCSP (either triple CsCl₂ purified adenovector or unpurified cell lysate from adenovector infected cells) and incubated with splenocytes from RAS or SPZ+CQ-immunized mice. Control targets were A20 cells infected with AdNull, AdGFP, and uninfected A20 cells. (A) IFNγ-secreting cells from mice immunized with RAS were measured by ELISpot. (B) CD8⁺ IFNγ⁺ cells from mice immunized with RAS were measured by ICS and flow cytometry. (C) Comparison of Ad-array PyCSP vector (AdgPyCSP) with AdPyCSP, which does not contain the recombination motifs flanking the expression cassette. CD8⁺ IFNγ⁺ cells from mice immunized with RAS were measured by ICS and flow cytometry. (D) Dose-response analysis for efficacy of SPZ+CQ vaccine regimen following immunization with 0, 200, 2,000, or 20,000 P. yoelii sporozoites with chloroquine treatment. Mice were assessed for sterile protection by blood smear. (E) AdPyCSP-infected cells can recall CD8⁺ T cell responses from mice immunized with 2,000 SPZ+CQ. Target A20 cells were infected with cell lysates containing the indicated Ad5 vectors and antigen-specific CD8⁺ T cell responses were measured by ICS and flow cytometry. Error bars indicate the SEM (n = 3). Asterisks indicate statistically significant differences compared with AdGFP (A), A20 (B and C), or AdNull (E) controls (p < 0.05 by ANOVA with the Bonferroni mean-comparisons test).