Characterization of the E7-specific CD8+ T cell immune responses in tumor-bearing mice treated with HPV-16 E7 DNA vaccine in combination with DMXAA administered at different time points. (A) Schematic diagram of the immunization regimen of the CRT/E7 DNA vaccine and/or DMXAA administered at different time points, either 3 days before the first vaccination (d-3) or 3 days after the first vaccination (d+3). 5-8 weeks old C57BL/6 mice were challenged with 1 × 105 TC-1 tumor cells subcutaneously, vaccinated with pcDNA3-CRT/E7 DNA vaccine via gene gun delivery and treated with DMXAA as indicated in Figure 4A. One week after last vaccination, splenocytes from tumor-bearing mice were harvested and characterized for E7-specific CD8+ T cells using intracellular IFN-γ staining followed by flow cytometry analysis. (B) Bar graph depicting the number of E7-specific IFNγ+ CD8+ T cells per 3 × 105 splenocytes ± SEM following DNA vaccination +/- DMXAA treatment. The data shown here are from one representative experiment of two performed.