Table 1.
Vaccination schedule.
Fig 1.
IN and DNA + IN regimens induce the highest serum IgG and IgA responses after three vaccinations.
Mice were vaccinated three times with the indicated vaccine modalities, serum samples taken 1 week after the last vaccination (week 8) were tested for antigen-specific IgG (a) and IgA (b) by ELISA. Data were analysed using one-way ANOVA with Tukey’s post test, * P <0.05, ** P < 0.01, *** P < 0.001.
Fig 2.
Detection of mucosal antigen-specific IgG and IgA responses in the vagina.
Mice were vaccinated three times and vaginal lavage samples were taken 1 week after the last vaccination and tested for antigen-specific IgG (a) and IgA (b) by ELISA. Results are expressed as specific antibody (ng)/total antibody (μg). One-way ANOVA with Tukey’s post test was used to analyse data. * P <0.05, ** P < 0.01, *** P < 0.001.
Table 2.
Mean humoral responses for each vaccination protocol.
Fig 3.
DNA vaccination induces Th1 skewed responses.
Levels of IgG1 and IgG2a were assessed in serum one week after the last vaccination (week 8) by ELISA (a) and IgG1:IgG2a ratios were calculated as a surrogate of Th1/Th2 bias (b).
Fig 4.
The highest frequency of IgA-secreting cells was observed after simultaneous DNA and IN vaccination.
The frequency of antibody secreting IgG (a) and IgA (b) splenocytes were assessed using commercial ELIspot kits. One-way ANOVA with Tukey’s post test was used to analyse data * P <0.05, ** P < 0.01, *** P < 0.001.
Fig 5.
DNA vaccination induces potent IFNγ responses to HIV-1 Env peptides.
Animals were vaccinated three times by the routes indicated. One week after the final vaccination splenocytes were cultured with HIV Env-specific peptide pools, and IFNγ-producing cells were measured using commercial ELIspot kits. Negative responses were assigned a value of 1 to allowing plotting on a log scale. One-way ANOVA with Tukey’s post test was used to analyse data * P <0.05, ** P < 0.01, *** P < 0.001.
Table 3.
Mean cellular responses for each vaccination protocol.
Fig 6.
IN vaccination induces potent IL-2 responses in splenocytes after stimulation with HIV-1 Env peptides.
Animals were vaccinated three times by the routes indicated. One week after the final vaccination splenocytes were cultured with two HIV Env-specific peptide pools, and IL-2-producing cells were measured using ELIspot kits. Negative responses were assigned a value of 1 to allowing plotting on a log scale. One-way ANOVA with Tukey’s post test was used to analyse data * P <0.05, ** P < 0.01, *** P < 0.001.
Fig 7.
DNA and IN vaccinations induces a distinct cytokine profile compared to IM vaccinations.
Cytokines were detected using multiplex bead assay capable of measuring 23 murine cytokines. Differences in cytokine production between groups were observed with selected cytokines IL-2 (a), IL-4 (b), IL-13 (c), IL-17 (d), IFNγ (e), MIP-1α (f), MIP-1β (g) and MCP1 (h). Kruskal-Wallis analysis with Dunn’s multiple comparison test was used to analyse data, * P <0.05, ** P < 0.01, *** P < 0.001.