T cell responses induced by tested vaccine candidates.

(A) Mice (n = 3 per group) were immunized once with 5 μg of DREP-C-ENV or 5 μg of DREP-C-GPN alone, or both constructs either mixed or given at separate sites. DREP constructs were given i.d. followed by EP. Splenocytes were assayed 10 days post-immunization with IFN-γ Elispot using peptides PADPNQEM (Env), VGPTPVNI (Pol1) and YYDPSKDLI (Pol2). (B-D) DREP-C-ENV and DREP-C-GPN induce T cells and antibodies that are boosted by multiple administrations. Mice (n = 5 per group) were immunized by i.d. EP one to three times with 5 or 0.1 μg of each DREP-C-ENV and DREP-C-GPN given at separate sites. One group was given DNA-C (5 μg of each construct) 3 times i.m. Boost immunizations were given with a 3 week interval between each administration. (B) Serum was assayed with ELISA for anti-gp140 IgG antibodies 3 weeks after the last immunization. Responses are shown as box plots, with whiskers representing 5–95 percentiles. (C) Splenocytes were assayed with IFN-γ Elispot 10 days and 3 weeks after last immunization using the peptides described above. (D) ICS for CD107a, IFN-γ, IL-2 and TNF-α was performed on splenocytes 10 days after the last immunization. The percentage of CD8+ T cells expressing a certain number of cytokines is shown. Specific markers expressed are shown in S2 Fig. Statistical analyses were performed to compare mice given a different number of administrations of the same dose of DREP-C. In addition, mice given the same number of administrations but with different doses/regimens were compared in separate statistical comparisons. Abbreviations: D-ENV, DREP-C-CN54ENV; D-GPN, DREP-C-ZM96GPN. * P < 0.05; ** P < 0.01