Figure 1: Immunization of rhesus macaques with plasmid DNA encoding MCP3-gp120 fusion primed viral-reactive T-cell immunity, which was further boosted by HIV-1 envelope peptide-cocktail vaccine. The DNA priming schedule included four rounds of immunization at 1-month intervals. Two rhesus macaques were immunized with 20 g of the plasmid DNA by gene gun injection at weeks 0, 4, 8, and 12. The boosting schedule included a peptide-cocktail vaccine that contains six highly conserved peptides from the HIV-1 envelope protein. After a rest period of 20 weeks following the final round of DNA vaccination to allow for the establishment of memory T cells, the peptide-cocktail boosts were administered by intranasal route along with a mutant cholera toxin on Weeks 32 and 37. Peripheral blood mononuclear cells collected after DNA vaccination revealed viral-reactive T-cell immunity as evidenced by the cell proliferation (a) and IFN- producing cells (b) in response to in vitro stimulation with cell-free, heat-inactivated SHI antigen-specific T-cell immunity was boosted by the peptide-cocktail vaccine, especially in #J160 showing significant elevation of IFN- producing cells in response to heat-inactivated SHI) (b). The immunogenicity of the peptide vaccine was confirmed by analysis of peptide-specific T-cell immunity (c). Vaccine-induced cytokine production was examined using LINCOplex multiple cytokine luminescent assay. Among the cytokines assayed, IL-6 was predominantly detected in both macaques after DNA vaccination at Week 12. Virus-reactive IL-6 secretion was more apparent in monkey #J160, particularly during the period of peptide-cocktail boost.