We previously reported that repeated inoculation of VC1, a macromolecular multicomponent peptide vaccine emulsified with Freund's adjuvant (VC1-F), induced high cytotoxic T lymphocyte (CTL) levels and a substantial level of multivalent antibodies which neutralized various human immunodeficiency virus type 1 (HIV-1) isolates. In the present study, we report that inoculation of VC1-F plus interleukin (IL)-12 expression plasmid can induce a higher antigen-specific CTL response compared to that with VC1-F alone. VC1-F plus IL-12 expression plasmid or VC1-F alone were inoculated to BALB/c mice twice at interval of 2 weeks. Two weeks after the second inoculation, spleen effector cells from these mice were examined. Stronger CTL responses against target cells were observed from the inoculation of VC1-F plus IL-12 plasmid than from that with VC-1F alone, but there was no difference in antibody induction. The inoculation of VC1 plus IL-12 plasmid also produced higher CTL activity than the inoculation of VC1 alone. These augmented CTL activities were not observed using target cells pulsed with non-HIV-specific peptides and different class I haplotype cells. These data demonstrate that co-inoculation of cell-mediated immune potent antigen and IL-12 plasmids can enhance the antigen-specific CTL response. This may be a potential approach for the induction of cellular immunization against HIV-1 and other diseases.