Post-exposure anti-rabies vaccination for individuals who have not previously been immunized against rabies includes a cell culture-derived vaccine and a one time injection of rabies immune globulin. Recent studies have shown DNA vaccinations to be highly effective in rabies pre-exposure experiments, but post-exposure protection has not been achieved. This failure is likely due to the slow onset of DNA vaccine induced antibody production. In an attempt to accelerate the onset of the antibody response, we manipulated variables, such as the route of vaccination and booster frequency. Anti-rabies virus antibody was detected 5 days after the initial DNA vaccination. Using this vaccination protocol and a single non-protective dose of anti-rabies immune serum, we questioned whether mice injected 6 h previously with rabies virus would be protected if a DNA vaccine was substituted for the cell culture-derived human diploid cell vaccine (HDCV). The DNA vaccine protected 87% of the mice (P = 0.00005, compared with unvaccinated control mice). Some 75% of mice receiving HDCV were protected (P = 0.00097, compared with unvaccinated control mice). Mice receiving only anti-rabies immune serum were not protected (P > 0.05 compared to unvaccinated control mice). Thus, post-exposure therapy, substituting a DNA vaccine for HDCV, did not compromise protection against rabies virus.