Improved immunogenicity and protective efficacy of a tuberculosis DNA vaccine encoding Ag85 by protein boosting

C57BL/6 mice were vaccinated with plasmid DNA encoding Ag85 from Mycobacter ium tuberculosis, with Ag85 protein in adjuvant, or with a combined DNA pri me-protein boost regimen. While DNA immunization, as previously described, induced robust Th1-type cytokine responses, protein-in adjuvant vaccination elicited very poor cytokine responses, which were 10-fold lower than those observed with DNA immunization alone. Injection of Ag85 DNA-primed mice wi th 30 to 100 mug of purified Ag85 protein in adjuvant increased the interle ukin-2 and gamma interferon (IFN-mu) response in spleen two- to fourfold. F urther, intracellular cytokine analysis by flow cytometry; also showed an i ncrease in IFN-gamma -producing CD4(+) T cells in DNA-primed-protein-booste d animals, compared to those that received only the DNA vaccination. Moreov er, these responses appeared to be better sustained over time. Antibodies w ere readily produced by all three methods of immunization but were exclusiv ely of tile immunoglobulin G1 (IgG1) isotype following protein immunization in adjuvant and preferentially of the IgG2a isotype following DNA and DNA prime-protein boost vaccination. Finally, protein boosting increased the pr otective efficacy of the DNA vaccine against an intravenous M. tuberculosis H37Rv challenge infection, as measured by CFU or relative light unit count s in lungs 1 and 2 months after infection. The capacity of exogenously give n protein to boost the DNA-primed vaccination effect underlines the dominan t role of Th1-type CD4(+) helper T cells in mediating protection.