Construction and Immunogenicity of the DNA Vaccine of Mycobacterium Tuberculosis Dormancy Antigen Rv1733c

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Abstract

We aimed to construct the DNA vaccine encoding Mycobacterium Tuberculosis (Mtb) dormancy antigen Rv1733c and investigate its immunogenicity in mice. The recombinant plasmid pcDNA-Rv1733c was transfected into P815 cells and its product was detected by indirect immunofluorescence. The mice were immunized once every 2 weeks by intramuscular injection of pcDNA-Rv1733c plasmid for a total of three times. The specific antibodies in the serum of the immunized mice were detected by enzyme-linked immunosorbent assay at the indicted time. Enzyme-linked immunosorbent spot was applied to determine the levels of IFN-γ, IL-2 and IL-4 secreted by splenic lymphocytes. Total cytotoxicity T lymphocyte (CTL) active of the splenic lymphocytes was detected by lactate dehydrogenase assay. Additionally, we analysed the percentages of CD4+ and CD8+ T cells in splenic lymphocytes using flow cytometry. The specific antibody was detected at 2 weeks after the first immunization, and the antibody titre was increased with time which was reached to 1:1600 at 8 weeks. The stimulation index of spleen lymphocytes and the levels of IFN-γ, IL-2 and IL-4 of pcDNA-Rv1733c-immunized mice were both higher than those of saline-immunized mice (< 0.05). However, no difference was found in the percentages of CD4+ and CD8+ T cells and the activity of CTL between the pcDNA-Rv1733c- and saline-immunized mice (> 0.05). So we got the conclusion that the plasmid pcDNA-Rv1733c DNA could induce specific humoral and cellular immunity in mice. Improving the immune effect of Rv1733c by several strategies, such as choosing appropriate immunization route and adjuvant, would be significant for Rv1733c as new tuberculosis vaccine.

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