Abstract: Objective To construct the recombinant plasmid of pcDNA3.1(+)/Gpd-Tp92 containing the outer membrane protein Gpd and Tp92 gene of T.pallidum,and observe its immune response in New Zealand white rabbits.Methods Gpd from pcDNA3.1(+)/Gpd was subcloned into the recombinant plasmid pcDNA3.1(+)/Tp92 by linking reactions to construct pcDNA3.1(+)/Gpd-Tp92,and then the recombinant plasmids were transfected into HeLa cells using liposome.After verifying the Gpd-Tp92 antigen fusion genes could be expressed in HeLa cells by immunocytochemistry,three kinds of DNA vaccines comprising pcDNA3.1(+)/Gpd-Tp92,pcDNA3.1(+)/Gpd,pcDNA3.1(+)/Tp92 were administered respectively to three groups of New Zealand white rabbits.The levels of the specific antibody in the sera of rabbits and the cytokine IFN-γin rabbits spleen lymphocyte culture supemant,after stimulating by special proteins, were quantitatively detected by ELISA.The proliferation response of spleen cells was detected by MTT assay.Results Gpd-Tp92 fusion gene constructed in pcDNA3.1(+) vector could express a fusion protein in Hela cells.The significant specific antibody IgG titers were detected and the highest titer was 1:1024 in rabbits of nucleic acid vaccine groups two weeks after three inoculations.The proliferation response of spleen cells and IFN-γlevels induced were significantly higher in rabbits injected with nucleic acid vaccines than those with pcDNA3.1(+) (P<0.05).There was no obvious difference of the levels of specific antibody induced between the double-gene nucleic acid vaccine group and mono-gene nucleic acid vaccine groups, except the more powerful and lasting cellullar immunologic response in double-gene vaccine group.Conelusion All above results establish a solid basis for future study of the biological activities of Gpd and Tp92, and benefit the development of the syphilitic DNA vaccine.

Key words: Treponema pallidum, Vaccines,DNA, Antibody formation, Immunity,cellular