[Cloning and expression of outer membrane protein gene Gpd from Treponema pallidum and preliminary studies on its immunogenicity in rabbits].
To construct the recombinant plasmid of Eukaryotic expression containing Gpd gene from Treponema Pallidum and study its immunogenicity in New Zealand White rabbits. Gpd gene was amplified from the genomic DNA of T. pallidum and cloned into appropriate site of pcDNA3. 1 ( + ) vector. After verified that the Gpd antigen gene could be expressed in HeLa cells by Western blot and immunocytochemistry, recombinant plasmids pcDNA3.1 ( + )-Gpd, control plasmid pcDNA3. 1 ( + ) or PBS buffer were administered in three groups of New Zeal and White rabbits. Booster immunizations were employed at 2-week interval for three times. ELISA was used for the quantitative detection of the specific antibody in the sera of rabbits. The proliferation response of spleen cells was detected by MTT assay. The results of the Western blot and immunocytochemistry showed that Gpd gene constructed in pcDNA3.1 ( + ) vector could express a fusion protein with a calculated molecular mass of 41kD in HeLa cells and react with positive blood serum from syphilis patients. The significant specific antibody IgG titers were observed and the highest titer was 1:1024 in rabbits after three times with pcDNA3.1 ( + )-Gpd. The proliferation response of spleen cells were significantly higher than that of rabbits injected with pcDNA3.1 ( + ) (p < 0.05). All above results establish a solid basis for future studying the biological activities of Gpd and benefit the development of the Syphilis DNA vaccine.
Zhao
Fei-jun
FJ
Institute of Pathogenic Biology, Nanhua University, Hengyang, China.
Wu
Yi-mou
YM
Zhang
Xiao-hong
XH
Liu
Shuang-quan
SQ
Yu
Min-jun
MJ
D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics* / Q000276:immunology; D001428:Bacterial Vaccines / Q000276:immunology*; D003001:Cloning, Molecular; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D008213:Lymphocyte Activation; D016133:Polymerase Chain Reaction; D011817:Rabbits; D014210:Treponema pallidum / Q000235:genetics / Q000276:immunology*; D019444:Vaccines, DNA / Q000276:immunology*
D000907:Antibodies, Bacterial; D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; D019444:Vaccines, DNA