Immunogenic Study On FMDV Structural Protein VP1 And C-terminal Of VP1 And Establishment Of Indirect VP1-ELISA Diagnose

Foot-and-mouth disease (FMD) is the causative agent of highly infectious and the economically most important animal viral disease in the world. For the contral of the disease, in developed countries, slaughtering of suspected and disease animal is often adopted, which can rapidly break down the disease. Whereas in contrast, in developing countries, systematic control measures based on inoculation of vaccine and detection methods is usually used. Now, ELISA is one of the most economical and effective methods. But conventional ELISA was based on inactive virus and it had the risk of disseminating virus into the surroundings. An alternative to this is to identify type specific gene sequences, express them in safe heterologous system and use them as diagnostic antigens. Moreover, primal antigen site gene of FMDV was expressed in prokaryotic cell and product was purified for inoculating animal. This study will lay a foundation for understanding sub-unit vaccine development.In this experiment, three genes, such as BoIFN-α, FMDV VP1 and C-terminal of VP1 were expressed in E.coli BL21(DE3)respectively, and emulsion vaccines were produced after purified and dialyted, then injected into the mouse by hypodermic multi-sites. T lymphocyte proliferation assays and sera antibody titers were evaluated respectively using MTT method and liquid-phase blocking ELISA. The result showed that recombinant VP1 and C-terminal of VP1 had a capability to induce both humoral and marginal cell-mediated immune responses in mice alone , while the two proteins co-incobated with rBoIFN-αrespectively presented overwhelming roles on the two responses.The complete gene encoding the structural protein of FMDV(VP1) was subcloned into expression vector pPROex-HT,resulting in the fusion expression plasmid.After transformed into E.coli BL21(DE3) and induced by IPTG,the fusion protein was expressed in high level.Western-blot was performed to confirm that the expressed fusion protein could specifically react with anti-serum against FMDV.The fusion protein was further purified and used as an antigen to establish a novel FMDV in pigs ELISA diagnose assay(VP1-ELISA).Comparison between VP1-ELISA and the government standard kit (liquid phase block ELISA) showed the two methods had 96.25 percent agreement by detecting 80 serum samples,indicating that the indirect VP1-ELISA was specific and sensitive.