Immunogenicity evaluation of Mycobacterium bovis Mb0950c protein and establishment of serological diagnostic method for bovine tuberculosis detection

Abstract:

[Objective] The purpose of this study is to express Mb0950c protein by prokaryotic expression system, and to evaluate its immunogenicity by mouse model, further to establish a serological indirect enzyme-linked immunosorbent assay (ELISA) for clinical detection of bovine tuberculosis. [Methods] The prokaryotic expression plasmid of pET32a-Mb0950c was constructed and transformed into BL21(DE3) to induce protein expression. The immunogenicity of the protein in mice was analyzed by flow cytometry (FCM) and ELISA. And then an indirect ELISA method based on Mb0950c was established and used in clinical trial. [Results] SDS-PAGE and Western blotting showed that Mb0950c protein was successfully obtained and had good immunogenicity. The analysis of FCM showed that Mb0950c protein upregulated the expression of CD69 on the surface of T cells. The results of ELISA showed that the protein could induce the secretion of IFN-γ and IL-4, at the same time, it could induce the body to secrete specific antibodies, and it mainly belongs to IgG1. Total 192 serum samples were detected by indirect ELISA based on Mb0950c and the results showed that the positive coincidence rate, negative coincidence rate and total coincidence rate were 65.7%, 97.9% and 72.4% respectively. [Conclusion] Mb0950c protein is expressed in the prokaryotic expression system and it induces Th1-and Th2-type immune response in mouse models. Based on this protein, an indirect ELISA method for serological detection of bovine tuberculosis has been established.