Prokaryotic expression and immunological characteristics of Mycobacterium tuberculosis Rv1886c

Abstract:

Abstract:［Objective］Ag85B (Rv1886c) is secreted during the early stages of infection by Mycobacterium tuberculosis．The purpose of this study was probed into the immune response against Ag85B in vivo．［Methods］Ag85B was prokaryotic expressed and identified，its immunological characteristics were evaluated with indirect-ELSIA，Sandwich-ELISA and enzyme-linked immunospot assay (ELISPOT)．［Results］ Ag85B was mainly expressed in form of inclusion body confirmed by SDS-PAGE． Western blot analysis shows that the fusion protein had good specific reaction with serum of tuberculosis patient and serum of mice immunized with LM-Ag85B．C57BL/6 mice were subcutaneously immunized with Ag85B，the production of IFN-γ and IL-4 in the spleen cells was determined by Sandwich ELISA，the level of IFN-γ was significantly higher than that of IL-4(P＜0.001) in the Ag85B immunization group，it indicated the protein induced Th1-tendency immune responses．Furthermore，purified protein derivative (PPD) used as coating antigen，antibody titer against Ag85B in murine serum reached 1:6400，it was demonstrated that Ag85B could also induce humoral immune responses． Additionally，C57BL /6 mice were intravenously immunized with M．tb H37Rv and bacillus Calmette-Guérin (BCG) respectively for 42 days，M．tb H37Rv group intended to induce Ag85B specific Th1 type immune response，and its ability of eliciting cellular immunity was significantly stronger than BCG group(P＜0.001)．［Conclusion］Ag85B can affectively induce strongly Th1-tendency immune response and humoral response．Whereas，BCG prime vaccination only can elicit low levels of Ag85B240－259 specific immune response．The study laid foundation for probing the pathogenic mechanism，the development of novel vaccine and the establishment of clinical diagnostic method．