[Prokaryotic Expression and Immunogenicity Analysis of a Fusion Protein Containing Cell Epitopes of 2660, 2460, 3875 and 3804 Genes].

Objective: To analyse the immunogenicity of a fusion protein containing cell epitopes of genes 2660, 2460, 3875 and 3804, and to evaluate the feasibility of using it as a novel target antigen for developing multi-stage TB vaccines.

Methods: Cell epitopes of 2660, 2460, 3875 and 3804 were fused in series to form a new antigen gene (named ). Then was cloned into the prokaryotic expression vector -Blunt E1. The fusion protein msv was expressed by -Blunt E1 under the induction of isopropyl-β-d-thiogalactoside (IPTG). Purified the protein by affinity chromatography and identified the protein by SDS-PAGE and Western blot. To evaluate the immunogenicity of the protein, the mice were immunized with the purified fusion protein, and the titer of the antibody in mice serum was evaluated by ELISA. Besides, splenocytes of immunized mice were separated and splenocytes proliferation was determined under the stimulation of the protein.

Results: The prokaryotic expression plasmid carrying gene was constructed successfully and msv protein could be expressed by the plasmid under the induction of IPTG. SDS-PAGE and Western blot results confirmed that a purified protein (relative molecular mass was 41.3×10) was obtained. ELISA result indicated that the titer of the antibody in msv immunized mice serum was about 1:81 920.The spleen lymphocyte proliferation assay showed that after immunization with msv protein, significant proliferation of antigen-sensitized lymphocytes was observed.

Conclusions: The fusion protein msv was successfully expressed and purified, which can induce humoral and cellular immunity in mice. It may be used as an antigen component for the development of TB vaccine in the future.