Specific T-cell Responses to CFP10, an Secreted Antigens of Mycobacterium Tuberculosis Protein, in Chinese HIV Positive Individuals

Abstract

Abstract Objective To construct prokaryotic expression vector of CFP-10 gene, and obtain recombinant protein, and the recombinant CFP-10 protein was taken as stimulus to detect specific T cell responses, to set up a method to faciliate to detect potential TB infection in China. Methods CFP-10 was cloned into inducible prokaryotic expression vector pET-32a (+) and transfected into E. coli BL21 (DE3). After IPTG induction, the product were verified with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot hybridization were carried out to verify the antigenicity; the recombinant CFP-10 protein was taken as stimulus to detect specific T cell responses in HIV (+) persons with or without clinical manifestation of TB diseases, and HIV (-) controls with or without TB diseases. Results The CFP-10 recombinant protein exsited in the form of inclusion body and accounted for 94% in total bacterial protein of E. coli and the molecular weight is 31 kD; Western blot confirmed the recombinant proteins had high antigenicity; our in-house ELISpot-IFN-γ assay with recombinant antigen derived from CFP-10 proteins showed significant higher frequencies in TB patients with or without HIV infection than that in the healthy controls and only HIV (+) group. Conclusions The recombinant CFP-10 genes can be expressed successfully in prokaryotic expression system of E. coli and recombinant proteins with high antigenicity were obtained, which will set foundation for further study on their immunogenicity and bioinformatics. Our results proved that it is indeed true that some HIV positive patient have high frequencies of TB specific T cell responses, which maybe a clue to find latent TB infection in this population.