Cloning, expression and biological activity of Mycobacterium tuberculosis fusion antigen ESAT-6 and CFP-10 / 军事医学

ABSTRACT

Objective To clone and express Mycobacterium tuberculosis fusion antigen ESAT-6 and CFP-10 ( EC) and to evaluate the biological activity of the fusion antigen EC in inducing specific cytokines secretion from THP -1 cells. Methods The fusion antigen EC gene was cloned into pET-30 a prokaryotic expression vector and expressed highly in E.coli BL21.Then, the THP-1 cells were stimulated with purified fusion antigen EC of different concentrations (10 and 20 μg/ml).Culture supernatants were collected after 12 h and 24 h, respectively.The secretion levels of IL-2, IL-4, IL-6, IL-8, IL-10, GM-CSF, TNF-αand IFN-γin THP-1 cell culture supernatants were detected using Bio-PlexProTM Assays kit.Results The M.tuberculosis fusion antigen EC was cloned and expressed successfully .The secretion levels of IL-6, IL-8 and TNF-αin EC infected THP-1 cells were significantly higher than those in THP-1 cells (P<0.05).The secretion levels of other cytokines did not change significantly .Conclusion The obtained M.tuberculosis fusion antigen EC has biological activity in inducing the THP-1 cells to secrete a higher level of IL-6,IL-8 and TNF-α.