Expression And Purification Of Recombinant Mycobacterium Tuberculosis CFP32 And Its Application In Serodiagnosis

Objective: To establish a recombinant plasmid of CFP32 of mycobacterium tuberculosis in E.coli,and to assess the serodiagnostic potential of CFP32 .Methods: The cfp32 gene was cloned by PCR, pET21a transformant expressed rCFP32 in E.coli, expression system was optimized and rCFP32 was purified by nickel column affinity chromatography. Enzyme-linked immunosorbent assay (ELISA) detection of human antibody to CFP32 after optimizing detection system.Sera from 160 cases clinical sera were tested for antibodies that recognize rCFP32.Results: The cfp32 gene was cloned from M.tuberculosis H37Rv and the IPTG-induced pET21a transformant expressed rCFP32 inclusion at a band size of 30 KD in E.coli by SDS-PAGE, and rCFP32 was purified by nickel column affinity chromatography.CFP32 was renatured by PBS and The yield of rCFP32 was 3.46 mg/mL.The ELISA assay for CFP32 antibody detection was established , Altogether,63.9% (62 of 97) of TB case patients had detectable antibodies to CFP32 while none of the healthy controls were positive,2 of the other lung disease controls were positive,its total specificity was 96.8%.Conclusions: The recombinant CFP32 proteins was successfully expressed in E.coli and purified,Elisa analysis showed recombinant CFP32 may be a candidate antigen for TB serodiagnosis.