结核分枝杆菌rESAT-6-CFP-10融合蛋白的制备及其血清学诊断价值

Production of recombinant fusion protein rESAT-6-CFP-10 and its application in tuberculosis serodiagnosis

目的 表达、纯化结核分枝杆菌rESAT-6-CFP-10（rEC）融合蛋白,评价其在结核病血清学诊断方面的应用价值.方法 用镍离子螯合亲和层析柱纯化融合蛋白,用ELISA法检测血清样本中的抗结核抗体,以37例PPD阴性患者正常血清A492＋2s为正常限值,评价rEC诊断的特异性及灵敏度.结果 rEC融合蛋白在大肠埃希菌中以可溶性非包涵体形式表达,表达浓度为0.19 mg/mL.rFC检测抗结核抗体在PPD阳性健康人群中的特异性为100%（30/30）,在痰涂片或细菌培养阳性和两法均阴性的结核病患者中灵敏度分别为75.56%（34/45）和67.3]%（35/52）.结论 结核分枝杆菌rEC融合蛋白以可溶性形式高效表达,具有较强的免疫反应性和较高的血清学诊断价值.

Objective To express and purify rESAT-6-CFP-10（ rEC） fusion protein of Mycobacterium tuberculosis H37Rv and evaluate its application in tuberculosis serodiagnosis. Methods The rEC fusion protein was purified by nickel-chelate affinity chromatography, and anti-tuberculosis antibody was analyzed by ELISA, the specificity and sensitivity of rEC were evaluated by the positive cut off value determined as A492 ＋ 2s in 37 serum samples from PPD-negative healthy people. Results The rEC fusion protein existed in soluble form in E. coli, and the concentration was 0.19 mg/mL. The specificity in PPD-positive healthy people was 100%（30/30） by anti-tuberculosis antibody detection of rEC, and the sensitivity in smear or germiculture positive and negative tuberculosis patients were 75.56%（34/45） and 67.31%（35/52）, respectively. Conclusions The rEC fusion protein can be highly expressed in soluble form in E. coli. It has good immunoreactivity and serodiagnosis value.