梅毒螺旋体抗原TpN17和TpN44.5表位的融合表达及其意义

Cloning and Expression of Treponema Pallidum Antigen TpN17 and an Epitope of TpN44.5 and Clinical Application

目的 :克隆并融合表达梅毒螺旋体特异性抗原TpN1 7和TpN4 4 .5的一个表位并应用于临床检测梅毒感染。 方法 :从梅毒基因组扩增出TpN1 7的全长基因 ,在其 5′端用聚合酶链反应 (PCR)加入TpN 4 4 .5的一个表位DNA片断 ,插入pGEX4T 2表达载体中 ,以亲和层析法纯化重组融合蛋白。将重组融合蛋白作为抗原制备酶联免疫吸附试验检测试剂盒 ,对正常人群、梅毒患者血清进行检测。 结果 :纯化后获得了相对分子质量为 4 5 0 0 0的特异性抗原。ELISA检测结果表明 :梅毒患者血清结果均阳性 ,正常人群血清均阴性。 结论 :本研究获得的TpN1 7和TpN4 4 .5融合抗原具有更好的灵敏度和特异性 ,为临床检测梅毒感染提供了新思路。

Objective: To investigate the cloning and expression of Treponema pallidum(TP) specific antigen TpN17 and an epitope of TpN 44.5 and the clinical application of this fusion antigen. Methods: TpN17 gene was amplified from the genome of TP and an epitope of TpN44.5 was spliced to the 5′ end of TpN17 gene.This modified TpN17 gene was cloned into the expression vector pGEX4T-2. The recombinant fusion antigen was purified by affinity chromatography and then an enzyme-linked immunosorbent assay (ELISA) kit was prepared. With this kit, the sera of 10 normal persons and 10 TP patients were tested. Results: The molecular weight of the purified fusion antigen was 45 000. Tested with ELISA,10 serum samples of the TP patients were positive and another 10 of the normal persons were negative. ELISA equipped with the GST-epi-TpN17 antigen showed higher sensitivity and specificity as compared with routine methods. Conclusion: The recombinant fusion TP specific antigen GST-epi-TpN17 was suitable for the preparation of ELISA kit in clinical examinations.