梅毒螺旋体TP15抗原在梅毒诊断中的应用价值

The application value of Treponema pallidum TP15 antigen in the diagnosis of syphilis

目的评估梅毒螺旋体TP15抗原在梅毒血清学诊断中的应用价值。方法聚合酶链式反应(PCR)扩增获得梅毒螺旋体TP15基因片段,构建TP15基因的原核表达载体,SDS-PAGE检测TP15蛋白表达水平,Ni-NTA亲和层析法纯化TP15蛋白,Western blot鉴定其免疫反应性;使用TP15重组蛋白建立间接酶联免疫吸附试验(ELISA),检测44份梅毒螺旋体颗粒凝集试验(TPPA)阳性血清和44份TPPA阴性血清,评价TP15-ELISA与TPPA的符合率。结果PCR扩增获得约0.5 kb的基因片段。成功构建了原核表达载体pET28b-TP15。TP15在大肠杆菌中以包涵体形式表达,分子量约20 kDa。Western blot结果表明TP15蛋白能够与梅毒TPPA阳性血清发生特异性反应,与TPPA阴性血清不发生反应。TPPA与TP15-ELISA检测梅毒特异性抗体的符合率比较,差异无统计学意义(P>0.05)。结论TP15抗原在梅毒血清学诊断中具有潜在的临床应用价值。

Objective To evaluate the value of treponema pallidum TP15 antigen in serological diagnosis of syphilis.Methods The TP15 gene fragment of Treponema treponema was amplified by polymerase chain reaction(PCR),and the prokaryotic expression vector of TP15 gene was constructed.The expression level of TP15 protein was detected by SDS-PAGE,the TP15 protein was purified by Ni-NTA affinity chromatography,and its immunoreactivity was identified by Western blot.Indirect enzyme-linked immunosor-bent assay(ELISA)was established by using TP15 recombinant protein,44 Treponema pallidum particle agglutination(TPPA)positive serum and 44 TPPA negative serum were detected,and the coincidence rate between TP15-ELISA and TPPA was evaluated.Results About 0.5 kb gene fragment was obtained by PCR amplification.The prokaryotic expression vector PET28B-TP15 was successfully constructed.TP15 is expressed as an inclusion body in E.coli with a molecular weight of about 20 kDa.Western blot showed that TP15 protein could react with TPPA positive serum,but not with TPPA negative serum.There was no statistically significant difference in the coincidence rate between TPPA and TP15-ELISA in detecting syphilis-specific antibodies(P>0.05).Conclusion TP15 antigen has potential clinical value in serological diagnosis of syphilis.