梅毒螺旋体重组蛋白rTp0608在梅毒血清学诊断中的初步应用  

Evaluation the value of Treponema pallidum recombinant protein Tp0608 in serological diagnosis of syphilis

作　　者：高飞 罗宏 谈园 李丽琴 张帅 张莲 GAO Fei;LUO Hong;TAN Yuan;LI Liqin;ZHANG Shuai;ZHANG Lian(Department of Dermatology,the First People′s Hospital of Changsha,Changsha 410005,China)

机构地区：[1]长沙市第一人民医院皮肤性病科,长沙410005

出　　处：《实用医学杂志》2019年第6期973-976,共4页The Journal of Practical Medicine

基　　金：湖南省科技厅一般项目(编号:2014TT2025)

摘　　要：目的建立以梅毒螺旋体重组蛋白Tp0608(rTp0608)为抗原的酶联免疫吸附试验(ELISA),进而评价其在梅毒血清学诊断中的价值。方法构建原核表达系统,镍离子金属鳌合亲和层析(Ni-NTA)纯化rTp0608,Western blotting鉴定目的蛋白并分析其免疫反应性。以该蛋白为包被抗原建立ELISA,检测梅毒患者及潜在交叉感染患者血清,并同梅毒螺旋体颗粒凝集实验(TPPA)进行比较。结果 Western blotting表明该蛋白能与梅毒阳性血清发生明显结合,但不能与健康人群及潜在交叉感染患者血清发生反应。rTp0608-ELISA阳性检测率为97.27%,与金标准的一致性为98.1%。结论 rTp0608-ELISA在检测梅毒患者血清时具有较高的灵敏性,且与金标准一致性较好。Objective We established enzyme linked immunosorbent assay(ELISA)based on recombi nant protein Tp0608 to evaluate the effect on syphilis serological diagnosis. Methods The full length oftp0608 gene was amplified by polymerase chain reaction(PCR) and prokaryotic expression system was constructed. Sodi um dodecylsulphate polyacrylamide gel electrophoresis(SDS PAGE) was used to analyze the expression level of the recombinant protein. Nickel nitrilotriacetic acid( Ni NTA) affinity chromatography was utilized to purify recom binant protein Tp0608. Western blotting was performed to identify the protein and detect the reactivity between the recombinant antigen and the serum samples which were from syphilis patients and individuals with potential cross infections. ELISA was established based on the antigen to determine the diagnostic potential using serum samples from 30 healthy individuals,5 leptospirosis patients,5 hepatitis B patients,5 hepatitis C patients and 110 syphi lis patients. The κ coefficients were used to compare the concordance between gold standard and the Tp0608 based ELISA. Results The purified recombinant protein Tp0608 had strong conjugation reactions to the sera with posi tive antibody againstTreponemapallidum, while there were no reactions observed between the recombinant antigen and serum samples from 5 healthy individuals,2 leptospirosis patients,2 hepatitis B patients,2 hepatitis C pa tients and 10 syphilis patients. The positive detection rate of Tp0608 based ELISA was 97.27%,which was similar to that of gold standard. Moreover,the ELISA was in perfect agreement with the gold standard with a κ value of 0.95. Conclusions ELISA based on recombinant antigen Tp0608 had high sensitivity in diagnosis of syphilis pa tients. The antigen can be a promising serodiagnostic candidate.

关 键 词：梅毒螺旋体 梅毒 rTp0608 血清学诊断 酶联免疫吸附试验 

分 类 号：R759.1[医药卫生—皮肤病学与性病学]