Objective: Bioinformatics analysis and epitope prediction of Rv3425 in Mycobacterium tuberculosis. Evaluating the diagnostic value of Rv342519-176 recombinant protein and CFP10-ESAT6 fusion protein in tuberculosis (TB). Methods: The gene coding 19～176 amino acid fragment of Rv3425 was amplified from Mycobacterium tuberculosis genome by PCR. Prokaryotic expression vector of Rv342519-176 was constructed and recombinant protein (rRv342519-176) and CFP10-ESAT6 fusion protein(rCFPl0-ESAT6) were expressed and purified. Establishing ELISA testing method with recombinant protein and fusion protein for antigen, and assessing the combined value of two proteins in TB-antibody detection. Results: The rRv342519-176 protein fragment with dominant epitope and rCFP10-ESAT6 were expressed effectively in E. coli. The ELISA result showed certain sensibility(39％) and high specificity (97.5％), when rRv342519-167 was antigen for clinical diagnosis in tuberculosis. The sensibility and specificity were respectively 37％ and 95％, when rCFP10-ESAT6 was antigen. Furthermore, two proteins conjoint analysis by ELISA showed higher sensibility(57％) and specificity(95％). Conclusion: The epitope of Rv3425 was predicted by bioinformatics. The expressed and purified rRv342519-167 and rCFP10-ESAT6 had a certain complementary antigen in TB-antibedy detection, and provided a reference with improving the diagnostic sensitivity in joint diagnostic antigen with keeping high specificity.%目的:对结核分枝杆菌Rv3425蛋白进行生物信息学分析及抗原表位预测,评价Rv342519-176重组蛋白与CFP10-ESAT6融合蛋白在结核抗体检测中的应用.方法:PCR扩增结核杆菌Rv3425蛋白19～176位的编码基因片段,原核表达并纯化Rv342519-176重组蛋白(rRv342519-176)和CFP10-ESAT6融合蛋白(rCFP10-ESAT6),建立以重组蛋白为抗原的ELISA方法,评价2种蛋白在结核抗体检测中的联合应用价值.结果:具有抗原表位的rRv342519-176与rCFP10-ESAT6在大肠杆菌中高效表达;ELISA结果显示,rRv342519-176的敏感性为39%,特异性为97.5%;rCFP10-ESAT6的敏感性为37%,特异性为95%;2种蛋白联合检测,敏感性提高到57%,特异性为95%.结论:生物信息学预测Rv3425的优势抗原表位,表达并纯化的rRv342519-176和rCFP10-ESAT6在结核抗体检测中具有一定的抗原互补性,在保持特异性的基础上可显著提高检测敏感性.
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