【摘要】 目的:构建表达结核分支杆菌ESAT6和MPT64融合蛋白,并评价其在血清学诊断中的价值。方法:PCR法扩增esat6和mpt64基因,构建原核表达载体pProEX HTb-EM,表达、纯化ESAT6-MPT64融合蛋白,Western-blot分析其抗原性。用该重组蛋白ELISA法检测结核病患者血清。结果:原核表达的ESAT6-MPT64融合蛋白约40 000,Western-blot证实重组蛋白与ESAT6和MPT64多克隆抗体特异结合。重组蛋白用于检测结核病患者血清,敏感性为67.7%,特异性为82%。结论:重组的ESAT6-MPT64融合蛋白有可能用于结核病免疫诊断试剂的制备。更多还原

【Abstract】 Objective To construct and express ESAT6-MPT64 fused protein of mycobacterium tuberculosis and evaluate its applicability in serological diagnosis of tuberculosis.Methods esat6 and mpt64 genes were amplified by polymerase chain reaction from genome of mycobacterium tuberculosis,and the genes were cloned into expression vector,and recombinant ESAT6-MPT64 protein was got by expression and purification.The antigenicity of the fused protein was analyzed with Western-blot with anti-ESAT6 and anti-MPT64 polyclonal antibodies.The fusion protein was used for serological diagnosis of inpatients.Results The DNA sequences of ESAT6 and MPT64 were identical with that of published by GenBank.The relative molecular mass of the fused protein was about 40 000 with SDS-PAGE analysis.The protein expression was confirmed with Western-blot analysis and the fused protein could react to both anti-ESAT6 and anti-MPT64 polyclonal antibody.The sensibility of the fused protein in serological diagnosis was 67.7% and specificity was 82%.Conclusion The recombinant ESAT6-MPT64 protein may be a potential candidate of diagnosis reagent of tuberculosis.更多还原