Em18.3重组蛋白的表达及其免疫诊断特性的研究

目的：在大肠杆菌中表达和纯化截短的泡球蚴rEm18.3抗原重组蛋白,对其免疫诊断特性进行鉴定.方法： IPTG诱导pET41a-Em18.3原核表达质粒,表达和纯化rEm18.3-GST重组蛋白,SDS-PAGE电泳及Western blot法对其进行初步鉴定,ELISA法确定其免疫诊断特性.结果： rEm18.3-GST重组蛋白得到成功表达,SDS-PAGE电泳显示相对分子量为45 kDa.Western blot结果表明,rEm18.3-GST重组蛋白能被AE病人阳性血清识别.对56例多房棘球蚴病（AE）、88例细粒棘球蚴病（CE）、24例其他寄生虫病、24例其他疾病和24例健康者共236份血清进行ELISA检测结果显示,对AE血清诊断的敏感性为10.7%,特异性为99.4%,阳性预测值85.7%,阴性预测值78.1%.结论： rEm18.3-GST免疫诊断价值较低,Em18抗原重要的表位可能位于1-40位氨基酸.

Expression and serological evaluation of diagnostic antigen Em18 for alveolar echinococcosis

Objective: To express and serologically evaluate the rEm18-3 antigen gene of Echinococcus multilocularis for diagnostic purpose. Methods: The constructed plasmid pET41a-Em18.3 was transferred into E. coli BL21 (DE3) for expression. The recombinant proteins were purified with His-Binding agarose by affinity chromatography. Twohundred and thirty six sera were used for evaluating diagnostic value of the recombinant Em18.3. Results: The expressed rEm18.3-GST recombinant protein ould be detected as a band of 45 kDa by SDS-PAGE and Western blot result confirmed that the recombinant protein could specifically react with the serum samples from patients with alveolar echinococcosis (AE). The molecular weight of the expression protein was 45 kDa. Serological evaluation by ELISA was carried out using sera from 56 patients with alveolar echinococcosis(AE), 88 with cystic echinococcosis(CE), 24 from other parasite patients, 44 from non-parasite patients, and 24 from healthy persons (HP). The results showed an overall sensitivity of 10.7% and specificity of 99.4%, positive predictive value 85.7% and negative predictive value 78.1%. Conclusion: The diagnostic value of rEm18.3 was not high