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目的研究非融合表达重组抗原用于棘球蚴病免疫诊断的效果。方法将泡球蚴主要表面抗原基因编码序列克隆于原核非融合表达载体pQE30(+)并进行表达。以十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)及免疫印迹(WB)试验对重组抗原鉴定。亲和层析纯化重组抗原,用于酶联免疫吸附法(ELISA)检测患者血清特异性抗体。结果SDS-PAGE及WB分析显示泡球蚴主要表面抗原基因在大肠埃希菌内得到了高效非融合表达。以此重组抗原进行ELISA试验,检测棘球蚴病68例,阳性率达97．1％,检测囊虫等其他感染性疾病80例及健康人血清20份,阴性符合率达98％。结论泡球蚴主要表面抗原基因在大肠埃希菌中获得了高效非融合表达,以该重组抗原建立的ELISA试验,对棘球蚴病诊断具有较高的敏感性和属特异性。
Objective To study the effect of non-fusion expression of recombinant antigen on immune diagnosis of hydatid disease. Methods The coding sequence of major surface antigen gene of Cysticercus cellulosae was cloned in prokaryotic non-fusion expression vector pQE30 (+) and expressed. The recombinant antigens were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting (WB). Purification of recombinant antigen by affinity chromatography for the detection of serum-specific antibodies by enzyme-linked immunosorbent assay (ELISA). Results SDS-PAGE and WB analysis showed that the major surface antigen genes of Cysticercus cellulosae were highly non-fusion in Escherichia coli. ELISA was carried out using this recombinant antigen, detecting 68 cases of hydatid disease, the positive rate of 97.1%, detecting 80 other cases of cysticercosis and other infectious diseases and 20 healthy human serum, the negative coincidence rate of 98%. Conclusion The major surface antigen gene of Echinococcus granulosus was highly efficient and non-fusion in Escherichia coli. The ELISA assay established by the recombinant antigen was highly sensitive and genus-specific for the diagnosis of hydatid disease.