【摘要】 目的以纯化的重组弓形虫RH株主要表面抗原SAG1、次黄嘌呤-黄嘌呤-鸟嘌呤磷酸核糖转移酶(HXG-PRT)和腺苷激酶(AK)作为诊断抗原,建立间接ELISA法检测抗弓形虫IgG抗体。方法诱导表达本实验室保种的SAG1/pET28a/BL21、HXGPRT/pET28a/BL21和AK/pET28a/BL-21,获得rSAG1、rHXGPRT和rAK三种重组抗原。分别用不同浓度的上述抗原包被聚苯乙烯酶标板,检测疑似弓形虫感染者血清。结果混合重组抗原ELISA法的最佳rSAG1包被浓度为500ng/ml、rHXGPRT和rAK均为200ng/ml;人血清稀释度为1∶20;血清1∶200稀释仍示阳性;特异性试验的抑制率为72.36%,变异系数11.2%。结论成功表达了弓形虫三种重组诊断抗原,并应用组合抗原(rSAG1+rHXGPRT+rAK)建立了ELISA检测方法,具有高度特异性和敏感性,本试剂具有弓形虫病辅助诊断价值。更多还原

【Abstract】 Objective Three recombinant antigens surface antigen1(SAG1),hypoxanthine-xanthine-guanine-phosphoribosyltransferase(HXGPRT),and adenosine kinase(AK)of Toxoplasma gondii were evaluated for their potentiality in diagnosis of suspected patients with Toxoplasmosis,who had been screened by commercial ELISA kit.Methods The expression vectors of Toxo-SAG1/pET28a /BL21,Toxo-HXGPRT/pET28a/BL21 and Toxo-AK/pET28a/BL-21,which had been stored in our laboratory,were simultaneously induced by IPTG and identified by western blotting.rSAG1,rHXGPRT and rAK were mixed at the ratio of 1∶1∶1 and the optimal working concentrations was optimized.Combination of the three recombinant antigens were used in ELISA for the detection of sera of sixty-one suspected pregnant women,who had been screened by commercial diagnostic kit.Results Working concentrations of rSAG1,rHXGPRT and rAK antigens(ng/ml)were 500,200 and 200 ng/ml respectively.The working dilution of sera was 1∶20 in pregnancies.The sensitivity,specificity and accuracy in human sera,were 1∶200,72.36% and 11.2% respectively.Conclusion This study demonstrates that combination of the recombinant major antigens in ELISA has high sensitivity,specificity and reliability.The method is useful in the serodiagnosis of pregnancies with suspected Toxoplasma infection.更多还原