弓形虫多表位基因重组抗原在弓形虫免疫检测中的应用

目的评价弓形虫多表位基因重组抗原在弓形虫感染诊断中的效果，探索多表位抗原在弓形虫免疫检测中的应用前景。方法以Ni—NTAAgarose和割胶电渗两步纯化法。获取弓形虫多表位基因的原核表达纯化产物rMAG，以适宜浓度的rMAG包被ELISA微孔板，分别检测弓形虫急性和慢性感染血清，评价试剂盒检测的敏感性和特异性。结果经Ni—NTAAgarose和割胶纯化，得到了纯度为95．86％的弓形虫多表位重组可溶性抗原。以3μg／ml的抗原浓度包被ELISA微孔板，对兔和小鼠的急慢性弓形虫感染血清进行了检测。结果检测小鼠血清141份，其中慢性感染弓形虫小鼠血清117份、正常小鼠血清24份，检测体系的敏感性为88．88％，特异性为91．67％，一致性为89．36％。检测兔血清24份，其中急性感染弓形虫兔血清18份，正常兔血清6份，阳性血清检出率为94．4％，总一致性为91．5％。结论以弓形虫多表位重组抗原构建的ELISA试剂盒既可以检测弓形虫急性感染血清．又可以检测弓形虫慢性感染血清．具有一定的应用前景。

Application of the Recombinant Antigen of MAG in Immunodiagnosis of Toxoplasmosis

Objective To evaluate the potential application of the recombinant multiple epitope antigen in diagnosis of toxoplasmosis. Methods Purifed recombinant MAG （rMAG） was collected by both immobilized metal affinity chromatography （Ni-NTA Agarose） and electroelution from SDS-polyacrylamide gels. rMAG was used to coat the 96-well plates for the screening of mice antisera against chronic infection of T.gondii and rabbits antisera against acute infection of T.gondii by ELISA. Results rMAG was prepared with a purity of 95.86%. 3 μg/ml of purifed rMAG was used to coat the 96-well plates. A number of 117 sera from 17 mice infected by T.gondii B36 strain and 24 from normal ones were detected by rMAG-ELISA kit. The results showed that the sensitivity and specificity of the kit for detecting chronic infection of T.gondii were 88.88% （104/117）and 91.67% respectively,with total consistency of 89.36%. A number of 24 rabbit sera, in which 18 were derived from rabbits infected with RH T.gondii and the other 6 from normal rabbits, were also detected by rMAG-ELISA kit. 94.4% （17/18） of the positive sera were detected by the kit, with total consistency of 91.5% [ （17＋5）/24]. Conclusion Purified rMAG can be used as good coating antigen to construct ELISA kit for diagnosis of both acute and chronic infection of T.gondii.