完整弓形虫P35重组蛋白IgM-酶联免疫吸附测定法检测弓形虫急性感染

为利用重组的完整弓形虫表面抗原P35 GST蛋白对弓形虫感染进行血清学诊断 ,构建了可表达P35 GST的JM10 9细胞株 .采用亲和层析对融合蛋白进行分离和纯化 ,用SDS 聚丙烯酰胺凝胶电泳 (SDS PAGE)和蛋白质印迹 (Westernblot)分析所表达的蛋白质 ,并用纯化的重组蛋白进行IgM 酶联免疫吸附测定法 (IgM ELISA)检测不同病人血清中的抗 P35抗体 .SDS PAGE分析发现P35 GST重组蛋白的大小约 6 0ku ,为一亲水性蛋白 ,蛋白质印迹分析表明该蛋白与弓形虫阳性感染病人的血清有特异性反应 .利用P35 GST为抗原 ,对 6 0例血清进行IgM ELISA分析 ,发现P35 GST可明显区分近期感染和既往感染 ,在弓形虫的诊断上有很大的应用前景 .

Detection of Toxplasma gondii Acute Infection Using Complete Recombinant P35 Surface Antigen and IgM-ELISA

In order to Detect Toxoplasma gondii acute infection in serum samples by complete recombinant P35 surface antigen protein, JM109 cell line which can express P35- GST protein was constructed. Then the recombinant protein was separated and purified using affinity chromatography. SDS- PAGE and Western blot were used to analyse the characters of this recombinant protein. Later P35- GST protein was used as antigen to detect Toxoplasma gondii infection by IgM- ELISA. The result of SDS- PAGE showed that the recombinant protein was about 60 ku and was an hydrophile protein. It reacted specifically with Toxoplasma gondii positive serum in Western blot analysis. 60 different serum samples were detected in IgM- ELISA tests using P35- GST as antigen. It was showed that P35- GST can separate acute infection, chronic infection with IgM and IgG positive, chronic infection with IgM negative and IgG postive significantly. P35- GST was very useful in detecting acute and chronic infection of Toxoplasma gondii . It can be concluded that P35- GST can effectively separate acute and chronic infection using serum samples.