日本血吸虫可溶性虫卵抗原组分的免疫学特异性分析

Analysis of Immunological Characteristics of Soluble Egg Antigen of Schistosoma japonicum

【目的】分析日本血吸虫可溶性虫卵抗原的特异组分,为建立牛血吸虫病免疫学诊断方法提供数据支持。【方法】用聚丙烯酰胺凝胶电泳(SDS-PAGE)、免疫印迹法(Western-blot)、葡聚糖凝胶层析和斑点酶联免疫吸附试验(Dot-ELISA),对日本血吸虫可溶性虫卵抗原进行了分离、鉴定。【结果】SDS-PAGE分离出10条蛋白条带,依次为100、80、60、55、45、28、27、17、15和13 KDa的小分子量蛋白;采用阳性病牛血清进行Western-blot,共鉴定出有3条特异带,分子量分别为50、28、17 KDa,28和17KDa条带较为明显,而50KDa条带较弱;葡聚糖凝胶层析分离出2个蛋白质高峰,即大分子量蛋白组分峰(Ⅰ峰)和小分子量蛋白组分峰(Ⅱ峰);再经Dot-ELISA鉴定表明,Ⅱ峰中高速离心后的上清蛋白是可用于牛血吸虫病诊断的特异性抗原。【结论】葡聚糖凝胶层析分离出的Ⅱ峰蛋白再经过高速离心后,其上清蛋白是能够用于牛日本血吸虫病诊断的特异性天然蛋白。

【Objective】In order to provide the data for establishment of diagnosis method on bovine Schistosomiasis by analyzing component of soluble egg antigen of Schistosoma japonicum.【Method】In this study,isolation and identification of SEA were performed with SDS-PAGE,western blot,Chromatographic separation with sephadex and Dot-ELISA.【Result】Totally ten proteins bands were detected by SDS-PAGE,including 100,80,60,55,45,28,27,17,15 and 13 KDa;Three proteins bands(50,28,17 KDa)were detected by western blot,strong signals were observed on 28 and 17 KDa protein bands,and weak signal was observed on 50 KDa protein band-two protein peaks[including protein molecules of high molecular weight(Ⅰ peak)and low molecular weight(Ⅱ peak),respectively]were observed with gel filtration on Sephadex G-200;High speed centrifugation was conducted for proteins separation of Ⅱ peak and supernatant proteins were specific antigens for diagnosis of bovine schistosomiasis confirmed by Dot-ELISA.【Conclusion】In summary,after high speed centrifugation,supernatant proteins of Ⅱ peak were specific and natural antigens for diagnosis of bovine schistosomiasis.