猪囊尾蚴丝氨酸蛋白酶抑制剂B6和4848优势抗原表位的分析与鉴定OA

Aiming at the problems of insufficient exposure of Cysticercus cellulosae serine protease inhibitors B6(Ts-serpin B6)and 4848(Ts-serpin 4848)antigenic epitopes and lack of high specificity,it is proposed to enhance the antigenic specificity by screening key antigenic epitopes.The sequences of Ts-serpin B6 and Ts-serpin 4848 genes were firstly analyzed by bioinformatics epitope analysis to screen out the dominant antigenic epitopes.After designing specific primers,the target gene fragments were amplified respectively by PCR.The purified PCR products were ligated into pET-28a(+)vector and then transformed into Escherichia coli DH5α receptor cells to construct segmented prokaryotic expression vectors for pET28a(+)-Ts-serpin B6 and pET28a(+)-Ts-serpin 4848,which were transformed into E.coli BL21(DE3)to induce express ion,and the target proteins were purified and then identified by SDS-PAGE and analyzed by Western-blot.Six recombinant target proteins,Ts-serpin B6(a),Ts-serpin B6(b),Ts-serpin B6(c),and Ts-serpin 4848(a),Ts-serpin 4848(b),Ts-serpin 4848(c),which existed in E.coli mainly in the form of inclusion bodies,were successfully expressed and purified.The results of Western-blot and ELISA showed that the recombinant proteins Ts-serpin B6(a)and Ts-serpin 4848(a)had stronger reactogenicity and specificity compared with other segmented recombinant proteins.This study clarified that Ts-serpin B6(a)and Ts-serpin 4848(a)are the dominant epitopes of the antigens,which provides a basis for the development of diagnostic reagents specific for C.cellulosae.