利用间接荧光抗体技术对微小隐孢子虫Cp16蛋白进行抗原定位

Localization of Cp16 Protein of Cryptosporidium parvum by Indirect Fluorescent Antibody Technique

Cp16基因是从核糖体展示文库中筛选的微小隐孢子黏附相关基因,为了确定Cp16蛋白是否是微小隐孢子虫卵囊或子孢子表面抗原,构建了Pet-28 a-Cp16原核表达载体,将其转化至BL21大肠杆菌中进行诱导表达,经纯化获得重组蛋白Cp16。将纯化的重组蛋白免疫BAILB/c小鼠,ELISA方法测定IgG抗体滴度。利用间接荧光抗体技术进行抗原定位的初步研究。结果表明:隐孢子虫卵囊表面及子孢子表面富含Cp16蛋白。

Cp16 gene,which was screened from a ribosome display library,is an adhesion gene of Cryptosporidium parvum.A Pet-28a-Cp16 vector was constructed to determine whether Cp16 protein was oocyst or sporozoite surface antigens.The expressive levels were determined by SDS-PAGE and Western Blotting.The recombinant protein Cp16 was purified.The experimental BALBc mice were immunized with the recombinant protein to detect the IgG antibody titers by ELISA.The localization of the protein（the expressive product of the target gene） in C.parvum was determined by fluorescent antibody technique.Results showed that Cp16 protein was localized on the surface of oocysts and sporozoites of C.parvum.