分子佐剂鞭毛素对弓形虫SAG1蛋白抗体应答的影响

发布时间：2018-06-20 15:02

本文选题：弓形虫 + 表面抗原(SAG)　；参考：《福建农林大学学报(自然科学版)》2017年03期

【摘要】：将沙门氏菌鞭毛素和弓形虫免疫优势表面抗原1(SAG1)融合表达,研究其在小鼠上激发的体液免疫应答.首先,采用PCR扩增SAG1片段.其次,将SAG1连接到原核表达载体pET-28a中构建表达质粒pET-28a-SAG1;连接到实验室已有的pET-28a鞭毛素质粒中构建表达质粒pET-28a-F-SAG1;质粒转化至大肠杆菌BL21(DE3)菌株中并进行表达.将纯化后的蛋白免疫小鼠,每隔2周免疫一次,共免疫3次,第3次免疫后15 d取小鼠血清.最后,采用蛋白印迹检测蛋白的大小;用酶联免疫吸附方法检测血清中多克隆抗体的效价.结果表明,诱导出的SAG1蛋白大小为30 ku,F-SAG1大小为70 ku.酶联免疫吸附方法检测的D450 nm均为阴性对照的两倍,表明该多克隆抗体具有较高效价.可见,鞭毛素可以作为分子佐剂促进弓形虫亚单位疫苗的体液免疫应答.
[Abstract]:Salmonella flagellin was fused with Toxoplasma gondii immune dominant surface antigen 1 SAG1 to study its humoral immune response in mice. First, the SAG1 fragment was amplified by PCR. Secondly, the expression plasmid pET-28a-SAG1 was constructed by ligating SAG1 into the prokaryotic expression vector pET-28a, and the expression plasmid pET-28a-F-SAG1 was constructed by ligating into the flagellum diathesis granules of pET-28a. Mice were immunized with purified protein once every 2 weeks for 3 times. The serum of mice was collected 15 days after the third immunization. Finally, the size of protein was detected by Western blotting, and the titer of polyclonal antibody in serum was detected by enzyme-linked immunosorbent assay (Elisa). The results showed that the induced SAG1 protein size was 30 kuo F-SAG 1 and 70 ku. The D450 nm detected by enzyme-linked immunosorbent assay (Elisa) was twice as high as that of the negative control, indicating that the polyclonal antibody had high titer. Therefore, flagellin can be used as a molecular adjuvant to promote humoral immune response of Toxoplasma gondii subunit vaccine.
【作者单位】：福建农林大学动物科学学院/福建省动物药物工程实验室;
【基金】：国家自然基金青年项目(31502058) 福建省自然科学基金面上项目(2015J01075) 福建省中青年教师教育科研项目(JA15159) 大学生创新创业训练计划项目(201610389148、201510389029)
【分类号】：S852.7

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