嵌合双串联OVA T细胞表位的RHDV VP60蛋白免疫特性研究  

Immunogenicity of Chimeric RHDV VP60 Protein Carrying Double CD_8^+ T Cell Epitopes of OVA

作　　者：宋艳华[1] 张燕[1] 胡波[1] 范志宇[1] 魏后军[1] 刘星[1] 黄兵[2] 薛家宾[1] 徐为中[1] 王芳[1] 

机构地区：[1]江苏省农业科学院兽医研究所农业部动物疫病诊断与免疫重点开放实验室国家兽用生物制品工程技术研究中心,江苏南京210014 [2]山东省农业科学院家禽研究所,山东济南250023

出　　处：《安徽农业科学》2015年第32期89-92,共4页Journal of Anhui Agricultural Sciences

基　　金：江苏省自然科学基金项目(BK20140740);现代农业产业技术体系建设专项(CARS-44);山东省现代农业产业技术体系项目(SDAIT-18-011-06)

摘　　要：为研究RHDV VP60作为载体容纳外源片段以及递呈外源T细胞表位的能力,将双串联卵清蛋白T细胞表位插入N端、C端以及替代N端氨基酸的方式获得3种嵌合蛋白,进行免疫试验。3种嵌合蛋白分别命名为DN1、DN2和DC,将蛋白进行浓缩、纯化及定量,利用透射电镜观察VLPs的形成,发现均可形成形态大小与VP60相似的VLPs。将3种嵌合蛋白分别腹腔注射C57BL/6雌性小鼠,利用间接ELISA检测首免后0、4和6周小鼠血清样品中VP60特异性抗体,结果表明嵌合蛋白组与VP60组均能够诱导产生较高水平的抗VP60特异性抗体,2组间无显著差异。首免后6周,利用合成的OVA多肽刺激脾淋巴细胞,ELISPOT检测特异性IFN-γ的分泌水平,结果表明3种嵌合蛋白均能诱导特异性IFN-γ的分泌。该研究扩展了VP60作为载体可携带的外源片段的耐受性及有效递呈T细胞表位的能力,为VP60-VLPs展示系统的研究提供了理论依据。Three chimeric RHDV-VLPs incorporating double CDs＋ T-cell epitopes derived from chicken ovalbumin （OVA） were constructed. Two integrations were performed by replacing aa. 2 - 13 （DN2） and 302 - 309 （DC） of VP60 with OVA epitopes respectively, and another one was generated by inserting the epitopes into N-terminus of VP60 （ DN1 ）. Three chimeric proteins were expressed in baculovirus system and self-assembled into VLPs correctly. Three chimeric proteins could induce high levels of anti-VP60 specific antibody and specific IFN-γ production in the murine model. The anti-VP60 specific antibody titers of chimeric proteins groups were comparable with that in group VP60. Chimeric proteins groups showed significant higher productions of specific IFN-γ than that in group VP60. Our result indicated that insertions or replacement of foreign epitopes at the specific locations of VP60 could effectively induce cellular immune responses. The manuscript should be very valuable for using RHDV-VLPs as a carrier for foreign sequences.

关 键 词：RHDV 衣壳蛋白 OVA T细胞表位 嵌合蛋白 免疫原性研究 

分 类 号：S858.29[农业科学—临床兽医学]