牛腺病毒3型六邻体蛋白的截短表达、单克隆抗体制备及初步应用  

Expression of truncated hexon protein and the preparation and preliminary application of monoclonal antibodies against Hexon protein of bovine adenovirus type 3

作　　者：严昊[1] 朱远茂[1] 史鸿飞[1] 王雪枝[1] 蔡红[1] 王姝[1] 马磊[1] 薛飞[1] 

机构地区：[1]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室/大动物传染病研究室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2014年第6期475-478,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：公益性行业(农业)科研专项经费(201003060-04);哈尔滨市科技攻关计划(2012AA6BN020);中央级公益性科研院所基本科研业务费项目(0302012002);国家科技支撑计划子课题(2012BAD12B03-3)

摘　　要：为制备牛腺病毒3型(BAV-3)的单克隆抗体(MAb),本研究采用原核表达系统截短表达了BAV-3六邻体蛋白(Hexon),并以纯化的重组蛋白免疫BALB/c小鼠,经细胞融合筛选出14株杂交瘤细胞株。Western blot与间接免疫荧光试验表明14株MAbs均具有良好的反应性。特异性试验表明这些MAbs与牛副流感病毒3型、牛病毒性腹泻病毒及牛传染性鼻气管炎病毒无交叉反应。免疫组化试验表明其中一株1F8亲和力最强,可以用于检测BAV-3。本研究为进一步鉴定BAV-3抗原表位及研发相应的诊断试剂奠定了基础。Bovine adenovirus type 3 （BAV-3） is considered one of the most important respiratory tract agents of cattle diseases. To prepare monoclonal antibodies （MAbs） against Hexon protein of BAV-3, the SP2/0 cells were fused with the splenic cells from BALB/c mice immunized with truncated Hexon protein expressed in E.coli and 14 hybridomas secreted MAbs against BAV-3 were obtained after screening by indirect ELISA based on BAV-3. All the MAbs were positively reacted with BAV-3 detected by western blot and indirect immunofluorescence assay. Further specific assays indicated that 14 MAbs were specific to BAV-3, but not cross-reacted with bovine parainfluenza virus type 3, bovine viral diarrhea virus and bovine herpesvirus type 1 by ELISA assays. In addition, the MAb 1F8 had the higher antibody affinity which was suitable for detecting BAV-3 in animal tissue sample by immunohistochemistry test. The preparation of the MAbs provided the basis for fiLrther epitope identifications of BAV-3 hexon protein and development of diagnostic methods for BAV-3 infection.

关 键 词：牛腺病毒3型 六邻体蛋白 单克隆抗体 特异性 

分 类 号：S852.65[农业科学—基础兽医学]