猪圆环病毒2型Rep'蛋白单克隆抗体制备及其抗原表位鉴定  

Preparation of the monoclonal antibody against the Rep' protein of porcine circovirus type 2 and its antigenic epitope identification

作　　者：刘丹[1,2] 刘建波[1] 吴洪丽[1] 王一平[1] 危艳武[1] 唐青海[1] 李胜斌[1] 魏萍 刘长明[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/猪传染病研究室,黑龙江哈尔滨150001 [2]东北农业大学动物医学学院,黑龙江哈尔滨1500301

出　　处：《中国预防兽医学报》2013年第5期405-409,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金(31101837);公益性行业(农业)专项(201203039);"863"家畜病毒病基因工程疫苗创制(2011AA10A208)

摘　　要：为制备猪圆环病毒2型Rep'蛋白单克隆抗体(MAb),本研究采用淋巴细胞瘤杂交技术制备其MAb,获得了1株能够稳定分泌抗PCV2-Rep'蛋白的杂交瘤细胞株,命名为3D1株。MAb亚类鉴定为IgG1/κ型,腹水效价达1∶819 200。Western blot分析表明,该MAb可与重组杆状病毒表达的PCV2-rRep'和PCV2-rRep蛋白发生特异性反应,具有良好的特异性和反应原性。采用MAb对PCV2感染细胞中Rep'蛋白抗原性进行了鉴定,证明该MAb能够与病毒Rep'蛋白产生特异性反应。采用合成肽扫描法对MAb对应的抗原表位鉴定,其核心序列为61FANFVKKQTFNKV73,位于PCV2-Rep'蛋白的N末端。制备的MAb及其抗原表位鉴定,为该病毒分子生物学及诊断技术的研究奠定了基础。Porcine circovirus type 2 （PCV2） is an important causative agent of postweaning multisystemic syndrome in pigs, and both replicase proteins （Rep＇ and Rep） encoded by PCV2-ORF1 are essential for the viral replication. The monoclonal antibody （MAb） to the Rep＇ protein was prepared by means of lymphocyte hybridoma technique. The MAb isotype was belong to be IgG1/K. The antibody titer of ascites was 1：819,200. Western blot and IPMA analysis showed that the MAb was able to react with both PCV2-rRep＇ and PCV2-rRep proteins expressed by baculovirus system and PCV2 infected cells. Furthermore, the Rep＇ protein antigenic epitope was mapped by the synthesized peptides to scan with the MAb. The epitope core sequence was identified as 6ZFANFVKKQTFNKV73, and it located at the nuclear localization signal region of the N terminal of Rep＇ protein. This MAb and its corresponding epitope mapping provide a basis for further study on the viral molecular biology and diagnosis technique.

关 键 词：猪圆环病毒2型 重组Rep’蛋白 单克隆抗体 抗原表位 

分 类 号：S852.65[农业科学—基础兽医学]