牛副流感病毒3型NP单抗的制备及初步应用  

Preparation and preliminary application of monoclonal antibody against nucleocapsid protein of bovine parainfluenza virus type 3

作　　者：吕闯[1] 朱远茂[1] 董秀梅[1] 蔡红[1] 于作[1] 高欲燃[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》2011年第12期970-973,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：哈尔滨市科技攻关项目(2010AA6AN019);中央级公益性科研院所基本科研业务费项目(ZGKJ201009)

摘　　要：为制备牛副流感病毒3型(BPIV3)核衣壳蛋白(NP)单克隆抗体(MAb),本研究利用原核表达并纯化的重组NP(rNP)免疫BALB/c小鼠,取免疫后小鼠脾细胞与骨髓瘤细胞SP2/0融合。采用以BPIV3为检测抗原的间接ELISA方法筛选阳性细胞克隆,经3次克隆纯化后获得1株稳定分泌抗NP特异性MAb的杂交瘤细胞株(5E5)并制备腹水,采用rNP及BPIV3包被的ELISA效价分别是2×106和1.28×105。间接ELISA、western blot、IFA试验表明该MAb具有良好的反应性和特异性。经抗体亚类鉴定该MAb亚类为IgG1/κ。特异性试验表明该MAb不与牛传染性鼻气管炎病毒、牛病毒性腹泻病毒反应。免疫组化试验表明该MAb可以检测BPIV3感染动物体内的病原。该MAb还可用于建立检测BPIV3病原及抗体的诊断方法,同时为研究NP的结构和功能提供了条件。To prepare monoclonal antibody （MAb） against nucleocapsid protein （NP） of bovine parainfluenza virus type 3 （BPIV3）, BALB/c mice were immunized with purified recombinant NP （rNP） expressed by E. coli and a hybridoma secreting MAb was screened from fusing the SP2/0 cells with the spleen cells of the immunized BALB/c mice by indirect ELISA coated with BPIV3. The titers of MAb in ascites were 2 × 10^6 and 1.28 × 10^5 as detected by rNP and BPIV3 coated ELISA, respectively. The MAb was specifically reacted with BP1V3 identified by indirect ELISA, western blot, immunofiuroescence assay. The specific tests indicated the MAb had no reaction with infectious bovine rhinotracheitis virus and bovine viral diarrhea virus. The BPIV3 was detected in experimentally infected animals in immunohistochemical test with the MAb. Therefore, this MAb could be used to establish diagnosis method for BPIV3 and further study on the structure and function of NP.

关 键 词：牛副流感病毒3型 核衣壳蛋白 单克隆抗体 特异性 

分 类 号：S852.65[农业科学—基础兽医学]