巴泰病毒单克隆抗体的制备及鉴定  

Preparation and Identification of Monoclonal Antibodies Against the Batai Virus

作　　者：朱翔宇 史宁[1] 李丽霞 蔡熙姮 刘昊[1,2] ZHU Xiangyu;SHI Ning;LI Lixia;CAI Xiheng;LIU Hao(Institue of Special Economic Animal and Plant Science,Chinese Academy of Agricultural Science,Changchun 130112,China;Foshan University,Foshan 528000,China;Jilin Wildlife Rescue and Rehabilitation Center,Forestry Department of Jilin Province,Changchun 130112,China)

机构地区：[1]中国农业科学院特产研究所,长春130112 [2]佛山科学技术学院,佛山528000 [3]吉林省林业厅野生动物救护繁育中心,长春130112

出　　处：《病毒学报》2020年第1期55-62,共8页Chinese Journal of Virology

基　　金：国家自然科学基金(项目号:31802199),题目:巴泰病毒通过网格蛋白介导的内吞途径入侵N2a细胞机制研究。

摘　　要：巴泰病毒(Batai virus,BATV)是一种人兽共患病毒,近年在全球广泛流行。我国也有人和动物感染的相关报道,而目前对BATV的研究仅局限于分子检测和全基因测序,还没有关于BATV单克隆抗体的相关研究,但制备特异性强、活性好的单克隆抗体也是防控BATV的关键。为了制备BATV单克隆抗体,本研究利用纯化的BATV作为抗原免疫雌性BALB/c小鼠,制备鼠源单克隆抗体。通过免疫小鼠后断尾取血,间接ELISA测其血清效价后,将SP2/0细胞与小鼠的脾细胞融合。ELISA方法筛选阳性细胞株,并进行亚克隆后制备单抗腹水,并利用辛酸-硫酸铵沉淀法纯化腹水。结果获得7株单克隆抗体,包括5株IgG型和2株IgM型。其中制备的3E2单克隆抗体效价最高为1∶32 000。经间接ELISA、间接免疫荧光和Western Blot检测表明,制备的3E2单抗具有较好的纯度、活性和特异性,为BATV快速检测方法的建立及致病机制研究奠定了基础。The Batai virus(BATV)has been reported to infect humans and animals in China. Research on the BATV is limited to molecular detection and whole-genome sequencing. Research on monoclonal antibodies(mAbs) against BATV is lacking. However,preparation of specific and active mAbs are important for the prevention and control of BATV infection. We prepared and identified mAbs against the BATV. First,we prepared a murine mAb by immunizing female BALB/c mice with the purified BATV NM/12 strain. Then,mice blood was collected from tail veins after three immunizations. Serum titers were measured by indirect enzyme-linked immunosorbent assay(ELISA). The spleen cells of mice were fused with SP2/0 cells. Positive cell lines were screened by indirect ELISA,and implanted into the abdominal cavities of mice to elicit ascites.Ascites fluid was purified by octanoic acid-ammonium sulfate precipitation. Finally,we obtained seven mAbs(five IgG and two IgM). The highest titer of 3 E2 mAbs was 1∶32,000. We achieved good purity,activity and specificity of mAbs by indirect ELISAs,immunofluorescence analyses and Western Blot. The present study lays a foundation for establishment of a rapid-detection method of the BATV,as well as research into its prevention and control.

关 键 词：巴泰病毒(BATV) 单克隆抗体(mAb) 杂交瘤细胞(Hybridoma cell) 

分 类 号：R373.3[医药卫生—病原生物学]