伪狂犬病病毒UL36蛋白单克隆抗体的制备和鉴定  

Preparation and identification of monoclonal antibody against pseudorabies virus UL36 protein

作　　者：陈玲超 王兴娅 李俊波 李佳佳 王晨 李松楠 王安铖 孟鑫 童武[1] 孔宁[1] 于海[1] 单同领[1] 童光志[1] 郑浩[1] CHEN Lingchao;WANG Xingya;LI Junbo;LI Jiaja;WANG Chen;LI Songnan;WANG Ancheng;MENG Xin;TONG Wu;KONG Ning;YU Hai;SHAN Tongling;TONG Guangzhi;ZHENG Hao(Shanghai Veterinary Research Institute,Chinese Academy of Agricultural Sciences,Shanghai 200241,China;College of Life Sciences,Anhui Agricultural University,Hefei 230036,China;College of Wildlife,Northeast Forestry University,Harbin 150040,China;College of Animal Science and Technology,AnhuiAgricultural University,Hefei 230036,China)

机构地区：[1]中国农业科学院上海兽医研究所,上海200241 [2]安徽农业大学生命科学院,安徽合肥230036 [3]东北林业大学野生动物学院,黑龙江哈尔滨150040 [4]安徽农业大学动物科技学院,安徽合肥230036

出　　处：《中国兽医科学》2024年第3期373-378,共6页Chinese Veterinary Science

基　　金：上海市科技创新行动计划项目(20392002500)。

摘　　要：伪狂犬病病毒UL36基因编码的皮层蛋白VP1/2在病毒粒子组装过程中发挥重要作用。本试验以UL36基因为研究对象,将截短的UL36基因克隆入载体pCold I,构建了重组原核表达质粒pCold I-UL36。经诱导表达,将包涵体形式表达的重组蛋白经过变性、亲和层析及复性后获得了纯度较高的蛋白。将纯化的蛋白免疫BALB/c小鼠,通过细胞融合和间接ELISA方法筛选出1株能稳定分泌针对UL36蛋白的单克隆抗体杂交瘤细胞株,被命名为UL36-6,抗体亚型鉴定结果为Ig G1和κ,制备的腹水抗体中和效价及纯度高,Western-blot和间接免疫荧光试验结果表明单克隆抗体具有很好的特异性。综上所述,本研究成功制备出1株针对PRV UL36蛋白的单克隆抗体,为深入研究UL36蛋白的生物学功能奠定了基础。VP1/2,a tegument protein encoded by the pseudorabies virus UL36 gene,plays an important role in the assembly of virions. In this study,UL36 gene was mainly used as the research object,the truncated UL36 gene was cloned into p Cold Ⅰ vector to construct the recombination prokaryotic expression plasmid p Cold I-UL36.After induced expression,the recombinant protein expressed in inclusion body form was denatured, affinity chromatography and renaturation to obtain high purity protein.Then,BALB/c mice were immunized with purified protein.A hybridoma cell strain named UL36-6,stably secreting antibodies against UL36 was obtained by cell fusion and indirect ELISA screening.The identification of antibody subtype showed that it was Ig G1 and κ.The ascites antibody had high neutralization titer and purity.The Western-blot and indirect immunofluorescence test results showed that the monoclonal antibody had good specificity.In conclusion,we successfully prepared a monoclonal antibody against PRV UL36 protein in the present study which will lay a foundation for further study of the biological function of UL36 protein.

关 键 词：伪狂犬病病毒 UL36基因 VP1/2 单克隆抗体 

分 类 号：S852.659.1[农业科学—基础兽医学]