A型塞内卡病毒VP1蛋白单克隆抗体的制备及鉴定  

Preparation and Identification of a Monoclonal Antibody against Senecavirus A VP1 Protein

作　　者：郭金硕 侯磊[2] 全荣[2] 王菁[2] 韦莉[2] 刘爵[2] GUO Jin-shuo;HOU Lei;QUAN Rong;WANG Jing;WEI Li;LIU Jue(College of Animal Science and Technology,Beijing University of Agriculture,Beijing 102206,China;Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry,Institute of Animal Husbandry and Veterinary Medicine,Beijing Academy of Agriculture and Forestry Sciences,Beijing 100097,China)

机构地区：[1]北京农学院动物科学技术学院,北京昌平102206 [2]北京市农林科学院畜牧兽医研究所畜禽传染病防控技术北京市重点实验室,北京海淀100097

出　　处：《中国兽医杂志》2022年第3期20-24,共5页Chinese Journal of Veterinary Medicine

基　　金：国家重点研发计划(2017YFD0500103)。

摘　　要：为制备抗A型塞内卡病毒(SVA)VP1蛋白的单克隆抗体,并初步应用于A型塞内卡病毒的检测,本研究构建了SVA VP1原核重组表达质粒pET30a-GST-VP1(528 bp)。将重组质粒转化大肠埃希菌BL21感受态细胞并大量表达VP1重组蛋白,利用纯化的VP1重组蛋白免疫BALB/c小鼠,取其脾细胞与骨髓瘤细胞(SP2/0细胞)进行融合,经过细胞亚克隆筛选,获得杂交瘤细胞。将其选为腹水生产细胞株,最后纯化单克隆抗体,通过Western blotting和间接免疫荧光试验对其进行鉴定。结果显示,本研究获得1株能够稳定分泌抗VP1重组蛋白单克隆抗体的杂交瘤细胞,其亚型重链为IgG2a、轻链为Kappa链;Western blotting和间接免疫荧光检测显示,该单克隆抗体能够特异性识别SVA病毒粒子。本研究成功制备的抗SVA VP1蛋白单克隆抗体可为进一步研究A型塞内卡病毒VP1蛋白的生物学功能及A型塞内卡病毒致病机制提供有效途径。This study aimed to prepare and identify monoclonal antibody against Senecavirus A(SVA) VP1 protein for diagnostic application.A prokaryotic recombinant expression plasmid pET30 a-GST-VP1(528 bp) was constructed and transformed into E.coli BL21 competent cells for VP1 expression.BALB/c mice were immunized with purified recombinant VP1 protein and hybridoma cells were obtained by fusion of spleen cells and myeloma cells(SP2/0 cells) and clonal screening.Monoclonal antibodies from selected clones were purified and identified by Western blotting and indirect immunofluorescence assay(IFA).A hybridoma cell line that stably secreted monoclonal antibody against VP1 protein was generated.This antibody was of IgG2 a subclass with Kappa light chain,and specifically recognized the SVA in Western blotting and IFA.This anti-SVA VP1 monoclonal antibody thus provides the opportunity to further investigate the biological function of Senecavirus A VP1 protein and the pathogenesis of Senecavirus A.

关 键 词：A型塞内卡病毒 VP1蛋白 杂交瘤细胞 单克隆抗体 

分 类 号：S855.3[农业科学—临床兽医学]