猪繁殖与呼吸综合征病毒谱系8毒株特异性单克隆抗体研制  

Development of monoclonal antibodies specific to the lineage 8 strains of porcine reproductive and respiratory syndrome virus

作　　者：任语馨 张杰[1] 高雁怩 孙杨杨[1] 张路捷 白娟[1] 姜平[1] REN Yuxin;ZHANG Jie;GAO Yanni;SUN Yangyang;ZHANG Lujie;BAI Juan;JIANG Ping(Nanjing Agricultural University/Key Laboratory of Animal Bacteriology,Ministry of Agriculture and Rural Affairs,Nanjing 210095,China)

机构地区：[1]南京农业大学/农业农村部动物细菌学重点实验室,江苏南京210095

出　　处：《畜牧与兽医》2025年第1期111-118,共8页Animal Husbandry & Veterinary Medicine

基　　金：江苏省自主创新研发基金项目(CX(22)1011);国家生猪产业技术体系专项(CARS-35);国家自然科学基金项目(32230103)。

摘　　要：猪繁殖与呼吸综合征病毒(PRRSV)流行毒株存在多种基因亚型(谱系)和抗原性差异,为了鉴别不同谱系流行毒株,选取PRRSV谱系8毒株BB0907(高致病性毒株)非结构蛋白2(NSP2)高变区基因片段(tNSP2),采用大肠杆菌表达制备获得NSP2重组蛋白,并免疫BALB/c小鼠。结果:研制出5株NSP2单克隆抗体(3D6、6B2、6C3、8C2和8H6),小鼠腹水ELISA抗体效价均达1∶216000;5株单抗轻链类型都为Kappa型,3D6、6B2和6C8的重链类型为IgG1,8C2的重链类型为IgG2b,8H6重链类型为IgG2a;通过构建tNSP2蛋白截短体,Western blot鉴定出4个抗原表位,即^(485)GPLNFPTPSE^(494)、^(555)FPLAPSQNMG564、575EVLSEISDIL^(584)和^(585)NDTPAPVS^(593);间接免疫荧光试验结果显示,这些单抗均能与PRRSV谱系8毒株发生特异性反应,但与谱系1和谱系5毒株不反应。本研究为PRRSV分离毒株抗原鉴别诊断和生物学研究提供了有意义的研究材料。Porcine reproductive and respiratory syndrome virus(PRRSV),which is a pathogen that seriously endangers Chinas breeding industry,has multiple genotypes(lineages)and antigenic differences in circulating strains.In order to identify the circulating strains of dif-ferent lineages,the NSP2 recombinant protein was prepared by using the NSP2 hypervariable region gene fragment(tNSP2)of the PRRSV lineage 8 epidemic strain BB0907(a highly pathogenic strain),and the NSP2 recombinant protein obtained was used to immunize BALB/c mice.As a result,five monoclonal cell lines that could stably secrete antibodies were developed by cell fusion.The titers of the antibodies in ascites fluid in mice were≥1∶216000.The heavy chains of 3D6,6B2 and 6C8 were IgG1,the heavy chain of 8C2 was IgG2,while the heavy chain of 8H6 was IgG2a,and the light chains of the five McAbs were of kappa subtype.Four epitopes were identified by the construc-tion of NSP2 protein truncates,which were^(485)GPLNFPTPSE^(494),^(555)FPLAPSQNMG564,575EVLSEISDIL^(584) and 585NDTPAPVS^(593).These mono-clonal antibodies were observed to be able to react specifically with 8 strains of the PRRSV lineage by indirect immunofluorescence assay,but they did not react with the lineage 1 and lineage 5 strains.This study providedsignificantresearch materials for the differential diagnosis of and biological research onisolated PRRSV antigens.

关 键 词：猪繁殖与呼吸综合征病毒 NSP2 单克隆抗体 B细胞表位 

分 类 号：S858[农业科学—临床兽医学]