首页|南非2型口蹄疫病毒VP1蛋白单抗的制备与特性研究

南非2型口蹄疫病毒VP1蛋白单抗的制备与特性研究

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为制备南非2型口蹄疫病毒型特异性单克隆抗体,本研究通过原核系统表达南非2型口蹄疫病毒VP1蛋白,亲和层析的方法纯化目的蛋白.将纯化的VP1蛋白使用弗氏佐剂乳化后,免疫BALB/c小鼠,待加强免疫后取血清效价最稳定的小鼠脾细胞与小鼠骨髓瘤SP2/0细胞进行融合,应用间接ELISA筛选针对南非2型VP1蛋白的单克隆抗体.采用特异性试验、叠加试验和抗体亚型鉴定等方法对筛选的单克隆抗体特性进行分析.结果发现,本研究成功表达南非2型VP1蛋白,筛选出2株能稳定分泌特异性针对南非2型VP1蛋白的杂交瘤细胞株(3B2和6F2),其亚型分别为IgG2a和IgG1,叠加试验结果表明2株单抗的叠加系数＞40％,说明两株单抗针对不同的抗原位点;单抗的效价可达1∶256 000,特异性试验证明2株单抗均不与O型、A型口蹄疫病毒及其他常见猪病病毒产生交叉反应.由此可见,本研究制备的单克隆抗体具有良好的特异性和反应性,为南非2型口蹄疫病毒定型诊断和未来疫苗的研制奠定了基础.
Preparation and Characterization of A VP1-Specific Monoclonal Antibody of South African Type 2 Foot-and-Mouth Disease Virus
In order to prepare South African type 2 foot-and-mouth disease virus(SAT2-FMDV)specific monoclonal antibodies,in this study,the VP1 protein of SAT2-FMDV was expressed by a prokaryotic system.The target protein was purified by affinity chromatography.The purified VP1 protein was emulsified with Freund's adjuvant and immunized to Balb/c mice.Mouse splenocytes with the most stable serum potency in serum were fused with mouse myeloma SP2/0 cells after reinforcement of the immunization.Indirect ELISA was applied to screen for monoclonal antibodies against the VP1 protein.of SAT2-FMDV.The characteristics of the screened monoclonal antibodies were analyzed by specificity tests,overlay tests,and identification of antibody subtype.The VP1 protein of SAT2-FMDVwas expressed in this study.Two hybridoma cell lines(3B2 and 6F2)were screened that could stably secrete antibodies specifically targeting the VP1 protein of SAT2-FMDV,and their subtypes were IgG2a and IgGl,respectively.A superposition assay showed that the stacking coefficients of the two monoclonal antibodies was＞40％,which indicated that the two monoclonal antibodies were directed against different antigenic sites.The potency of the monoclonal antibodies was up to 1∶256 000.A specificity assay used to characterize the monoclonal antibodies revealed no cross-reaction with O-type or A-type foot-and-mouth disease viruses or other common viruses that cause diseases in pigs.The monoclonal antibodies prepared in this study had good specificity and reactivity.Our data lay a foundation for the SAT2-FMDV diagnosis and vaccine development.

South African type 2 foot-and-mouth disease virusVP1 proteinSpecific monoclonal antibody

朱昱茜、石正旺、罗俊聪、陈婕、林永玉、席韬、张帆、石鑫泰、郑海学、包世俊、田宏

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甘肃农业大学动物医学院,兰州 730070

中国农业科学院兰州兽医研究所兰州大学动物医学与生物安全学院家畜疫病病原生物学国家重点实验室,兰州 730000

南非2型 口蹄疫病毒 VP1蛋白 特异性单克隆抗体

科技部国家重点研发计划项目科技重大专项-国家重点实验室重组项目2022年甘肃省创新联合体项目十四五广东省揭榜挂帅项目国家生猪技术创新中心先导科技项目

2021YFD180030022ZD6NA00122ZD6NA0122023SDZG02NCTIP-XD/C03

2024

10.13242/j.cnki.bingduxuebao.004493

病毒学报
中国微生物学会

病毒学报

CSTPCD北大核心
影响因子：1.046
ISSN：1000-8721
年,卷(期)：2024.40(4)
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