烟草花叶病毒单克隆抗体的制备和鉴定  

Preparation and identification of monoclonal antibody against Tobacco mosaic virus

作　　者：吕琦[1] 吴峰[1,2] 林洁[1] 谭仲夏[3] 秦西云[3] 夏振远[3] 莫笑晗[3] 马岚[2] 

机构地区：[1]云南大学生命科学学院,云南昆明650091 [2]清华大学深圳研究生院生命与健康学部,广东深圳518005 [3]云南省烟草农业科学研究院,云南玉溪653100

出　　处：《云南大学学报（自然科学版）》2013年第1期93-98,共6页Journal of Yunnan University(Natural Sciences Edition)

基　　金：中国烟草总公司云南省分公司科技计划项目(07A07)

摘　　要：经工程菌表达与纯化,得到了纯度95%以上的TMV-CP-F重组蛋白,配合提取的TMV天然病毒颗粒作为免疫原.通过杂交瘤技术获得了14株能分泌特异针对TMV外壳蛋白的单克隆抗体杂交瘤细胞株.经鉴定14株细胞所分泌的抗体亚类为IgG1型,抗体轻链均为κ型.经ProteinA一步法亲和层析纯化所得抗体经鉴定相对分子质量在149.36～157.23 ku之间,抗体纯度在80%以上.经间接ELISA测定,14株抗体均与TMV-CP重组蛋白和TMV病毒有良好特异性反应.所制备的抗TMV抗体的特异性高,可用于与其相关的免疫检测研究和应用.Recombinant TMV coat protein（TMV-CP） was expressed by engineered bacteria and then purified（purity was above 95 %）.It was used as immunogen together with the natural TMV particles for immunization.After hybridoma screening,14 hybridoma cell lines which stably secreted monoclonal antibodies against recombinant TMV-CP were obtained.The IgG subclasses and light-chain isotypes of all these antibodies were identified as IgG1 and κ respectively.The antibodies were purified by one-step recombinant protein-A affinity chromatography.Their molecular weight was among 149.36-157.23 ku,and the purity was above 80 %.The ELISA results showed that all of these 14 monoclonal antibodies reacted with recombinant TMV-CP and natural TMV particles specifically.The TMV monoclonal antibodies have high specificity and it can be used for relevant researches and immunoassays.

关 键 词：烟草花叶病毒 单克隆抗体 间接法ELISA 

分 类 号：S432.41[农业科学—植物病理学]