作　　者： (韦传宝）; (邓辉）;

机构地区： 皖西学院生物与制药工程学院,六安237012

出　　处： 《中国免疫学杂志》 2017年第9期1355-1358,1360,共5页

摘　　要： 目的:建立检测侵染生姜的烟草花叶病毒的ELISA方法。方法:用RT-PCR方法检测侵染生姜的烟草花叶病毒,挑选仅含烟草花叶病毒的生姜叶片,采用离心方法提取叶片中的烟草花叶病毒,通过12%SDS-PAGE和5%~20%梯度SDS-PAGE二次制备电泳分离出烟草花叶病毒的外壳蛋白CP,将含CP的聚丙烯酰胺凝胶磨细后免疫小鼠,获得抗血清。用Western blot检测抗血清特异性,ELISA检测抗血清的结合性,通过亲和层析纯化抗血清中的Ig G。以待测生姜叶片液为抗原包被,纯化的抗血清IgG为一抗,羊抗小鼠IgG为二抗,建立检测侵染生姜的烟草花叶病毒的间接ELISA。结果:抗血清中的Ig G仅仅与烟草花叶病毒的CP结合,并能够结合天然烟草花叶病毒颗粒。建立的ELISA对花叶病生姜叶片的检测结果与RT-PCR检测结果一致。结论:成功地建立了检测侵染生姜的烟草花叶病毒的ELISA方法。 Objective In order to establish ELISA method for testing tobacco mosaic virus （TMV） infecting Zingiber officinale Rose. Methods： Tobacco mosaic virus（TMV） infecting Zingiber officinale Rosc was tested by RT-PCR. Zingiber officinale Rosc leaves which only contained TMV were choiced. TMV particle was purified by centrifugation method. TMV CP was purified through preparation electrophoresis including 12% SDS-PAGE first and then 5% -20% gradient SDS-PAGE. Polyacrylamide gel contained TMV CP w,as ground into suspension. Mice were immuned with the suspension and antiserum was obtained. Antiserum quality was tested by Western blot and ELISA test. IgG was purified through affinity chromatography method. IgG solution was concentrated and dialyzed to a suitable concentration. The IgG then mixed with glycerol. Results ： IgG in antiserum only combined with TMV CP protein and it could combine with nature TMV particle CP protein. Its quality was up to standard. Conclusion： Establishment of ELISA method for testing TMV infecting Zingiber offcinale Rosc is successfull by using this IgG.

关 键 词： 烟草花叶病毒 生姜 方法