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分泌抗芜菁花叶病毒特异性单克隆抗体杂交瘤细胞系的建立及单克隆抗体特性分析

Establishment of hybridoma cell line secreting specific monoclonal antibodies against turnip mosaic virus and analysis of properties of the McAb.

作者信息

Yang F, Cai S, Xu L, Feng L

机构信息

Biotechnology Research Center, CAAS (Chinese Academy of Agricultural Sciences), Beijing.

出版信息

Chin J Biotechnol. 1992;8(4):247-54.

PMID:1343828
Abstract

Monoclonal antibodies (McAb) of hybridomas derived from the fusing of the mouse splenocytes immunized by TuMV with BALB/c mouse myeloma cells (SP2/0-Ag14). Five kinds of hybridoma cell line were produced by indirect-ELISA screening and cloning three times with limiting dilution. Four kinds of hybridoma produced antibodies respectively reactive to TuMV C1, C3, C4 and C5. One kind was reactive to all five strains of TuMV. In indirect-ELISA and sandwich-ELISA tests, TuMV specific monoclonal antibodies did not react with CaMV, CMV, TMV, PVX, and PVY. Antibody titers of ascitic fluids were about 1:256,000 to 2,048,000 in indirect-ELISA. The biological, physical, and chemical properties of the hybridoma cell lines and McAb were identified. The identification of TuMV strains, the specificity and stability of McAb, the coat proteins, and the antigenic site of TuMV were discussed and analyzed with SDS-PAGE and western-blotting.

摘要

用芜菁花叶病毒(TuMV)免疫的小鼠脾细胞与BALB/c小鼠骨髓瘤细胞(SP2/0-Ag14)融合得到的杂交瘤产生的单克隆抗体(McAb)。通过间接酶联免疫吸附测定(ELISA)筛选并经三次有限稀释克隆,获得了五种杂交瘤细胞系。四种杂交瘤分别产生了对TuMV C1、C3、C4和C5有反应的抗体。一种对所有五种TuMV毒株都有反应。在间接ELISA和夹心ELISA试验中,TuMV特异性单克隆抗体与花椰菜花叶病毒(CaMV)、黄瓜花叶病毒(CMV)、烟草花叶病毒(TMV)、马铃薯X病毒(PVX)和马铃薯Y病毒(PVY)均无反应。间接ELISA中腹水的抗体效价约为1:256,000至2,048,000。对杂交瘤细胞系和单克隆抗体的生物学、物理和化学性质进行了鉴定。用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)和蛋白质免疫印迹法(western-blotting)对TuMV毒株的鉴定、单克隆抗体的特异性和稳定性、外壳蛋白以及TuMV的抗原位点进行了讨论和分析。

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