【摘要】 目的为了获得高效价和高纯度的O型口蹄疫病毒单克隆抗体。方法用O型口蹄疫乳鼠毒(LD50=10-8.5)感染免疫小鼠,按有限稀释法和ELISA方法筛选出阳性杂交瘤细胞株,制备单抗腹水,所收腹水用50%和45%的饱和硫酸铵沉淀,再用proteinG亲和层析柱进行亲和层析纯化单体,并用SDS-PAGE和单抗检测方法分析其纯化抗体的纯度和活性。结果经3次亚克隆获得了一株杂交瘤细胞株(10E6),抗体亚类鉴定为IgG2a;SDS-PAGE电泳显示,纯化的抗体纯度较高,只有IgG2a的重链和轻链,而没有其它杂蛋白带;单抗ELISA检测方法显示,纯化的抗体具有良好的活性,其效价为1∶102400,高于腹水效价。结论用proteinG亲和层析法获得了高纯度和高效价的单克隆抗体,将为O型口蹄疫病毒的基础研究和应用研究奠定基础。更多还原

【Abstract】 The aim of this study is to prepare a monoclonal antibody against foot and mouth disease virus(FMDV).BALB/c mice was immunized with FMDV O type with LD50 of 10-8.5,and the positive clones were screened by indirect ELISA and limiting dilution assay.The monoclonal antibody produced in the ascitic fluids of mice was purified with 45% and 50% saturated ammonia sulfate and affinity column with protein G and the purity and titer of the monoclonal antibody were assayed with indirect ELISA and SDS-PAGE.The result showed that one strain of hybridoma cell designated as 10E6 was obtained and its monoclonal antibody secreted was proved to be IgG2a subclass.As demonstrated by SDS-PAGE,the purity of this antibody was rather high,and only heavy and light chain of IgG2a were present without any other protein band in the SDS-PAGE analysis.The titer of this antibody present in the ascitic fluids of mice revealed by indirect ELISA was 1:102,400.It is evident that the monoclonal antibody against FMDV with high purity and high titer can be obtained by purification through affinity column with protein G.And the monoclonal antibody would provide a basis for the basic studies and clinical use in foot-and-mouth virus infection.更多还原