【摘要】 为制备牛支原体表面脂蛋白P48的单克隆抗体,应用纯化的牛支原体武威株脂蛋白P48基因的原核表达产物免疫BALB/c小鼠,继而取免疫小鼠的脾细胞与小鼠骨髓瘤细胞SP2/0融合,经克隆和筛选,获得2株能分泌抗牛支原体脂蛋白P48抗体的杂交瘤细胞融合株,并分别命名为C7及E5。ELISA和W estern-blot检测结果表明,2株单克隆抗体均可与牛支原体特异性地结合,而与牛的其他常见病原菌无任何反应。亚类鉴定结果表明,2株单克隆抗体重链均为Ig G 1,轻链均为λ型。稳定性试验结果表明,2株杂交瘤细胞均可稳定分泌单克隆抗体。本研究结果为建立牛支原体有效的检测方法和深入研究脂蛋白P48的生物学功能奠定了基础。更多还原

【Abstract】 To prepare monoclonal antibodies(Mc Abs) against lipoprotein P48 from Mycoplasma bovis(M.bovis), The lipoprotein P48 gene from M.bovis Wuwei strain was expressed in a prokaryotic expression system, purified, and used as antigen to immunize BALB/c mice. The spleen cells from the immunized mice were fused with mouse myeloma cell line SP2/0. Two positive hybridoma cell lines secreting Mc Abs against P48, named as C7 and E5, were selected by indirect enzyme-linked immunosorbent assay(ELISA)based on limited dilution method. Western-blot and ELISA showed that the Mc Abs can react with the recombinant lipoprotein protein P48 and M.bovis total protein specifically, but they did not react with other bovine pathogenic bacteria. The immunoglobulin subtypes of Mc Abs were identified and the results showed that both Mc Abs were Ig G1, and light chains were λ. Two hybridoma cell lines could secret Mc Abs against lipoprotein P48 stably in vitro. The study results laid a foundation for establishing detection method for M.bovis and further studying biological functions of the lipoprotein P48 from M.bovis.更多还原