口蹄疫病毒非结构蛋白3AB单克隆抗体的制备及其对应表位区分析

Preparation of monoclonal antibodies against nonstructural protein 3AB of foot-and-mouth disease virus and analysis of their antigenic epitope domains

为鉴定口蹄疫病毒(FMDV)的非结构蛋白3AB的抗原表位,本研究以原核表达并纯化的FMDV 3AB重组蛋白免疫BALB/c小鼠,采用淋巴细胞杂交瘤技术制备杂交瘤细胞,通过间接ELISA进行筛选,获得6株能够稳定分泌抗3AB蛋白特异性单克隆抗体(MAb)的杂交瘤细胞。这6株MAbs亚类鉴定均为IgG1型,轻链均为κ链。间接免疫荧光试验结果表明,这6株MAbs均能够识别FMDV 3AB蛋白。采用制备的MAb对分段表达的3AB蛋白进行western blot分析,结合位点分别位于3AB的第aa 55～aa 70、aa 64～aa 79、aa 130～aa 145区段。该结果为进一步探索3AB蛋白的结构和功能以及建立诊断方法奠定了基础。

To identify linear epitopes on nonstructural protein 3AB of foot-and-mouth disease virus（FMDV）,BALB/c mice were immunized with purified recombinant 3AB expressed in E.coli and the splenocytes of the BALB/c mice were fused with myeloma cells SP2/0.Six hybridoma cell lines stably secreting monoclonal antibodies（MAbs） were screened by indirect ELISA against the protein 3AB.The subtypes of the 6 MAbs were IgG1 and κ light chain.All the MAbs were able to react with FMDV 3AB detected by indirect immunofluorescence assay.Western blot analysis indicated that the MAbs were reacted with overlapping short peptides of FMDV 3AB C-terminus aa 55 to aa 70,aa 64 to aa 79 and aa 130 to aa 145 expressed in E.coli,respectively.It was concluded that the MAbs against the protein 3AB could be useful tools for futher studying on the structure and function of FMDV 3AB,as well as for development of diagnostic methods.