【摘要】 目的:制备抗血清I型马立克氏病病毒(MDV-1)VP22的单克隆抗体(mAb),并鉴定其免疫学特性。方法:在原核系统中表达VP22羧基端区域(94-243aa),获得融合表达产物GST-VP22C。将该表达产物切胶免疫小鼠,利用杂交瘤技术制备抗MDV-1 VP22C的mAb,并通过ELISA、间接免疫荧光(IFA)、Western blot鉴定其特性。结果:获得了2株可稳定分泌抗MDV-1 VP22C的mAb的杂交瘤细胞,命名为3F7、4E4。IFA鉴定表明,两株mAb均能与感染MDV-1的成纤维细胞反应,其中,3F7 mAb染色呈现MDV空斑,而4E4mAb呈现整个单层的细胞核荧光。ELISA和Western blot分析表明,3F7能与杆状病毒表达的VP22反应,4E4不具备该特性。对3F7 mAb进一步鉴定,确定了该mAb针对的具体位置在94-193aa处,是蛋白转导域的预测位置。结论:成功地制备了抗MDV-1 VP22C的mAb,为深入研究VP22蛋白的转导功能提供了有用的试剂。更多还原

【Abstract】 AIM:To prepare and characterize monoclonal antibodies(mAb) against VP22 carboxyl terminus of CVI988/Rispens strain of Marek’s disease virus serotype 1.METHODS: Carboxyl terminus of CVI988 VP22(94aa～243aa) was expressed in prokaryotic system.mAbs against VP22 were prepared by hybridoma technology from BALB/c mice immunized with the fusion protein GST-VP22C and characterized by ELISA,indirect fluorescence assay(IFA) and Western blot.RESULTS: Two hybridoma cell lines stably secreting mAb against VP22C were obtained and designated as 3F7 and 4E4.mAb 3F7 could react with VP22 expressed in all the plaques,while mAb 4E4 stained all the cells nuclei in MDV-infected CEF cells.It was also found that 3F7 could react with VP22 expressed in Sf9 cells and denatured VP22 by Western blot analysis.In addition,it was further showed that the epitope of mAb 3F7 was located within the domain between 94aa and 193aa,the predicted site of protein transduction domain of VP22.CONCLUSION: The preparation of the mAb is very important to further research in protein transduction domain of MDV-1 VP22.更多还原