多囊蛋白1胞内区单克隆抗体的制备及初步应用

Preparation of monoclonal antibodies against polycystin-1 intracellular region

目的 :制备多囊蛋白 1胞内区单克隆抗体 ,并用以观察多囊蛋白 1在肾组织中的分布与表达。 方法 :以重组多囊蛋白 1胞内段融合蛋白为抗原 ,采用杂交瘤技术 ,制备产生针对多囊蛋白 1胞内区的单克隆抗体的杂交瘤细胞株 ,并利用所产生的单克隆抗体 ,采用标准 En Vision免疫组织化学方法 ,对多囊肾组织、正常成人肾组织和胎肾组织进行染色。结果 :获得 4株稳定分泌抗体且效价为 1∶ 10 6 左右的杂交瘤细胞株 ,传代至第 5 0代仍能稳定分泌抗体 ,效价不变 ;多囊蛋白 1在正常成人肾组织的近端小管、远端小管和集合管的上皮细胞呈弱阳性表达 ,在胎儿肾脏的肾小管上皮细胞呈强阳性 ,在多囊肾组织的囊肿衬里上皮细胞表达明显增强。 结论 :成功制备了多囊蛋白 1胞内区单克隆抗体 ,为深入了解多囊蛋白

Objective: To prepare the monoclonal antibodies against polycystin 1 intracellular region and to study the distribution and expression of polycystin 1 in kidney tissues. Methods: Using the recombinant fusion protein containing polycystin 1 intracellular region as antigen, the hybrid cells secreting the monoclonal antibodies against polycystin 1 intracellular region were established by hybridoma technique. The distribution and expression of polycystin 1 in polycystic kidney, fetal kidney and adult kidney were investigated by immunohistochemical methods(standard EnVision method) with the monoclonal antibodies. Results: Four cell lines of hybrids steadily secreting the monoclonal antibodies against polycystin 1 intracellular region were established. The antibody titers were 1∶10 6. The 50th generation of these cell lines of hybrids still can secret the monoclonal antibodies and the titers remain similar. Polycystin 1 was weakly expressed in tubules and collecting ducts of normal kidney, the strong staining was seen at tubules of fetal kidney, and very strong staining of cyst lining epithelium of polycystic kidney was observed. Conclusion: The monoclonal antibodies against polycystin 1 intracellular region will be a useful tool in the studies of polycystin 1 structure and function.