PAGE-4单克隆抗体的制备及表达分布

目的:制备抗PAGE-4 mAb并鉴定其特异性及敏感性,应用该抗体在蛋白质水平上分析PAGE-4在正常与疾病组织中的表达. 方法:以PAGE-4为抗原免疫BALB/c小鼠,利用杂交瘤技术建立稳定分泌抗PAGE-4 mAb的杂交瘤细胞系. 采用Western Blot及ELISA等方法鉴定抗体的特异性和敏感性. 应用正辛酸-饱和硫酸铵方法纯化抗体. 利用免疫组织化学方法对PAGE-4在正常与疾病组织中的表达进行分析. 结果:获得4株分泌特异性抗PAGE-4 mAb的杂交瘤细胞株,mAb具有较高的特异性与敏感性. 在正常男女生殖系统组织如:睾丸、前列腺、子宫中以及正常胰腺组织中有PAGE-4的表达,在前列腺癌、子宫内膜腺癌中PAGE-4高表达. PAGE-4在蛋白质水平上的表达格局与以往有关PAGE-4在mRNA水平上的报道一致. 结论:成功获得抗PAGE-4 mAb,为研究PAGE-4定位及功能提供了重要工具.

Preparation and characterization of monoclonal antibodies against PAGE-4 and distribution of PAGE-4 in normal and malignant tissues

AIM: To prepare anti-PAGE-4 monoclonal antibody and to analyze the expression of PAGE-4 in normal tissues and some tumors. METHODS: BALB/c mice were immunized with recombinant PAGE-4 protein and the hybridoma cell lines secreting monoclonal antibodies (mAbs) against PAGE-4 were established using routine hybridoma technique. Western Blotting, ELISA and immunohistochemical staining were performed to identify the specificity and sensitivity of the PAGE-4 mAbs as well as the expression of PAGE-4 in normal tissues and some tumors. RESULTS: Four hybridoma cell lines secreting mAbs to PAGE-4 with high specificity and sensitivity were obtained. PAGE-4 was strongly expressed in spermatogonia and some of primary spermatocytes of testis, normal prostate and uterus in human reproductive tissues, and this molecule was also localized in normal pancreas. The expression of PAGE-4 molecule, a novel member of Cancer/Testis antigen family, was observed in such tumor tissues as those of prostate adenocarcinoma, uterus adenocarcinoma and clear cell renal adenocarcinoma. CONCLUSION: The mAbs against PAGE-4 were prepared which will facilitate further investigation of the distribution and function of PAGE-4 molecule.