抗人晚期糖基化终产物受体胞外段不同表位单克隆抗体的制备

南方医科大学学报 ›› 2004, Vol. 24 ›› Issue (02): 129-132.

抗人晚期糖基化终产物受体胞外段不同表位单克隆抗体的制备

朱平¹, 唐磊¹, 赵善超², 卢晓¹, 候凡凡², 富宁¹

1. 第一军医大学免疫教研室, 广东, 广州, 510515;
2. 第一军医大学南方医院肾内科, 广东, 广州, 510515

出版日期:2004-02-20 发布日期:2004-02-20
基金资助:
收稿日期:2004-1-12。
基金项目:国家自然科学基金重点项目(30330300);广东省团队项目(010717)
作者简介:朱平(1975-),女,河南南阳人,助理实验师
通讯作者:富宁,电话:020-61648221,E-mail:zfzn@fimmu.com

Preparation of monoclonal antibodies against different epitopes on extracellular domain of human receptor for advanced glycation end product

ZHU Ping¹, TANG Lei¹, ZHAO Shan-chao², LU Xiao¹, HOU Fan-fan², FU Ning¹

1. 第一军医大学免疫教研室, 广东, 广州, 510515;
2. 第一军医大学南方医院肾内科, 广东, 广州, 510515

Online:2004-02-20 Published:2004-02-20

摘要/Abstract

摘要： 目的制备抗人晚期糖基化终产物受体(RAGE)胞外段单克隆抗体。方法用纯化人重组RAGE胞外段(氨基酸序列23~342)免疫Balb/c小鼠,取其脾细胞与NS-1融合制备杂交瘤,经筛选和两次克隆化,从制备的腹水中纯化单克隆抗体。结果和结论 获得两株杂交瘤细胞B2.2和E10,其分泌单抗的亚型均为IgG2b。经ELISA、Western blot和流式细胞仪鉴定,证明两株单抗均可结合重组RAGE及表达于细胞表面的RAGE,且两株抗体识别的位点为远隔表位,所建立的双夹心法可用于检测可溶性重组RAGE。这两株抗体将为研究RAGE相关疾病提供有效的工具。

Abstract: Objective To prepare and characterize monoclonal antibodies (mAb) against recombinant human receptor for advanced glycation end product (rhRAGE). Methods BALB/c mice were immunized with recombinant extracellular domain(amino acid 23-342) of human advanced glycation end product (RAGE), and hybridoma was generated with mouse spleen cells and myeloma NS-1 cells. After three fusions and cloning, two hybridoma cell lines secreting monoclonal antibodies to RAGE were obtained and monoclonal antibodies were purified from the ascites followed by characterization with indirect enzyme-linked immunosorbent assay (ELISA), flow cytometry and Western blotting. Results and Conclusion Two hybridoma cell lines secreting anti-RAGE mAb were established and designated as B2.2 and E10, respectively. Both of mAb B2.2 and E10 belonged to IgG2b isotype and could bind to recombinant RAGE and natural RAGE expressed on THP-1 cells. In addition, B2.2 and E10 could recognize different epitopes of RAGE, which were conformed to be capable of detecting soluble recombinant RAGE in sandwich ELISA. These two mAbs against different epitopes of rhRAGE would be useful for study of glycation end product and RAGE-related diseases.

中图分类号:

R392.11

朱平¹, 唐磊¹, 赵善超², 卢晓¹, 候凡凡², 富宁¹. 抗人晚期糖基化终产物受体胞外段不同表位单克隆抗体的制备[J]. 南方医科大学学报, 2004, 24(02): 129-132.

ZHU Ping¹, TANG Lei¹, ZHAO Shan-chao², LU Xiao¹, HOU Fan-fan², FU Ning¹. Preparation of monoclonal antibodies against different epitopes on extracellular domain of human receptor for advanced glycation end product[J]. Journal of Southern Medical University, 2004, 24(02): 129-132.

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