人抑制素βA亚基单克隆抗体的制备和鉴定  

Preparation and identification of human inhibin β~A subunit monoclonal antibody

作　　者：黄新[1] 夏辉明[1] 张东升[1] 高晓东[1] 宫亚欧[1] 鲁桂琛 孙颖 

机构地区：[1]河南医科大学病理生理学教研室,郑州4500522 [2]中国医学科学院药物研究所合成室,北京100050

出　　处：《河南医科大学学报》1999年第4期5-7,共3页Journal of Henan Medical University

基　　金：卫生部科学研究基金 ;河南省自然科学基金资助项目

摘　　要：目的：制备人抑制素βＡ（ＩＮＨβＡ） 亚基多肽单克隆抗体（单抗，ＭｃＡｂ）。方法和结果：用固相多态合成法合成了人抑制素βＡ亚基１ － ２８，８２ － １１４ 多肽片段。以粗提的猪卵泡液抑制素（ＩＮＨ） 和合成肽段纯品作为抗原免疫ＢＡＬＢ／ｃ 小鼠，经细胞融合与筛选，获得４ 株杂交瘤细胞株，ＥＬＩＳＡ检测腹水效价为１×１０－ ４ ～１ ×１０－ ５ ，抗体特异性试验表明与α亚基片段及其他多肽激素无交叉反应。表位分析表明３ 株ＭｃＡｂ（ 抗ＩＮＨβＡ（８２－ １１４）） 可能针对同一抗原位点。相对亲和力试验表明４Ｇ３＞ ３Ｂ７＞ ６Ｃ５ ＞２Ａ３。抗原竞争试验表明腹水中抗体能明显被人卵泡液和猪卵泡液中和， 而绝经期妇女血清不能中和抗体活性。Aim:To prepare the McAbs againsting the INH βA subunit and to identify them.Methods and Results:The fragment of inhibin subunits βA(1-28)and βA(82-114) have been synthesized manually by stepwise solid phase procedure on Pam resin support in Boc chemistry, and purified by HPLC.Inhibin isolated from pocine follicle fluid and synthetic polypeptide fragment were used as antigen.Intraperitoneal immunizationwas performed on BALB/c mice, then boosted intraspleenic injection was performed 3 days prior to fusion. After fusion,selection and cloning procedures, 4 hybridoma celllines were successfully obtained. Theascites titer, specificity of McAb were identified.Epitope specificity analysis proved that the 3 McAbs (against INHβA(82-114)) recognized the same epitope. Relative affinity of 4 hybridoma cell lines were 4G3>3B7>6C5>2A3. Antigen competitive inhibition test shows that antibodies can be blocked by preincubating with human follicle fluid or pocine follicle fluid, but the serum of postmenopausal woman can not block immunoreaction. Conclusions :The cell lines estabolished can secret the INH βA subunit antibodies having high specificity.

关 键 词：抑制素βA亚基 人工合成多肽 单克隆抗体 免疫 

分 类 号：R392－33[医药卫生—免疫学]