肝癌组织γ-谷氨酰转移酶单克隆抗体制备及ELISA方法的建立  

Preparation of monoclonal antibody against γ-glutamyltransferase of primary hepatic cancer tissue strongly bound to datura stramonim(DSA) lectin and establishment of ELISA method

作　　者：王念跃[1] 赵伟[1] 刘晨 张永臣 李勇[3] 端木浩[1] 

机构地区：[1]东南大学医学院附属南京第二医院,江苏南京210003 [2]北京晶美基因谷科技有限公司单克隆抗体制备中心,北京101113 [3]中国人民解放军南京军区总医院全军医学检验中心,江苏南京210002

出　　处：《中国生化药物杂志》2008年第2期89-92,共4页Chinese Journal of Biochemical Pharmaceutics

基　　金：南京市医学重点科技发展项目(No.ZKX0303)

摘　　要：目的应用纯化的人肝癌组织中蔓陀罗凝集素(DSA)强结合的γ-谷氨酰转移酶(GGT)制备单克隆抗体(McAb),并建立ELISA检测方法。方法应用亲和色谱法纯化肝癌组织中DSA强结合的GGT;采用单克隆抗体技术获得其McAb;protein A-Sepharose亲和色谱法纯化McAb,过碘酸钠法标记HRP法后建立ELISA方法。结果获得5株能特异性识别DSA强结合GGT的McAb细胞株,其中3株细胞所分泌McAb具有较高的特异性,应用所获得McAb进行HRP标记后,建立的ELISA方法,其检测灵敏度为10μg/L,批内及批间平均变异系数为8.9%和11.5%。结论成功制备了DSA强结合的GGT McAb杂交瘤细胞系,并建立了ELISA免疫学检测方法,为临床应用打下基础。Purpose To prepare monoclonal antibody（McAb） against γ-glutamyltransferase（GGT） of primary hepatic cancer tissue strongly bound to datura stramonim（DSA） lectin and establish an ELISA method. Methods The GGT from primary hepatic cancer tissue strongly bound to DSA Lectin（ DSA-GGT）was purified by Lectin affinity chromatography and the McAb was obtained by monoclonal antibody technique; the McAb was purified by the protein A-sepharose affinity chromatography and an ELISA examination method was established after the McAb labelled to horseradish peroxidase （HRP） by sodium periodate method. Results Results 5 strains which can produce the McAb specifically recognizing γ-glutamyltransferase strongly bound to DSA wereacquired, and the specificity of McAbs was high; the sensitivity of ELISA method which was built up after the purified McAb effectively labeled with HRP was 10μg/L. The average intra assay and inter assay precision were 8.9% and 11.5 % respectively. Conclusion the McAb hybridoma cell lines against γ-glutamyltransferase strong binding with DSA were successfully prepared and the examination method of the ELISA was successfully built up, which lay the foundation for the clinical application.

关 键 词：Γ-谷氨酰转移酶 单克隆抗体 肝细胞癌 色谱 ELISA 

分 类 号：Q78[生物学—分子生物学]