人乳腺珠蛋白单克隆抗体制备及生物特性鉴定  

Preparation of monoclonal antibody against human mammaglobin and identification of its biological characterization

作　　者：杨锡琴[1] 吕晓娟[2] 王国华[1] 黄焰[2] 宋晓国[1] 陈堃[1] 修冰水[1] 冯晓燕[1] 张贺秋[1] 

机构地区：[1]军事医学科学院基础医学研究所,北京100850 [2]解放军307医院,北京100071

出　　处：《中国实验诊断学》2012年第11期1965-1968,共4页Chinese Journal of Laboratory Diagnosis

基　　金：国家高技术研究发展计划(863)重大项目(SQ2010AA220970001)

摘　　要：目的构建重组hMAM蛋白,通过杂交瘤技术获得特异性的抗hMAM的单克隆抗体,并鉴定其生物特性。方法克隆表达hMAM 8-93AA、20-52AA、43-70AA、56-82AA、67-93AA区段的抗原,以纯化的pBV220/hMAM(8-93AA)为免疫原,杂交瘤技术筛选出能稳定分泌抗hMAM单克隆抗体的细胞株。采用Western blotting和免疫组织化学染色方法鉴定其表位和特异性。结果获得6个高纯度的重组抗原,通过pBV220/hMAM(8-93AA)免疫小鼠后,获得2株稳定分泌抗hMAM抗体的杂交瘤细胞株。Western blotting结果显示MAb1、MAb2分别是识别hMAM20-52AA、hMAM52-67AA片段的特异性单克隆抗体。免疫组织化学染色结果显示该抗体仅在乳腺组织中呈阳性反应,在其他肿瘤组织中不反应。结论通过杂交瘤技术成功制备了两株稳定分泌且高特异性的抗hMAM不同位点的单克隆抗体细胞株,为进一步研制高灵敏的乳腺癌早期检测试剂奠定了基础。Objective Preparing hMAM recombinant protein to obtain specific anti-hMAM monoclonal antibody by hybridoma technology,and identify their biological characterization.Methods Cloning and expression of the fragment of hMAM 8-93AA,20-52AA,43-70AA,56-82AA,67-93AA antigens,purified pBV220/hMAM（8-93AA） as immunogen,hybridoma technology can be selected monoclonal antibody secreting anti hMAM cell lines.Using Western blotting and immunohistochemical staining to identify the epitope and specificity.Results Six recombinant antigen of high purity were obtained.Two hybridoma cell lines were obtained after the mice were immunized by pBV220/hMAM（8-93AA）.Western blotting showed MAb1,MAb2 were identified hMAM20-52,hMAM52-67AA fragment-specific monoclonal antibodies.Immunohistochemical staining showed that the antibody only in the breast tissue was positive,in other tumor tissue was negative.Conclusion Two high-secreting specific anti-hMAM different sites of monoclonal antibody cell lines have been successfully prepared by hybridoma technique to lay the foundation of further develop a highly sensitive early breast cancer detection kit.

关 键 词：人乳腺珠蛋白 杂交瘤技术 单克隆抗体 

分 类 号：R737.9[医药卫生—肿瘤]