CAJ | 학술논문

目的制备并鉴定抗多形上皮黏蛋白1(MUC1)单克隆抗体,将抗体应用于人乳腺癌组织、纤维腺瘤、囊性增生症及正常乳腺组织的临床鉴别,采用免疫组化方法进行检测.方法用合成的MUC1重复序列短肽与钥孔血蓝蛋白(KLH)及牛血清白蛋白(BSA)偶联后作为抗原免疫Balb/c小鼠,杂交瘤技术筛选出稳定分泌单克隆抗体的细胞株.制备单克隆抗体,以ELISA、Western blot、流式细胞术、免疫荧光法对抗体进行特异性鉴定及亚类分析;将抗体应用于人乳腺癌组织、纤维腺瘤、囊性增生症及正常乳腺组织的临床鉴别,采用免疫组化方法进行检测.结果获得4株稳定分泌抗MUC1的杂交瘤细胞株,第1株反应性最强,效价1∶10 7以上.抗体亚型3株为IgG1,1株为IgM,轻链均为κ型.经Western blot、流式细胞术、免疫荧光、免疫组化检测证实抗体与乳腺癌细胞株MCF-7特异性结合,单抗检测MUC1在乳腺癌组织与纤维腺瘤、囊性增生症及正常乳腺组织之间表达差异明显(均P＜0.01).结论抗MUC1单克隆抗体经检测效价高、特异性强,为乳腺癌早期诊断和靶向治疗的进一步研究奠定了基础.
목적 제비병감정항다형상피점단백1(MUC1)단극륭항체,장항체응용우인유선암조직、섬유선류、낭성증생증급정상유선조직적림상감별,채용면역조화방법진행검측.방법 용합성적MUC1중복서렬단태여약공혈람단백(KLH)급우혈청백단백(BSA)우련후작위항원면역Balb/c소서,잡교류기술사선출은정분비단극륭항체적세포주.제비단극륭항체,이ELISA、Western blot、류식세포술、면역형광법대항체진행특이성감정급아류분석;장항체응용우인유선암조직、섬유선류、낭성증생증급정상유선조직적림상감별,채용면역조화방법진행검측.결과 획득4주은정분비항MUC1적잡교류세포주,제1주반응성최강,효개1∶10 7이상.항체아형3주위IgG1,1주위IgM,경련균위κ형.경Western blot、류식세포술、면역형광、면역조화검측증실항체여유선암세포주MCF-7특이성결합,단항검측MUC1재유선암조직여섬유선류、낭성증생증급정상유선조직지간표체차이명현(균P＜0.01).결론 항MUC1단극륭항체경검측효개고、특이성강,위유선암조기진단화파향치료적진일보연구전정료기출.
Objective To prepare the anti-human MUC1 monoclonal antibody (mAb) and to apply it in human breast cancer tissues,fibro adenoma,cystic hyperplasia and normal breast tissues,using immunohistochemistry assay.Methods Synthetic MUC1 repetitive sequence short peptide coupled with KLH protein and BSA protein as antigen to immunize Balb/c mice,cell lines capable of secreting monoclonal antibodies were screened by hybridoma cell fusion technique.The specificities and subclasses of the prepared monoclonal antibodies were identified and analyzed by ELISA,Western blot,flow cytometry and immunofluorescence test.The antibodies were used in the detection of clinical human breast cancer,benign fibro adenoma and normal breast tissue by immunohistochemical methods.Results Four hibridoma cell lines secreting specific anti-MUC1 mAbs were obtained,The titers of the most powerful antibody were above 1 ∶ 107.3 strains were of IgG1 antibody,one was of IgM,and the light chain of which was categorized into chain to.The mAb could specifically bind to the MCF-7 cells,which was verified by Western blot,flow cytometry analysis,fluorescence immunoassay and immunohistochemistry test.Significant differences of expression (all P ＜ 0.01) of the detection of MUC1 monoclonal antibodies were found in breast cancer tissue,fibro adenoma,cystic hyperplasia and normal breast tissues.Conclusions The mAb against MUC1 developed in this study is of high titers and specific for breast cancer,exhibiting a favourable prospect for early diagnosis and targeted therapy of breast cancer.