【摘要】 目的:建立针对多药耐药人乳腺癌细胞膜表面差异抗原的抗体库,并鉴定抗角蛋白CK8特异性的单克隆抗体。方法:采用细胞免疫和杂交瘤技术制备单克隆抗体库,经免疫沉淀得到抗原-抗体复合物,SDS-PAGE分离目的抗原,质谱测序,Western blot验证及激光共聚焦显微镜观察抗原分子的细胞膜定位。结果:成功筛选到稳定分泌抗CK8分子的单克隆抗体的杂交瘤细胞株(7F9)。结论:文中所研制的单克隆抗体具有与乳腺癌细胞膜表面CK8分子特异结合的活性,在上皮来源肿瘤标志物CK8的临床检测及CK8相关生物学活性研究中具有广泛应用前景。更多还原

【Abstract】 Objective:To identify tumor associated antigens or the drug resistance-related proteins presented on membrane of a multidrug resistant human breast cancer cell line.Methods:BALB/c mice were immunized with intact multidrug resistant breast cancer cell lines and the splenocytes from immunized mice were fused with mouse myeloma cell SP2/0.Hybridoma cells were screened by ELISA.Several monoclonal antibodies were developed and followed by protein G affinity chromatograph,One of these McAb was selected for further investigation,which showed strongly membrane staining by laser confocal microscopy.Antigen-antibody complex was obtained by immunoprecipitation and separated by SDS-PAGE.The specific antigen was identified by MALDI-TOF/TOF MS sequencing and verified by reciprocal immunoprecipitation and western blot analysis using commercial monoclonal antibody.Results:One of hybridoma cell lines(7F9) secreting specific McAb against CK8 were developed.Conclusion:The novel monoclonal antibody may be used as the tool for research of biological activities about CK8 and specific detection of membrane protein CK8 which is apotential tumor marker of epithelial derived tumors.更多还原