三株肺腺癌单克隆抗体的筛选和初步鉴定  

Screening and preliminary identification of three monoclonal antibodies to lung adenocarcinoma

作　　者：曲婵娟[1,2] 邵俊国[3] 易玲[1,2] 闫卓红[1,2] 王小珏[1,2] 韦攀健[1,2] 周丽娟[1,2] 张海青[1,2] 张洪涛[1,2] 

机构地区：[1]北京市结核病胸部肿瘤研究所 [2]首都医科大学附属北京胸科医院,北京101149 [3]河北医科大学附属第四医院,石家庄050011

出　　处：《中国免疫学杂志》2015年第7期922-926,共5页Chinese Journal of Immunology

基　　金：国家自然科学基金(30572123;81273209)资助

摘　　要：目的：制备肺腺癌单克隆抗体,筛选具有较高肺癌亲和性的单克隆抗体。方法：通过杂交瘤技术制备一组肺转移腺癌H2009细胞特异性单克隆抗体,以一定通量的细胞ELISA正、负筛选,初步确定抗体的肺癌特异性,经过色谱技术获得高纯抗体,免疫印迹确定抗体识别细胞膜蛋白分子量大小,应用流式细胞分析和免疫组化进一步分析对肺癌组织结合的特异性。结果：筛选出19株高结合肺腺癌细胞系,弱结合或不结合正常肺上皮细胞的杂交瘤,类别鉴定均为Ig M,通过亲硫色谱纯化杂交瘤培养上清获得高纯度抗体,其中三株抗体7C1、9H4和11C10均识别细胞膜蛋白,分子量介于20-110 k D,条带清晰,经病理组织验证7C1和9H4具有较好的肺癌组织亲和性,而对正常肺上皮阴性或弱染色;单抗11C10除结合肺癌上皮组织外,对肺正常上皮和乳腺正常上皮细胞也呈一定阳性反应,结合蛋白低水平泛表达于上皮来源细胞。结论：通量筛选出三株杂交瘤,初步确定具有较好的肺腺癌特异性,为深入探讨它们识别的抗原及相关的肺癌免疫研究奠定了基础。Objective： To prepare and screen monoclonal antibodies with a relatively high specific to lung adenocarcinoma. Methods： Mice were immunized with a lung adenocarcinoma cell line,which derived from metastatic lymph node,followed by hybridoma technique and a throughput screening via cellular ELISA against lung adenocarcinomas cell lines and normal bronchial epithelial cell line. Thiophilic chromatography was used to purify the hybridoma culture supernatant. Specific of monoclonal antibodies to lung adenocarcinoma were further evaluated by FACS analysis and Immunohistochemistry. The proteins which recognized by monoclonal antibodies were preliminarily identified by Western blot. Results： 19 hybridomas were selected on the basis of ELISA assay binding to H2009,but no or weakly binding to bronchial epithelial cell line. All of the m Abs were Ig M iostype. Among them 7C1,9H4 and 11C10 were then analyzed by Western blot,be identified to bind cell membrane proteins with molecular weight between 20-110 k D. A further immunohistochemical staining showed that 7C1 and 9H4 were more specific to lung adenocarcinoma,while the 11C10 with a less strength but perceptible specific to epithelial tissue. Conclusion： By throughput screening and the specific analysis to lung adenocarcinoma suggest the three m Abs are worthwhile to be further evaluated,and critically their recognized antigens to be identified.

关 键 词：肺腺癌 支气管上皮细胞 杂交瘤制备 免疫印迹 免疫组化 

分 类 号：R392[医药卫生—免疫学]