抗Der p 2单克隆抗体细胞株的建立与鉴定  

Establishment and Identification of Hybridomas Secreting Monoclonal Antibodies to Dermatophagoides pteronyssinus Ⅱ

作　　者：张红绪[1] 胡萍[1] 胡庚熙[2] 刘丹丹[1] 邢文会[1] 

机构地区：[1]河南师范大学生命科学学院,河南新乡453007 [2]中国科学院上海生命科学研究院生物化学与细胞生物学研究所,上海200031

出　　处：《河南师范大学学报（自然科学版）》2008年第4期136-139,共4页Journal of Henan Normal University(Natural Science Edition)

基　　金：河南省科技攻关项目(0624410030);河南省自然科学基金(0611033200);河南省动物重点学科资助

摘　　要：目的：建立Der p 2（屋尘螨Ⅱ类变应原）杂交瘤细胞株，并对其分泌的Der p 2单克隆抗体进行鉴定．方法：用重组Der p 2蛋白作为免疫原，免疫BALB／c小鼠，取脾细胞与Sp2／0小鼠骨髓瘤细胞融合，经筛选和3次克隆化，制备出单克隆抗体．采用间接ELISA法、Western blot法确定单克隆抗体的免疫球蛋白的类型和亚类、相对亲和力、特异性，对单克隆抗体进行鉴定．结果：获得3株能稳定分泌高效价Der p 2单克隆抗体的杂交瘤细胞株，186为IgG1类，1G11、4B1为IgG2a类．间接ELISA法检测细胞上清效价为10^4～10^5，腹水效价为10^5～10^6．Western blot试验证明3株单抗与Der p 2蛋白均有特异性反应，结论：成功制备了3株抗Der p 2的单克隆抗体，均具有良好的特异性和亲和力，为进一步建立Der p 2蛋白检测和纯化的方法奠定了基础．Objective： to establish Der p 2 （Dermatophagoides pteronyssinus Ⅱ ） hybridoma cell lines, and to perform elementary identification of monoclonal antibodies （Me Abs）. Methods： BALB/c mice are immunized with recombinant protein Der p 2, and hybridoma is generated with mouse spleen cells and myeloma Sp2/0 cells. After fusions and three cloning, three hybridoma cell lines secreting monoclonal antibodies against Der p 2 have been obtained. The monoclonal antibodies are identified by the immunoglobulin class, relative affinity and characterization determined by indirect enzyme-linked immunosorbent assay （ELISA） and Western blot. Results： three hybridoma cell lines that stably secreted Der p 2 Me Abs are obtained： 1B6 belonging to IgG1, 1G11 and 4B1 belonging to IgG2a. The titers of cell supernatant are 10^4-10^5, and the titers of ascites were 10^5-10^6 , which are showed by indirect ELISA. Characterization is determined by Western blotting. Conclusion： three monoclonal antibodies against Der p 2 protein with high specificity and affinity have been generated successfully, which may provide a basis for further establishing methods to detect and purify Der p 2 protein.

关 键 词：Der P 2 单克隆抗体 杂交瘤 细胞融合 

分 类 号：Q813.2[生物学—生物工程]