抗Der p 2单克隆抗体细胞株的建立与鉴定

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,1B6为IgG1类,1G11、4B1为IgG2a类.间接ELISA法检测细胞上清效价为104~105,腹水效价为105~106.Westernblot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.

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

Objective: to establish Der p 2(Dermatophagoides pteronyssinus Ⅱ) hybridoma cell lines,and to perform elementary identification of monoclonal antibodies(Mc 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 Mc Abs are obtained: 1B6 belonging to IgG1,1G11 and 4B1 belonging to IgG2a.The titers of cell supernatant are 104~105,and the titers of ascites were 105~106,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.