人白细胞介素15单克隆抗体的制备及鉴定

目的 :用rhIL 15与沙门氏菌裸菌相偶联制备IL 15单克隆抗体 (monoclonalantibody ,McAb) ,以便为疾病的诊断、治疗和发病机制的研究提供可靠的生物制剂。方法 :将纯化的rhIL 15蛋白与沙门氏菌裸菌相偶联制成免疫原 ,用脾内、腹腔、静脉三种途径相结合免疫BALB c小鼠 ,以PEG为促融剂 ,将脾细胞与SP2 0细胞进行融合 ,HAT选择培养 ,间接ELISA筛选阳性克隆 ,通过多次克隆化 ,获得稳定分泌特异性McAb的杂交瘤细胞株 ,并对所获得的细胞株及分泌的McAb特性进行了分析。结果 :获得 4株能稳定分泌特异性McAb的杂交瘤细胞系 (hybridomacell,Hc) ,ELISA法检测其效价分别为 1:10 6 、1:10 7、1:10 7、1:10 7。Dotblot检测IL 15的灵敏度为 1 5ng ,其中一株 (1 7E4 )尚可用于Westernblot检测。结论 :成功制备了 4株IL 15McAb杂交瘤细胞系。

Preparation and identification of human interleukin-15 monoclonal antibody

Objective:To produced mouse monoclonal antibodies against IL 15 that should be a valuable reagent for diagnosis and investigation into the pathological significance of IL 15 protein expression Methods:rhIL 15 was conjugated to naked bacteria of salmonella typhimurium as immunogen to immunize BALB/c mice The spleen cells were fused with SP2/0 cells using polyethlene glycol, suspended in selective growth medium (HAT) Clones secreting specific monoclonal antibody were screened by indirect enzyme linked immunosorbent assay(ELISA) After several cloning,several hybridoma cell clones stably producing anti IL 15 monoclonal antibody were obtainied The McAb was identified by Western blot with goat polyclonal antibody raised against mouse Results:Four hybridoma cell strains which could produce monoclonal antibodies to IL 15 were obtained The chromosome number of hybridoma cells was between 90 108, there were metacentric chromosome and submetacentric chromosome in hybridoma cells Monoclonal antibodies were able to inhibit and/or neutralize the effects of IL 15, while naked bacteria could not The sensitivity of Dot blot was 1 5 ng Conclusion:Monoclonal antibodies to rhIL 15 have been prepared and characterized