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目的制备抗hMOF的单克隆抗体并对其生物学特性进行鉴定。方法以重组GST-hMOF融合蛋白作为免疫原免疫BALB/c小鼠,取其脾细胞与SP2/0细胞融合,经多次筛选及克隆化,建立可以稳定分泌抗hMOF单克隆抗体的杂交瘤细胞株。用ELISA、Western blot、免疫组化对此抗体特性进行鉴定。结果筛选到1株能稳定分泌抗hMOF单克隆抗体的细胞株4C1C8,亚类鉴定单抗为IgG1。ELISA法测定饱和硫酸铵盐析抗体效价为1∶409600,抗体相对亲和力为7.65×106L/mol;Western blot显示抗体能特异识别免疫原;细胞免疫组化显示此单抗可以特异的识别正常细胞株HL7702表达的hMOF。结论成功制备抗hMOF单克隆抗体,为进一步研究hMOF功能及机制奠定了基础。
Objective To prepare anti-hMOF monoclonal antibody and identify its biological characteristics. Methods The recombinant GST-hMOF fusion protein was used as immunogen to immunize BALB / c mice. The spleen cells were fused with SP2 / 0 cells. After multiple screening and cloning, the hybridoma cells secreting anti-hMOF monoclonal antibody Strain. The characteristics of this antibody were identified by ELISA, Western blot and immunohistochemistry. Results A strain of 4C1C8 cell line stably secreting anti-hMOF monoclonal antibody was screened. The subclass of McAb was IgG1. The titer of the saturated ammonium sulfate salting-out antibody was 1:409600 by ELISA and the relative affinity of the antibody was 7.65 × 106 L / mol. Western blot showed that the antibody could specifically recognize the immunogen. Immunocytochemistry showed that the monoclonal antibody could specifically recognize normal cells Strain HL7702 expressed hMOF. Conclusion The successful preparation of anti-hMOF monoclonal antibody laid the foundation for further study of hMOF function and mechanism.