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目的:用OAS1蛋白免疫小鼠,获得OAS1特异性单克隆抗体,为OAS1的含量测定提供基础。方法:通过全基因合成的方法获得目的基因序列,转化大肠杆菌BL21细胞诱导His-OAS1及OAS1蛋白表达,纯化后用作抗原免疫小鼠,取脾融合,筛选稳定分泌抗体的阳性细胞株,制备并纯化单抗,通过SDS-PAGE,ELISA,Western blot等方法进行检测。结果:体外高效表达OAS1蛋白,并成功制备特异性单克隆抗体,效价在5×10-11mol/L以上,亲和常数为3.37×108 L.mol-1。结论:获得高亲和力OAS1单克隆抗体,为其含量的检测奠定了基础。
OBJECTIVE: To immunize mice with OAS1 protein to obtain OAS1 specific monoclonal antibody, which provides the basis for the determination of OAS1. Methods: The gene sequence was obtained by whole gene synthesis. The recombinant plasmid was transformed into E. coli BL21 cells to induce the expression of His-OAS1 and OAS1 protein. After purified, the recombinant plasmid was used as antigen to immunize mice. The monoclonal antibodies were purified and detected by SDS-PAGE, ELISA and Western blot. RESULTS: OAS1 protein was highly expressed in vitro and specific monoclonal antibodies were successfully prepared. The titer was above 5 × 10-11mol / L and the affinity constant was 3.37 × 108 L. mol-1. Conclusion: Obtaining high affinity OAS1 monoclonal antibody lays a foundation for the determination of its content.