【摘要】 目的表达、纯化PPM1A-His融合蛋白及制备鼠多克隆抗体。方法将pBEX1-PPM1A-His原核表达质粒在大肠杆菌BL21(DE3)pLysS内进行诱导表达,用纯化的PPM1A-His融合蛋白免疫Balb/c小鼠制备多克隆抗体,并用ELISA、免疫组化和免疫荧光检测抗体的灵敏度和特异性。结果成功表达并纯化了PPM1A-His融合蛋白,纯化后纯度可达90%;ELISA法测定抗体效价为1∶100000;免疫组化和免疫荧光结果显示所制备的抗体可特异性检测肝和肝癌细胞的PPM1A。结论获得的PPM1A鼠多克隆抗体有较高的效价和特异性,为PPM1A在肝癌的相关研究打下了基础。更多还原

【Abstract】 Objective To express and purify PPM1A-His fusion protein and prepare PPM1A polyclonal antibody. Methods pBEX1-PPM1A-His was expressed in E.coli BL21 (DE3) pLysS under induction of IPTG. The recombinant protein was purified using Ni-NTA spin column. A polyclonal antibody was developed by immunizing Balb/c mice with the purified fusion protein,and the sensitivity and specificity of the antibody were tested by ELISA,immunohistochemisity,and immunofluorescence technique. Results A high titer (1∶100 000),high sensitivity,and high specificity polyclonal antibody which could recognize human PPM1A in liver tissues and hepatocellular carcinoma was prepared successfully. Conclusion PPM1A mouse polyclonal antibody possesses high sensitive and specificity,which can be used for further study of PPM1A in HCC or other diseases.更多还原