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目的制备癌-睾丸抗原OY-TES-1多克隆抗体,并进行鉴定。方法以大肠杆菌表达的MBP/OY-TES-1融合蛋白作为抗原,免疫新西兰白兔,制备抗血清。经蛋白A凝胶纯化后,采用ELISA法检测抗血清的效价,Western blot和免疫组化法检测多克隆抗体的特异性。结果制备的抗血清经ELISA检测,效价不低于1∶128000,纯化后,其效价不低于1∶8000。Western blot显示,抗OY-TES-1多抗可与MBP/OY-TES-1融合蛋白、MBP、OY-TES-1重组蛋白和睾丸组织总蛋白特异性结合;免疫组化检测显示,精子头部呈现阳性反应。结论已成功制备了效价高、特异性强的癌-睾丸抗原OY-TES-1多克隆抗体,为深入研究OY-TES-1的生物学功能提供了工具。
Objective To prepare polyclonal antibody of OY-TES-1, a cancer-testis antigen, and identify it. Methods The New Zealand white rabbits were immunized with MBP / OY-TES-1 fusion protein expressed in E.coli as antigens to prepare antiserum. After purification by Protein A gel, the antiserum titer was detected by ELISA, and the specificity of polyclonal antibody was detected by Western blot and immunohistochemistry. Results The antiserum was prepared by ELISA. The titer was no less than 1:128000. After purification, its titer was no less than 1: 8000. Western blot showed that anti-OY-TES-1 polyclonal antibody could specifically bind to MBP / OY-TES-1 fusion protein, MBP, OY-TES-1 recombinant protein and total protein in testis tissue. Immunohistochemistry showed that sperm head Ministry showed a positive reaction. Conclusion The polyclonal antibody OY-TES-1 with high titer and specificity has been successfully prepared, which provides a tool for further study on the biological function of OY-TES-1.