前列腺素D合成酶单克隆抗体的制备及其在精子中的定位

Lipocalin-type prostaglandin D synthase:preparation of monoclonal antibodies and localization on human sperm

目的制备小鼠抗人Lipocalin型前列腺素D合成酶的单克隆抗体,探讨Lipocalin型前列腺素D合成酶在精子表面的定位与分布。方法用毕氏酵母表达的Lipocalin型前列腺素D合成酶作抗原免疫BALBc小鼠,制备杂交瘤,产生单克隆抗体,ELISA与Westernblot分析鉴定其特异性,免疫组织化学法探讨Lipocalin型前列腺素D合成酶在人精子表面的分布与定位。结果获得了1株抗人Lipocalin型前列腺素D合成酶的单克隆抗体,其效价为1∶1000,抗体亲和力为2×108molL,小鼠IgG亚型为IgG1,Westernblot结果显示该抗体能特异性识别Lipocalin型前列腺素D合成酶,组化结果显示Lipocalin型前列腺素D合成酶定位于人精子表面。结论成功制备了1株鼠抗人Lipocalin型前列腺素D合成酶的单克隆抗体,精子的表面有Lipocalin型前列腺素D合成酶的分布。

Objective:To prepare monoclonal antibodies against lipocalin type prostaglandin D synthase and to explore the distribution of lipocalin type prostaglandin D synthase on human sperm.Methods:BALB/c mice were immunized with lipocalin type prostaglandin D synthase,monoclonal antibodies were prepared by hybridoma technique,the biological characteristics of antibodies were analyzed by ELISA and Western blot,the distribution of lipocalin type prostaglandin D synthase on sperm were performed by immunohistochemistry.Results:A strain of hybridoma cell was obtain,and the IgG isotype of the antibody was identified as IgG1,with affinity constat(K) of 2×108 and titer over 103.Western blot demonstrated specific recognition of lipocalin type prostaglandin D synthase by the obtained monoclonal antibody.The result of immunohistochemistry displayed that lipocalin type prostaglandin D synthase was mainly distributed on the surface of human sperm.Conclusion:A hybridoma cell secreting anti-lipocalin type prostaglandin D synthase monoclonal antibodies stably was established.On the surface of human sperm,there is lipocalin type prostaglandin D synthase's distribution. [