人精子结合蛋白HBRP的抗体制备及定位

目的制备人类新的精子结合蛋白HBRP的单克隆抗体,确定其在精子的定位及其与精子功能相关的作用。方法分别利用PCR、Western blot、动物免疫、血清纯化等方法得到HBRP单克隆抗体,用ELISA方法检测抗体滴度;利用间接免疫荧光方法观察HBRP在精子的定位。结果经纯化鉴定获得HBRP融合表达蛋白;首次获得HBRP单克隆抗体;应用制备的单克隆抗体,确定HBRP主要结合在精子尾部的中段和主段。结论 HBRP结合在人成熟精子尾部的中段和主段,提示其参与精子运动功能的调控。

Antibody preparation and localization of human sperm binding protein

Objective To prepare the monoelonal antibody of novel human HBRP （bovine seminal plasma protein-related protein）, verify its localization and function in sperm cell. Methods Constructing the expression vector of PGEX-4T-1/hbrp, inducing the expression of hbrp gene and characterizing the protein by Western blot, immunization of the animals and purification of serum were performed to prepare the monoclonal antibody. ELISA experiment was performed to assay antibody titer. Indirect immunofluorescence is used to observe the localization of HBRP in sperm cell. Results HBRP fusion protein was expressed and purified in recombinant construction. HBRP monoclonal antibody was obtained by immunonization of animals and purification of serum. It was found that HBRP mainly located in the middle and the principle section of sperm cells by using HBRP monoclonal antibody. Conclusions Location of HBRP in the middle and the principle section of sperm cells suggest that HBRP might be involved in the regulation of sperm motility.