重组人卵透明带蛋白及其多克隆抗体对精子-透明带结合的影响

为了探讨毕赤酵母表达的重组人卵透明带ZP3蛋白(recombinant human zona pellucida-3 protein,rhZP3)及其多克隆抗体对小鼠和人精卵结合的影响,采用不同浓度的rhZP3以及空白培养液分别处理小鼠精子,然后再与小鼠卵子进行结合实验,观察经过不同处理的精子对透明带黏附及体外受精率的影响;用rhZP3以及空白培养液分别处理人精子,然后再与人卵子进行结合实验,观察经过不同处理的精子对透明带黏附的影响;用抗rhZP3抗体与阴性血清分别处理小鼠和人卵子,再与精子进行结合实验,观察多克隆抗体对精子粘附以及小鼠体外受精率的影响。实验结果表明,rhZP3和抗rhZP3多克隆抗体既能抑制人的体外精卵结合,也能抑制小鼠体外精卵结合,提示rhZP3具有天然透明带的特性,有发展成避孕疫苗和作为检测透明带抗体检测试剂的可能。

Effect of the recombinant human zona pellucida protein and its polyclonal antibodies on the sperm-zona pellucida binding

To investigate the effect of purified recombinant human zona pellucida protein-3(rhZP3) expressed in Pichia pastoris and its polyclonal antibodies on the sperm-zona pellucida binding,the mouse sperm treated with culture medium containing different concentration of rhZP3 or without were incubated with zona-intact oocytes and matched the number of sperm bound per zona pellucida and ratio of fertilization.The rhZP3-exposed and non-exposed human sperm were incubated with zona-intact oocytes and observed the number of sperm bound per zona pellucida.Meanwhile,the oocytes of humans and mouse pre-treated with anti-rhZP3 polyclonal antibodies and negative serum control were co-incubated with homogeneous sperm respectively,and the number of sperm bound per zona pellucida as well as the ratio of mouse in vitro fertilization were estimated.The experimental data demonstrated that both rhZP3 and its polyclonal antibodies could interfere with the sperm-zona pellucida binding of humans and mouse.These results imply that rhZP3 exhibits the same characteristics as those of the natural human zona pellucida,thus affording the possibility for developing contraceptive vaccines and kits which may be used to detect the presence of antibodies against zona pellucida.