Hemizona Assay in Rhesus Monkeys:Influences of SpermMotility and Oocyte Maturational Stages onSperm/Egg Binding

The zona pellucidae is the first important position in sperm/oocyte interactions.Sperm binding to the zona pellucidae is an initial and critical step in fertilization,which is relevant to both of sperm motility and oocyte maturational stage. However,until recently there was no ideal model to predict fertilization potential of spermand to reveal the mechanism of sperm/oocyte interactions. The hemizona assay (HZA)was first established in human in vitro fertilization(IVF) by using the two matchingzona hemispheres with functional equality which provides many obvious advanages incomparative research of sperm/oocyte binding and interaction. In human IVF, the re-sults have certainly verified the feasibility of the HZA as a test to predict

人卵透明带蛋白ZP3a和ZP3b肽段的免疫原性及其抗血清体外抑制人精子-半透明带结合

分析大肠杆菌表达的重组人卵透明带-3(r-huZP3)蛋白两个肽段(r-huZP3a22-176及r-huZP3b177-348)的免疫原性,比较两者抗血清体外抑制人精子-半透明带结合的能力。以制备性聚丙烯酰胺凝胶电泳(PAGE)纯化的大肠杆菌表达蛋白为抗原, 主动免疫新西兰兔产生抗huZP3a22-176及huZP3b177-348抗血清;通过ELISA测定比较两者的抗体应答水平:以蛋白印迹和免疫组化方法测定两者抗血清同重组表达蛋白、天然人卵ZP以及卵巢组织的反应性;通过竞争性半透明带结合试验 (hemizona assay,HZA)观察两者抗血清体外抑制人精子-ZP结合能力。结果显示:未与大分子蛋白载体耦联的r-huZP3a22-176 和r-huZP3b177-348抗原都在免疫兔中产生了较高的抗体滴度,而且它们的抗血清可识别大肠杆菌表达的各自重组ZP3肽以及人卵细胞表面天然ZP,两者抗血清也都能在体外抑制人精子-ZP结合。由此可见,r-huZP3a22-176及r-huZP3b177-348蛋白具有良好免疫原性,所产生抗血清也都显示出细胞和组织特异性。因此,单一或合并两个huZP3肽段均可作为抗原研制检测不明原因性不孕妇女中是否存在抗自身ZP抗体的诊断试剂盒,另外它们的抗血清也能用于鉴定已知huZP3表位肽段的最小B-细胞表位基序。