Cariño Cecilia, Prasad Sarvamangala, Skinner Sheri, Dunbar Bohn, Chirinos Mayel, Schwoebel Eric, Larrea Fernando, Dunbar Bonnie
Department of Reproductive Biology, The National Institute of Medical Sciences and Nutrition Salvador Zubirán, Mexico City, Mexico.
Reprod Biomed Online. 2002 Mar-Apr;4(2):116-26. doi: 10.1016/s1472-6483(10)61928-1.
The mammalian zona pellucida (ZP) consists of three glycoproteins (ZP1, ZP2 and ZP3), which are variably conserved among species at the genomic and amino acid levels. In order to evaluate the expression of ZP during ovarian development, a population of antibodies was selected that recognize species conserved antigenic domains of the three ZP proteins. Domain specific antibodies were selected from sera of rabbits immunized with all three native pig ZP proteins by elution of antibodies bound to each of the three human ZP recombinant proteins expressed from cDNAs, using the baculovirus expression system in insect cells. Immunoblot analysis was used to characterize the specificity of the antibodies and immunohistochemistry was used to evaluate the stage specific expression of ZP proteins during ovarian follicular development of the mouse, baboon and human. This study demonstrates that the conserved domains of all three ZP proteins are localized in the oocyte extracellular ZP matrix as well as in a subset of granulosa cells. However, this expression does vary among species with respect to the stage and cell type during early stages of ovarian follicular development. These antibodies should serve as excellent markers for evaluating early stages of human ovarian follicular development and in the development of contraceptive agents.
哺乳动物透明带（ZP）由三种糖蛋白（ZP1、ZP2和ZP3）组成，它们在基因组和氨基酸水平上在不同物种间的保守程度各异。为了评估卵巢发育过程中ZP的表达情况，我们筛选了一组抗体，这些抗体能够识别三种ZP蛋白的物种保守抗原结构域。通过从用所有三种天然猪ZP蛋白免疫的兔血清中，利用杆状病毒表达系统在昆虫细胞中表达的三种人ZP重组蛋白，洗脱与之结合的抗体，从而筛选出结构域特异性抗体。免疫印迹分析用于鉴定抗体的特异性，免疫组织化学用于评估小鼠、狒狒和人卵巢卵泡发育过程中ZP蛋白的阶段特异性表达。本研究表明，所有三种ZP蛋白的保守结构域都定位于卵母细胞的细胞外ZP基质以及一部分颗粒细胞中。然而，在卵巢卵泡发育早期，这种表达在不同物种的阶段和细胞类型方面确实存在差异。这些抗体应可作为评估人类卵巢卵泡发育早期阶段以及开发避孕药物的优良标志物。
