Longo F J
Department of Anatomy, University of Iowa, Iowa City 52242.
Dev Biol. 1989 Jan;131(1):37-43. doi: 10.1016/s0012-1606(89)80036-3.
Monoclonal antibodies have been generated to determine the fate of sperm-specific surface components of the sea urchin, Arbacia punctulata, subsequent to gamete fusion. Monoclonal antibody-7 (MAB-7) reacted with two polypeptide bands having apparent molecular masses of 33 and 35 kDa derived from the sperm plasma membrane; similar reactivity was not detected in egg preparations. Eggs and oocytes were prepared for electron microscopy at periodic intervals following insemination and reacted with MAB-7 followed by antimouse antibodies conjugated to colloidal gold. In samples prepared 30 to 60 sec postinsemination (PI) colloidal gold was confined to the surface of fused sperm with only a few gold particles associated with the egg plasma membrane. In later samples (2-10 min PI) label was present on increasingly greater areas of the egg/oocyte surface away from the site of gamete fusion so that by 20 min PI particles were located along one hemisphere of inseminated eggs and oocytes. These results demonstrate the incorporation of sperm plasma membrane polypeptides and their intermixing with egg plasmalemmal components subsequent to gamete membrane fusion. From measurements of labeled oocytes at different times PI, the diffusion coefficient for sperm surface polypeptides detected by MAB-7 was estimated to be 0.7 to 2.4 x 10(-9) cm2/sec.
已制备出单克隆抗体，以确定海胆（刺冠海胆）配子融合后精子特异性表面成分的命运。单克隆抗体 -7（MAB -7）与源自精子质膜的两条表观分子量分别为33 kDa和35 kDa的多肽带发生反应；在卵的制备物中未检测到类似的反应性。在授精后的不同时间间隔制备卵和卵母细胞用于电子显微镜观察，并与MAB -7反应，随后与缀合有胶体金的抗小鼠抗体反应。在授精后30至60秒（PI）制备的样品中，胶体金局限于融合精子的表面，只有少数金颗粒与卵质膜相关。在随后的样品中（授精后2 - 10分钟），标记物出现在远离配子融合位点的卵/卵母细胞表面越来越大的区域，以至于到授精后20分钟时，颗粒沿着受精的卵和卵母细胞的一个半球分布。这些结果表明配子膜融合后精子质膜多肽的掺入及其与卵质膜成分的混合。通过在不同授精后时间测量标记的卵母细胞，估计MAB -7检测到的精子表面多肽的扩散系数为0.7至2.4×10^(-9) cm²/秒。
