Immunocytochemical and ultrastructural study on the condition of Fertilization.

采用冷冻破碎法研究了几种哺乳动物附睾各部位精子的膜内颗粒（Imps）的变化。一般来说，顶体周围质膜（PAPM）可以区分大小imp。在仓鼠、大鼠、家兔和公猪的精子中，小的IMP暂时形成六角形排列，但在人类精子中没有发现，其IMP密度较低。根据这些观察，imp模式的形成可能需要以下条件：(1)相同大小的大量IMP的存在，(2)IMP的运动，(3)IMP聚集因素的存在，如它们运动的障碍。从附睾起始段提取的仓鼠精子，其膜甾醇复合物（FSCs）的密度为390/m ^2，后来增加到约500/m ^2。当精子通过头弯曲时。FSC密度的增加可能有助于膜的稳定。用Tyrode溶液孵育金仓鼠精子，观察其体外获能过程中精子质膜的变化。孵育2小时后，PAPM中的FSC密度降低了70-80%。孵育3小时后，当精子被能化时，大部分小IMP消失，PAPM内形成较宽的IMP自由区。从小鼠分离的生精细胞中提取蛋白，制备单克隆抗体。其中一种抗体13D3识别hsc7l， hsc7l属于7OkDa热休克蛋白家族。间接免疫荧光显示，hsc7l在幼生精细胞的细胞质中普遍存在，但定位于精子的中间部分。

Changes in the intramembranous particles (Imps) of the sperm plasma membrane during epididymal transport Spermatozoa taken from the various parts of the epididymis were studied by freeze fracture in several mammalian species. In general, large and small IMPs could be distinguished in the peri-acrosomal plasma membrane (PAPM). In hamsters, rats rabbits and boars, hexagonal arrangement was temporally formed by the small IMPs, but it was not found in the human spermatozoa, whose IMP- density was low. From these observations, following conditions may be needed for the IMP-pattern formation; (1) the presence of similarly sized numerous IMPs, (2) the movement of the IMPs, (3) the presence of IMP gathering factors such as a barrier for their movement.Increase of membrane sterol density in the hamster spermatozoa during their epididymal passage The density of filipin-sterol complexes (FSCs),was 390/mum^2 in the PAPM of the hamster spermatozoa taken from the initial segment of the epididymis and it became to about 500/mum^2. when the spermatozoa passed the caput flexure. The increase of FSC density may contribute to stabilize the membrane.Changes of the sperm plasma membrane during in vitro capacitation in the golden hamster Spermatozoa were incubated in Tyrode solution, and they were examined by freeze fracture. At two hours after incubation, FSC density in the PAPM reduced by 70-80%. At three hours after incubation, when the spermatozoa were capacitated, the majority of small IMPs had disappeared, and wide IMP free- area were formed in the PAPM.Production of monoclonal antibody Monoclonal antibodies were produced against extracted proteins from the isolated spermatogenic cells in mice. One of such antibody, 13D3, recognized hsc7l, which belongs to 7OkDa heat shock protein family,. Indirect immunofluorescence showed that hsc7l was present throughout the cytoplasm in the young spermatogenic cells, but localized in the middle piece of the spermatozoa.