Purification and partial characterization of plasma membranes from bovine spermatozoa.
Purification and partial characterization of plasma membranes from bovine spermatozoa.
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牛精子质膜的纯化和部分表征。
DOI：
10.1095/biolreprod29.4.987
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发表时间：
1983
影响因子：
3.6
通讯作者：
Garbers,DL
中科院分区：
文献类型：
--
作者：
Noland,TD;Olson,GE;Garbers,DL
对牛附睾精子进行氮气空化（600 psi，10 min）以去除质膜。通过电子显微镜检查的空化细胞显示，质膜被优先删除的顶体周围和鞭毛区域。核膜、线粒体膜和顶体膜保持完整并附着在精子上，但细胞质液滴经常被破坏，其内部的膜结合囊泡被释放。较低的压力（小于200 psi）在去除顶体周围质膜方面相对无效，而中等压力（400 psi）从约70%的精子中去除了该膜。没有明显的选择性去除的顶体和鞭毛质膜观察作为空化pressure.The空化细胞的功能分离的质膜微分，然后通过线性蔗糖密度梯度离心。两个不同的膜群体，解决了蔗糖梯度，并指定带I和带II。带I只包含从质膜产生的球形囊泡。完整细胞的表面标记证实了质膜是带I的起源。包括带II的较高密度的膜是异质的，由球形和扁平囊泡组成。当纯化的细胞质液滴空化和离心的蔗糖梯度上只有带IIwerceived.These研究表明，氮空化牛附睾精子可以导致显着的污染质膜组分的细胞质液滴膜，除非适当的差速离心分离膜组分。
Bovine epididymal spermatozoa were subjected to nitrogen cavitation (600 psi for 10 min) to remove plasma membrane. Examination of the cavitated cells by electron microscopy revealed that the plasma membrane was preferentially removed from the periacrosomal and flagellar regions. Nuclear, mitochondrial and acrosomal membranes remained intact and attached to the spermatozoa, but the cytoplasmic droplets were frequently disrupted and their internal membrane-bound vesicles were released. Lower pressures (less than 200 psi) were relatively ineffective in removing the periacrosomal plasma membrane, while an intermediate pressure (400 psi) removed this membrane from about 70% of the spermatozoa. No apparent selectivity for removal of the periacrosomal and flagellar plasma membrane was observed as a function of cavitation pressure.The cavitated cells were separated from the plasma membranes by differential followed by linear sucrose density gradient centrifugation. Two distinct membrane populations were resolved on sucrose gradients and were designated Band I and Band II. Band I contained only spherical vesicles which arose from the plasma membrane. Surface labeling of intact cells confirmed the plasma membrane as the origin of Band I. The membranes of higher density comprising Band II were heterogeneous consisting of both spherical and flattened vesicles. When purified cytoplasmic droplets were cavitated and centrifuged on the sucrose gradient only Band II was obtained.These studies indicate that nitrogen cavitation of bovine epididymal spermatozoa can result in significant contamination of plasma membrane fractions by cytoplasmic droplet membranes unless appropriate differential centrifugation is used to separate the membrane fractions.