Henkel R, Stalf T, Miska W
Andrologische Forschungsabteilung, Universität Giessen, Germany.
Biol Chem Hoppe Seyler. 1992 Aug;373(8):685-9. doi: 10.1515/bchm3.1992.373.2.685.
Flagella of human spermatozoa were separated from the sperm head by sonication at 25 kHz and subsequent density gradient centrifugation in Percoll. For isolation of the outer dense fibers (ODF), the flagellar membrane and fibrous sheath were dissolved under reducing conditions in the cationic detergent cetyltrimethylammonium bromide (CTAB) for 30, 60 and 90 min, respectively. The isolation steps were monitored by phase-contrast microscopy and electron microscopy. After SDS-PAGE and silver staining two protein bands at 55 and 67 kDa could be detected. An identification of these proteins as phosphoproteins, either with molybdate/methylgreen or rhodamine B, was not possible. The obtained results indicate that the ODF proteins might have more passive elastic than active function with respect to motility of spermatozoa.
通过25千赫兹的超声处理以及随后在Percoll中进行密度梯度离心，将人类精子的鞭毛与精子头部分离。为了分离外致密纤维（ODF），分别在还原条件下将鞭毛膜和纤维鞘溶解于阳离子去污剂十六烷基三甲基溴化铵（CTAB）中30分钟、60分钟和90分钟。通过相差显微镜和电子显微镜监测分离步骤。经过SDS-PAGE和银染后，可检测到55 kDa和67 kDa处的两条蛋白带。无法用钼酸盐/甲基绿或罗丹明B将这些蛋白鉴定为磷蛋白。所得结果表明，就精子的运动性而言，ODF蛋白可能具有更多的被动弹性而非主动功能。
