Leibniz Institute for Zoo and Wildlife Research, Berlin, Germany.
Lipids Health Dis. 2009 Nov 11;8:49. doi: 10.1186/1476-511X-8-49.
Detergents are often used to isolate proteins, lipids as well as "detergent-resistant membrane domains" (DRMs) from cells. Different detergents affect different membrane structures according to their physico-chemical properties. However, the effects of different detergents on membrane lysis of boar spermatozoa and the lipid composition of DRMs prepared from the affected sperm membranes have not been investigated so far.
Spermatozoa were treated with the selected detergents Pluronic F-127, sodium cholate, CHAPS, Tween 20, Triton X-100 and Brij 96V. Different patterns of membrane disintegration were observed by light and electron microscopy. In accordance with microscopic data, different amounts of lipids and proteins were released from the cells by the different detergents. The biochemical methods to assay the phosphorus and cholesterol contents as well as 31P NMR to determine the phospholipids were not influenced by the presence of detergents since comparable amounts of lipids were detected in the organic extracts from whole cell suspensions after exposure to each detergent. However, matrix-assisted laser desorption and ionization time-of-flight mass spectrometry applied to identify phospholipids was essentially disturbed by the presence of detergents which exerted particular suppression effects on signal intensities. After separation of the membrane fractions released by detergents on a sucrose gradient only Triton X-100 and sodium cholate produced sharp turbid DRM bands. Only membrane solubilisation by Triton X-100 leads to an enrichment of cholesterol, sphingomyelin, phosphatidylinositol and phosphatidylethanolamine in a visible DRM band accompanied by a selective accumulation of proteins.
The boar sperm membranes are solubilised to a different extent by the used detergents. Particularly, the very unique DRMs isolated after Triton X-100 exposure are interesting candidates for further studies regarding the architecture of sperm.
清洁剂通常用于从细胞中分离蛋白质、脂质以及“抗清洁剂膜域”（DRM）。不同的清洁剂根据其物理化学性质影响不同的膜结构。然而，迄今为止，尚未研究不同清洁剂对猪精子膜的溶血作用以及受影响的精子膜制备的 DRM 的脂质组成的影响。
选择 Pluronic F-127、胆酸钠、CHAPS、Tween 20、Triton X-100 和 Brij 96V 等清洁剂处理精子。通过光镜和电子显微镜观察到不同的膜崩解模式。根据显微镜数据，不同的清洁剂从细胞中释放出不同数量的脂质和蛋白质。由于在每种清洁剂暴露后，从全细胞悬浮液的有机提取物中检测到相当数量的脂质，因此用于测定磷和胆固醇含量的生化方法以及用于确定磷脂的 31P NMR 不受清洁剂的影响。然而，应用于鉴定磷脂的基质辅助激光解吸电离飞行时间质谱受到清洁剂的存在的严重干扰，这些清洁剂对信号强度产生特殊的抑制作用。在蔗糖梯度上分离清洁剂释放的膜部分后，只有 Triton X-100 和胆酸钠产生明显混浊的 DRM 带。只有用 Triton X-100 进行的膜溶解才会导致胆固醇、鞘磷脂、磷脂酰肌醇和磷脂酰乙醇胺在可见的 DRM 带中富集，同时伴有蛋白质的选择性积累。
所使用的清洁剂对猪精子膜的溶解程度不同。特别是在用 Triton X-100 暴露后分离出的非常独特的 DRM 是进一步研究精子结构的有趣候选物。
