Hazlewood C F, Kellermayer M
Department of Physiology and Molecular Biophysics, Baylor College of Medicine, Houston, TX 77030.
Scanning Microsc. 1988 Mar;2(1):267-73.
Nonionic detergents, Triton X-100 and Brij 58, removed, within 2-5 minutes, lipid membrane of suspended thymus lymphocytes and monolayer H-50 cells grown in culture. Studies of hydration, ionic asymmetry, and ionic and protein release kinetics were conducted on these membraneless cellular preparations. The hydration of nuclei isolated by Triton X-100 procedures appears to be influenced strongly by the monovalent ionic concentration of the buffer bathing the organelles. The putative monovalent ionic concentration of the cellular aqueous phase (i.e., 150 meq/L) caused nuclei to swell and coalesce. Monovalent ionic concentrations of 30 meq/L or less caused minimal changes in volume and in morphology. Triton X-100 treatment led to rapid mobilization and solubilization of membrane and cytoplasmic lipids and proteins, and the cellular potassium was reduced to very low levels. Brij 58 treatment of the lymphocytes for 5 minutes led to loss of membrane structure. Potassium, however, was retained at significant levels for over 10 minutes. Potassium and protein release kinetic studies in the H-50 monolayer cells following Brij treatment revealed that potassium and the detergent mobilized proteins may be co-compartmentalized and that 10 minutes or more are required before their release is completed. These results support the view that most of the potassium and "diffusible" proteins are not fully dissolved in the cellular water. Furthermore, the integrity of the membranes does not appear to be essential for the retention of the ions and the proteins.
非离子型去污剂Triton X - 100和Brij 58在2 - 5分钟内去除了悬浮的胸腺淋巴细胞和培养的单层H - 50细胞的脂质膜。对这些无膜细胞制剂进行了水合作用、离子不对称性以及离子和蛋白质释放动力学的研究。通过Triton X - 100程序分离的细胞核的水合作用似乎受到洗涤细胞器的缓冲液单价离子浓度的强烈影响。细胞水相假定的单价离子浓度(即150 meq/L)会导致细胞核肿胀并聚集。30 meq/L或更低的单价离子浓度引起的体积和形态变化最小。Triton X - 100处理导致膜和细胞质脂质及蛋白质迅速动员和溶解,细胞内钾含量降至极低水平。用Brij 58处理淋巴细胞5分钟导致膜结构丧失。然而,钾在10多分钟内仍保持在显著水平。对Brij处理后的H - 50单层细胞进行的钾和蛋白质释放动力学研究表明,钾和去污剂动员的蛋白质可能共分隔,并且在它们完全释放之前需要10分钟或更长时间。这些结果支持这样一种观点,即大多数钾和“可扩散”蛋白质并未完全溶解在细胞水中。此外,膜的完整性对于离子和蛋白质的保留似乎并非必不可少。