MATURATION OF HUMAN WHITE MATTER MYELIN . FRACTIONATION OF MYELIN MEMBRANE PROTEINS
MATURATION OF HUMAN WHITE MATTER MYELIN . FRACTIONATION OF MYELIN MEMBRANE PROTEINS
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DOI：
10.1021/bi00852a042
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发表时间：
1968-01-01
期刊：
影响因子：
2.9
通讯作者：
TAVASTSTJERNA, MG
中科院分区：
文献类型：
--
作者：
ENG, LF;CHAO, FC;TAVASTSTJERNA, MG
材料与方法尸检取脑组织。个体的年龄从10周到92岁不等。除2例MS患者外，其余患者均无中枢神经系统病变，且经临床观察多年，其脑组织均呈典型病变。仅使用大体上无斑块的白色物质进行研究。牛脑和坐骨神经从当地屠宰场新鲜获得。Wistar大鼠（16日龄）用作脑和脊髓的来源。Triton X-100（烷基苯氧基聚乙氧基乙醇）由Rohm and哈斯，费城，宾夕法尼亚州慷慨提供。马心细胞色素C购自Calbiochem，洛杉矶，加利福尼亚州。髓鞘的制备。用Laatsch等人的方法从人和牛的白色物质中制备髓鞘。(1962; Gerstl等人，1967）并通过在连续蔗糖梯度中离心进一步纯化（Autilio等，1964; Eng and Noble，1968）.中性盐和酸提取髓鞘。首先在玻璃组织匀浆器中使用马达驱动的特氟隆研杵将髓磷脂与0.11 mM乙酸铵溶液（pH 6.9）冷均质化30分钟。在以平均105，000 g离心1小时后，获得髓磷脂沉淀和澄清的上清液。使用该程序，然后用0.5 M乙酸铵溶液（pH 6.9）提取髓磷脂30 min。通过反复冻干（3 - 5次）除去上清液中的水和乙酸铵。在最终冻干后，4455
Materials and MethodsHuman brain tissue was obtained at autopsy. The age of the individuals ranged from 10 weeks to 92 years. Their history suggested, andthe gross and histological examination confirmed, the absence of lesions in the CNS in all except two cases with MS. The MS cases had been under clinical observation for many years, and their brains showed typical lesions. Only white matter grossly free of plaques was utilized for study. Bovine brains and sciatic nerve were obtained fresh from a local slaughterhouse. Wistar rats (16-day old) were used as the source for brain and spinal cord. Triton X-100 (alkylphenoxypolyethoxyethanol) was generously supplied by Rohm and Haas, Philadel-phia, Pa. Horse heart cytochrome C was purchased from Calbiochem, Los Angeles, Calif. Preparation of Myelin. Myelin was prepared from human and bovine white matter by the procedure of Laatsch et al.(1962; Gerstl et al., 1967) and further purified by centrifugation in a continuous sucrose gradient (Autilio et al., 1964; Eng and Noble, 1968). Neutral Salt and Acid Extraction of Myelin. Myelin was first homogenized in the cold with 0.11 m ammo-nium acetate solution (pH 6.9) for 30 min in a glass tissue homogenizer using a motor-driven Teflon pestle. On centrifugation at an average of 105,000 g for 1 hr, a myelin pellet and a clear supernatant were obtained. Using this procedure, the myelin was then extracted with 0.5 m ammonium acetate solution (pH 6.9) for 30 min. Water and ammonium acetate in the supernatant were removed by repeated lyophilization (three to five times). Following the final lyophilization, the 4455