NEVILLE D M
J Biophys Biochem Cytol. 1960 Oct;8(2):413-22. doi: 10.1083/jcb.8.2.413.
A procedure is described for isolating cell membranes from rat liver homogenates. 20 gm. of rat liver was homogenized in a Dounce homogenizer in ice cold water buffered to pH 7.5 with NaHCO(3), rupturing all of the cells and most nuclei. The diluted homogenate was filtered through cheesecloth to remove precipitated nucleoprotein and centrifuged at 1500 g, 10 minutes, to sediment a crude membrane fraction. The membrane containing sediment was recentrifuged 3 times in conical tubes (1220 g, 10 minutes), the top layer of the 2-layered sediment being retained. Flotation in a sucrose solution d = 1.22 freed the preparation from contaminating cell fragments and nuclear membranes not previously disintegrated. The floating material approximately 0.4 ml. was quite homogeneous and consisted of thin amorphous membranes. Electron micrographs revealed numerous double profiles similar in shape and dimensions to apposed liver cell membranes in intact tissue.
本文描述了一种从大鼠肝脏匀浆中分离细胞膜的方法。取20克大鼠肝脏，在冰水中用Dounce匀浆器匀浆，用NaHCO₃将其缓冲至pH 7.5，使所有细胞和大多数细胞核破裂。将稀释后的匀浆通过粗棉布过滤以去除沉淀的核蛋白，然后在1500g条件下离心10分钟，使粗膜部分沉淀。将含有膜的沉淀物在锥形管中再离心3次（1220g，10分钟），保留两层沉淀物的顶层。在密度为1.22的蔗糖溶液中浮选，可使制剂中不含先前未分解的污染细胞碎片和核膜。浮选得到的约0.4毫升物质相当均匀，由薄的无定形膜组成。电子显微镜照片显示，许多双轮廓在形状和尺寸上与完整组织中相邻的肝细胞膜相似。
