Casale T B, Friedman M, Parada N, Plekes J, Kaliner M
J Membr Biol. 1984;79(1):33-9. doi: 10.1007/BF01868524.
A simple and rapid method of isolating plasma membranes from human peripheral lung tissue is described. The method involves homogenization of tissue in 0.25 M sucrose-buffered medium followed by differential and sucrose density gradient centrifugation. Enzymatic and morphological characterization of the plasma membrane fraction revealed minimal contamination by nonplasma membrane fragments. The isolated plasma membranes showed an 18-fold purification of 5'-nucleotidase activity compared to the original homogenate. Electron-microscopic studies of the plasma membrane fraction revealed the presence of small membrane vesicles having a trilaminar membrane structure. To further examine the purity of the plasma membrane preparation, the binding of the H1 receptor antagonist, 3H pyrilamine, to the plasma membrane-enriched fraction was compared to the binding to crude membrane preparations. Both the plasma membrane-enriched fraction and the crude membrane preparation had similar Kd's for the histamine antagonist, but the plasma membrane-enriched fraction had a threefold greater binding capacity, reflecting the relative enrichment of plasma membranes of the preparation. Thus, a method has been developed for the isolation of plasma membranes from human peripheral lung which should provide material for a variety of biochemical and pharmacological studies.
本文描述了一种从人外周肺组织中分离质膜的简单快速方法。该方法包括在0.25M蔗糖缓冲培养基中对组织进行匀浆，然后进行差速离心和蔗糖密度梯度离心。对质膜部分的酶学和形态学表征显示，非质膜片段的污染极少。与原始匀浆相比，分离得到的质膜显示5'-核苷酸酶活性纯化了18倍。对质膜部分的电子显微镜研究显示存在具有三层膜结构的小膜泡。为了进一步检查质膜制剂的纯度，将H1受体拮抗剂3H吡苄明与富含质膜的部分的结合与与粗膜制剂的结合进行了比较。富含质膜的部分和粗膜制剂对组胺拮抗剂的Kd相似，但富含质膜的部分的结合能力大三倍，反映了制剂中质膜的相对富集。因此，已经开发出一种从人外周肺中分离质膜的方法，该方法应为各种生化和药理学研究提供材料。
