Haaparanta T, Gustafsson J A, Glaumann H
Arch Biochem Biophys. 1983 Jun;223(2):458-67. doi: 10.1016/0003-9861(83)90610-0.
A procedure is presented for the isolation of lysosomes, mitochondria, and microsomes from the rat ventral prostate with relatively good yield. Homogenization was performed with a Polytron homogenizer or in combination with the Potter-Elvehjem device. Reasonably pure mitochondria and lysosomes could only be obtained using a Metrizamide gradient, whereas it was possible to prepare pure microsomal fractions by differential centrifugation in sucrose. The purity of the lysosomes and mitochondria was 90 and 85%, respectively, as judged by the presence of different marker enzymes. These findings were confirmed by ultrastructural analyses. Electron micrographs of the isolated lysosomes showed intact lysosomes surrounded by a single membrane. The lysosomes contained intramatrical vesicles with lipid-like material. Vesicles derived from the endoplasmic reticulum in the microsomal fraction ranged from 70 to 90% depending on the centrifugal force used to sediment the mitochondrial fraction. Electron micrographs of the microsomal fraction showed that about 40% of the vesicles were inverted and turned "inside-out", i.e., having their ribosomes attached to the inside of the vesicles. By fractionation of ethylenediaminetetraacetate treated microsomes on a sucrose gradient a partially purified fraction was isolated which consisted of 65% of inverted microsomes.
本文介绍了一种从大鼠腹侧前列腺中分离溶酶体、线粒体和微粒体的方法，其产量相对较高。使用Polytron匀浆器或与Potter-Elvehjem装置联合进行匀浆。只有使用Metrizamide梯度才能获得纯度较高的线粒体和溶酶体，而通过在蔗糖中进行差速离心可以制备纯的微粒体部分。根据不同标记酶的存在情况判断，溶酶体和线粒体的纯度分别为90%和85%。这些发现通过超微结构分析得到了证实。分离出的溶酶体的电子显微镜照片显示，完整的溶酶体被单层膜包围。溶酶体含有带有类脂物质的基质内小泡。微粒体部分中源自内质网的小泡占70%至90%，这取决于用于沉淀线粒体部分的离心力。微粒体部分的电子显微镜照片显示，约40%的小泡是倒置的，即“内外翻转”，也就是说它们的核糖体附着在小泡内部。通过在蔗糖梯度上对经乙二胺四乙酸处理的微粒体进行分级分离，分离出了一个部分纯化的部分，该部分由65%的倒置微粒体组成。
