Eriksson L C
Biochim Biophys Acta. 1978 Mar 21;508(1):155-64. doi: 10.1016/0005-2736(78)90197-9.
A modified procedure for preparing the microsomal fraction from rat liver was developed with the aim of increasing the recovery without increasing the degree of contamination. 87% of the membranes of the microsomal fraction isolated from the first mitochondrial (10 000 X g) supernatant originates from the endoplasmic reticulum, representing a 35% yield. By gentle resuspension of the 10 000 X g pellet followed by differential centrifugation a second crop of microsomes can be prepared which, together with the first crop, gives a 55% total recovery of microsomal markers. 87% of the protein in this second crop also originates from the endoplasmic reticulum and this fraction has properties similar to those of the first crop. Contaminating membranes include Golgi membranes (0.6% of the total protein), mitochondria (2.5%), lysosomes (5%) and plasma membranes (5%). Collecting further crops increases the contamination. Subfractionation studies revealed almost identical distributions of ribosome-rich, ribosome-poor and smooth membranes in the two crops of microsomal fractions. The results obtained after treatment of the animals with phenobarbital or methylcholantrene were similar to those obtained with control animals; but in the case of methylcholantrene treatment the second crop represents a larger portion of the total membranes of the endoplasmic reticulum.
为了在不增加污染程度的情况下提高回收率，开发了一种改良的从大鼠肝脏制备微粒体组分的方法。从第一次线粒体（10000×g）上清液中分离出的微粒体组分的87%的膜来自内质网，产率为35%。通过轻柔重悬10000×g沉淀，然后进行差速离心，可以制备第二批微粒体，将其与第一批微粒体一起，微粒体标志物的总回收率达到55%。第二批微粒体中87%的蛋白质也来自内质网，且该组分的性质与第一批相似。污染膜包括高尔基体膜（占总蛋白的0.6%）、线粒体（2.5%）、溶酶体（5%）和质膜（5%）。收集更多批次会增加污染。亚分级研究表明，两批微粒体组分中富含核糖体、核糖体较少和光滑膜的分布几乎相同。用苯巴比妥或甲基胆蒽处理动物后得到的结果与对照动物相似；但在甲基胆蒽处理的情况下，第二批微粒体占内质网总膜的比例更大。
