Studies on protein and nucleic acid metabolism in virus-infected mammalian cells. 3. Methods for the disruption of Krebs II mouse-ascites-tumour cells

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Abstract
Krebs II ascites-tumour cells can be ruptured by ultrasonics, by osmotic shock or by the high shear of a Dounce homogenizer. Cells ruptured in a Ca2+- and Mg2+ -free medium by a combination of osmotic shock and a low-shear homogenizer gave high yields of undamaged nuclei, mitochondria and microsomes. Nuclei prepared by the osmotic method and separated from contaminating whole cells by a novel procedure, were free from succinoxidase and had low RNA:DNA ratio. Microsomes prepared by osmotic-shock methods in the absence of Mg2+ ions were less active in incorporating radioactive valine than were microsomes prepared in the presence of Mg2+ ions by ultrasonics or by a high-shear homogenizer. The Krebs II ascites-tumour cell is unusually robust and cannot be ruptured by methods which are adequate for other types of cell.