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- W146759131 abstract "This chapter explains the procedures for the isolation of nuclei, mitochondria, and lysosomes from cultured mammalian cell lines. Cultured Chinese hamster ovary (CHO) cells are routinely disrupted by low-pressure nitrogen cavitation. The cell pellets concentrate at the bottom of the bottles. All centrifugations are timed from when the rotor reaches running speed. The cavitated cells are homogenized with four strokes of a motor-driven pestle of a Potter–Elvehjem homogenizer. The motor drive is set so that the Teflon pestle almost stops turning as it reaches the bottom of the homogenizer. For the subsequent isolation of cytoplasmic organelles, nuclei and unbroken cells are rapidly separated from cytoplasmic organelles by differential sedimentation at low centrifugal force. Intact organelle membranes prevent substrate access to lumenal enzymes. Hence, organelle-specific enzyme activity is generally measured in the presence of nonionic detergent; comparison of enzyme activity in the presence and absence of detergent provides a quick method to assess organelle intactness." @default.
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- W146759131 date "1990-01-01" @default.
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- W146759131 title "[16] Isolation of subcellular organelles" @default.
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- W146759131 doi "https://doi.org/10.1016/0076-6879(90)82018-w" @default.
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