Improve method for isolation of rat liver plasma membrane |
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Authors: | Alfred E. Brown M.P. Lok John Elovson |
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Affiliation: | Department of Biology, University of California, San Diego, La Jolla, California 92037 U.S.A. |
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Abstract: | An improved method for the isolation of plasma membrane from rat liver is presented.Gentle homogenization of perfused livers in buffered isotonic KCI, followed by direct flotation of a low-speed nuclear pellet through a discontinuous sucrose density gradient results in a 32% yield, and 25-fold enrichment for the plasma membrane marker, phosphodiesterase I, in a crude plasma membrane fraction. This fraction contains less than 1% of the mitochondria, and endoplasmic reticulum present in the original homogenate, but is more heavily contaminated with lysosomes and Golgi membrane.Vigorous mechanical disruption of this material, followed by a second discontinuous sucrose density gradient, gives a light plasma membrane fraction with an 80-fold purification and 20% yield of phosphodiesterase I over the original homogete (with further reduction of contaminants). |
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Keywords: | HEPES MOPES morpholinopropane-sulfonic acid |
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