Isolation of plasma membranes from Drosophila embryos
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Cell surface components probably play an important role in early embryonic development. However, hardly any information is available on the structure or regulation of expression of the corresponding genes. As a first step in approaching this issue, we devised a procedure to obtain enriched plasma membranes from embryonic Drosophila cells. Membranes are fractionated according to two independent physical parameters: size, using velocity gradient centrifugation and density, using isopicnic gradient centrifugation. The final membrane fraction is enriched by 6 to 8 fold with respect to the plasma membrane enzyme marker Na+/K+ ATPase and substantially depleted of the mitochondrial enzyme marker cytochrome C oxidase. Two-dimensional polyacrylamide gel electrophoresis of the purified membranes reveals enrichment for specific proteins and electron microscopy reveals membrane vesicles in abundance. The enriched fraction should be suitable for the preparation of antibody probes that recognize cell surface components.
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