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. 2000 Oct 30;151(3):507–518. doi: 10.1083/jcb.151.3.507

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© 2000 The Rockefeller University Press

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).

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Characterization of in vitro–generated COPI-coated vesicles. Transport vesicles were generated from either CHO (left) or rat liver (right) Golgi membranes in the presence of bovine brain cytosol, ATP-regenerating system, and GTPγS. COPI vesicles were purified on a continuous sucrose gradient. The gradients were fractionated from the bottom into 18 fractions. Aliquots of fractions 3–16 of the isopycnic gradients were chloroform/methanol-precipitated, and analyzed by running on a 13% SDS-PAGE and Western blotting, using antibodies against β-COP and p23 (top). COPI vesicles are recovered in fractions 6–8, corresponding to an average sucrose concentration of 41% (wt/wt). Below are electron micrographs of COPI-coated vesicles present in fractions 6–8. Bars, 90 nm.