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High‐resolution density gradient electrophoresis of subcellular organelles and proteins under nondenaturing conditions
Author(s) -
Tulp Abraham,
FernandezBorja Mar,
Verwoerd Desirée,
Neefjes Jacques
Publication year - 1998
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150190812
Subject(s) - vesicle , isoelectric focusing , chromatography , electrophoresis , endosome , density gradient , free flow electrophoresis , isoelectric point , chemistry , resolution (logic) , organelle , transferrin , endocytic cycle , gel electrophoresis , biochemistry , membrane , polyacrylamide gel electrophoresis , gel electrophoresis of proteins , enzyme , endocytosis , physics , quantum mechanics , artificial intelligence , computer science , intracellular , cell
We have developed a density gradient electrophoresis device (DGE) and used it for the preparative separation of various endocytic organelles that are hard to separate by other means. Our separation by DGE of late endosomal vesicles, recycling vesicles, early endosomes and plasma membranes is unmatched. Using the same DGE device, we performed preparative high‐resolution rate zonal separation of proteins using amphoteric buffers as originally described by Bier ( Electrophoresis 1993, 14 , 1011–1018). Isoforms of bovine β‐lactoglobulin, human apo‐transferrin, and bovine erythrocyte carbonic anhydrase that have isoelectric points within 0.8 pH units were readily separated even in the absence of nonionic detergents. The DGE apparatus is inexpensive and has unique separation abilities for vesicles and proteins.