Fractionation of subcellular membrane vesicles of epithelial and nonepithelial cells by OptiPrep density gradient ultracentrifugation.

Research output: Contribution to journalArticle

Abstract

Density gradient ultracentrifugation (DGUC) is widely used for physical isolation (enrichment rather than purification) of subcellular membrane vesicles. It has been a valuable tool to study specific subcellular localization and dynamic trafficking of proteins. While sucrose has been the main component of density gradients, a few years ago synthetic OptiPrep (iodixanol) began being used for separation of organelles because of its iso-osmotic property. Here, we describe a detailed protocol for density gradient fractionation of various mammalian subcellular vesicles, including endoplasmic reticulum (ER), Golgi apparatus, endosomes, and lipid rafts, as well as apical and basolateral membranes of polarized epithelial cells.

Original languageEnglish (US)
Pages (from-to)97-110
Number of pages14
JournalMethods in molecular biology (Clifton, N.J.)
Volume440
StatePublished - 2008

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Ultracentrifugation
Cell Count
Epithelial Cells
Membranes
Endosomes
Golgi Apparatus
Protein Transport
Endoplasmic Reticulum
Organelles
Sucrose
Lipids
iodixanol

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

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abstract = "Density gradient ultracentrifugation (DGUC) is widely used for physical isolation (enrichment rather than purification) of subcellular membrane vesicles. It has been a valuable tool to study specific subcellular localization and dynamic trafficking of proteins. While sucrose has been the main component of density gradients, a few years ago synthetic OptiPrep (iodixanol) began being used for separation of organelles because of its iso-osmotic property. Here, we describe a detailed protocol for density gradient fractionation of various mammalian subcellular vesicles, including endoplasmic reticulum (ER), Golgi apparatus, endosomes, and lipid rafts, as well as apical and basolateral membranes of polarized epithelial cells.",
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