The isolation of detergent-resistant lipid rafts for two-dimensional electrophoresis.

Ki Bum Kim, Jae Seon Lee, Young-Gyu Ko

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

Because lipid rafts are plasma membrane platforms mediating various cellular events such as in signal transduction, immunological response, pathogen invasion, and neurodegenerative diseases, protein identification in the rafts could provide important information to study their function. Here, we present an optimized method to isolate detergent-resistant lipid rafts that are subsequently analyzed by two-dimensional electrophoresis (2-DE). Lipid rafts were isolated based on their two distinct biochemical properties such as Triton X-100 insolubility and low density. To solubilize completely the proteins embedded in lipid rafts, sample lysis buffer (9 M urea, 2 M thiourea, 100 mM DTT, 2% CHAPS (w/v), 60 mM n-octylbeta-D-glucopyranoside, 2% IPG buffer) was applied to the isolated rafts. This method was found to be the most suitable choice for obtaining 2-DE profile of lipid raft proteome from various cells and tissues. We expect that this method could provide the way to dissect the function of raft-associated proteins and to gain a comprehensive insight upon various cellular events mediated through lipid rafts, the specialized domains in cell surface.

Original languageEnglish
Pages (from-to)413-422
Number of pages10
JournalMethods in molecular biology (Clifton, N.J.)
Volume424
Publication statusPublished - 2008 May 27

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Detergents
Electrophoresis
Lipids
Buffers
Thiourea
Proteins
Octoxynol
Proteome
Neurodegenerative Diseases
Urea
Signal Transduction
Cell Membrane

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Cite this

The isolation of detergent-resistant lipid rafts for two-dimensional electrophoresis. / Kim, Ki Bum; Lee, Jae Seon; Ko, Young-Gyu.

In: Methods in molecular biology (Clifton, N.J.), Vol. 424, 27.05.2008, p. 413-422.

Research output: Contribution to journalArticle

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