Proteomic analysis of detergent-resistant membrane rafts

Josip Blonder, Martha L. Hale, David A. Lucas, Carl F. Schaefer, Li Rong Yu, Thomas P. Conrads, Haleem J. Issaq, Bradley G. Stiles, Timothy D. Veenstra*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

76 Scopus citations


A combined, detergent- and organic solvent-based proteomic method for the analysis of detergent-resistant membrane rafts (DRMR) is described. These specialized domains of the plasma membrane contain a distinctive and dynamic protein and/or lipid complement, which can be isolated from most mammalian cells. Lipid rafts are predominantly involved in signal transduction and adapted to mediate and produce different cellular responses. To facilitate a better understanding of their biology and role, DRMR were isolated from Vero cells as a Triton X-100 insoluble fraction. After detergent removal, sonication in 60% buffered methanol was used to extract, solubilize and tryptically digest the resulting protein complement. The peptide digestate was analyzed by microcapillary reversed-phase liquid chromatography-tandem mass spectrometry. Gas-phase fractionation in the mass-to-charge range was employed to broaden the selection of precursor ions and increase the number of identifications in an effort to detect less abundant proteins. A total of 380 proteins were identified including all known lipid raft markers. A total of 91 (24%) proteins were classified as integral α-helical membrane proteins, of which 51 (56%) were predicted to have multiple transmembrane domains.

Original languageEnglish
Pages (from-to)1307-1318
Number of pages12
Issue number9
StatePublished - May 2004
Externally publishedYes


  • Detergent-resistant membrane rafts
  • Gas-phase fractionation
  • Membrane proteomics
  • Reversed-phase liquid chromatography
  • Vero cells


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