2-D differential membrane proteome analysis of scarce protein samples

Proteomics. 2006 Aug;6(16):4506-13. doi: 10.1002/pmic.200600169.

Abstract

Proteome studies with small sample amounts are difficult to perform, especially when membrane proteins are the focus of interest. In our study a new method for the analysis of scarce membrane protein samples combining large gel 2-D-CTAB/SDS-PAGE with fluorescence dye saturation labelling (satDIGE) was developed, allowing a highly sensitive differential analysis of different cell states. After Triton X-114 phase partitioning, enriched membrane protein samples of T cells were labelled at cysteine residues using fluorescence dyes and separated by large gel 2D-CTAB/SDS-PAGE. For a differential analysis 3 mug protein was found to be sufficient to detect proteins in a widespread well-separated diagonal spot pattern.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cells, Cultured
  • Electrophoresis, Gel, Two-Dimensional
  • Fluorescent Dyes / chemistry
  • Humans
  • Membrane Proteins / analysis*
  • Membrane Proteins / isolation & purification
  • Mice
  • Molecular Sequence Data
  • Proteome*
  • Spectrometry, Mass, Electrospray Ionization

Substances

  • Fluorescent Dyes
  • Membrane Proteins
  • Proteome