Short GeLC-SWATH: a fast and reliable quantitative approach for proteomic screenings

Proteomics. 2015 Feb;15(4):757-62. doi: 10.1002/pmic.201400221. Epub 2015 Jan 14.

Abstract

The quantification of large proteomes across multiple samples has become the major focus of proteomics. In addition to the advantages of in-gel digestion, the extensive time and sample handling required have precluded the use of this type of method for large quantitative screens. Therefore, an adaptation of the in-gel digestion method, termed short-GeLC, is proposed as a faster and more reproducible sample preparation method for quantitative approaches. The proposed methodology was compared with two well-established procedures for sample preparation, GeLC-MS and the classic liquid digestion followed by LC-MS, using a membrane protein-enriched sample. The results show that the short-GeLC approach substantially reduces the amount of sample handling and the overall time required for analysis compared with the gel-based methods without compromising the overall results at the protein identification level. Furthermore, the short-GeLC approach in combination with the SWATH acquisition method leads to the best quantitative results: more proteins were quantified, and the reproducibility was improved. Finally, this method performed well even on challenging samples enriched in membrane proteins.

Keywords: In-gel digestion; Membrane proteins; Proteomic screenings; Quantification; SWATH; Technology.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cerebral Cortex / chemistry
  • Chromatography, Liquid / methods*
  • Membrane Proteins / analysis
  • Membrane Proteins / chemistry
  • Membrane Proteins / metabolism
  • Peptide Fragments / analysis*
  • Peptide Fragments / chemistry
  • Peptide Fragments / metabolism
  • Proteome / analysis*
  • Proteome / chemistry
  • Proteome / metabolism
  • Proteomics / methods*
  • Rats
  • Reproducibility of Results

Substances

  • Membrane Proteins
  • Peptide Fragments
  • Proteome