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

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.