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Digital Microfluidics for Automated Proteomic Processing
Published on: November 6, 2009
Assessing a novel microfluidic interface for shotgun proteome analyses
An Staes1, Evy Timmerman, Jozef Van Damme
1Department of Medical Protein Research, VIB, Ghent, Belgium.
Journal of Separation Science
|July 12, 2007
Summary
New HPLC-Chips enhance proteomic analysis sensitivity and protein identification by improving chromatography and peptide charging for mass spectrometry. This method offers greater proteome coverage with minimal user intervention.
Area of Science:
- Proteomics
- Analytical Chemistry
- Biotechnology
Background:
- Microfluidic interfaces coupled with mass spectrometry offer increased sensitivity and reduced manipulation for proteomics.
- Conventional nano-liquid chromatography (LC) interfaces have limitations in sample loading capacity.
Purpose of the Study:
- Evaluate a novel HPLC-Chip with larger enrichment columns for gel-free proteome studies.
- Compare the performance of the HPLC-Chip against a conventional nano-LC-MS/MS interface.
Main Methods:
- Analysis of tryptic digest of human T-cell proteome using strong cation exchange chromatography.
- Peptide fractionation and subsequent MS/MS analysis on an ion trap mass spectrometer.
- Comparison of HPLC-Chip and conventional nano-LC-MS/MS interfaces.
Main Results:
- The HPLC-Chip effectively handled complex peptide mixtures, identifying more peptides and proteins than the conventional interface.
- The HPLC-Chip demonstrated a preference for producing doubly charged tryptic peptides.
- MS/MS spectra from doubly charged ions were more readily identified by MASCOT.
Conclusions:
- The HPLC-Chip enhances proteome coverage in mass spectrometry-based proteomics.
- Improved chromatographic conditions and peptide charging profile contribute to increased identification rates.
- The HPLC-Chip represents a significant advancement for gel-free proteome studies.
