On-chip protein sample desalting and preparation for direct coupling with electrospray ionization mass spectrometry.
A membrane-based desalting step integrated in a MS microchip is presented: drugs, peptides and proteins are adsorbed on a hydrophobic poly(vinylidene difluoride) membrane, which allows the washing out of salts. The integration with microfluidics permits a controlled elution of analytes from the membrane and their direct mass spectrometric analysis by electrospray ionisation MS. The desalting process is demonstrated with picomole amounts of propanolol, insulin and cytochrome c. Moreover, this stop-and-go desalting process is tolerant to high concentrations of urea, and to the presence of reductants such as dithiothreitol. This particular feature allowed the chemical tagging of cysteines in beta-lactoglobulin A with iodoacetamide. Finally, the integration of chemical tagging, on-chip desalting and MS microchip paves the way for the development of high-throughput analytical procedure for structural proteomics.
Lion
Niels
N
Laboratoire d'Electrochimie Physique et Analytique, Institut de Chimie Moléculaire et Biologique, Ecole Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland.
Gellon
Jean-Olivier
JO
Jensen
Henrik
H
Girault
Hubert H
HH
D000327:Adsorption; D003545:Cysteine / Q000737:chemistry; D045304:Cytochromes c / Q000737:chemistry; D004229:Dithiothreitol; D007202:Indicators and Reagents; D007328:Insulin / Q000737:chemistry; D007460:Iodoacetamide; D007782:Lactoglobulins / Q000737:chemistry; D011433:Propranolol / Q000737:chemistry; D040081:Protein Array Analysis; D011506:Proteins / Q000737:chemistry*; D012492:Salts; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014508:Urea
D007202:Indicators and Reagents; D007328:Insulin; D007782:Lactoglobulins; D011506:Proteins; D012492:Salts; D014508:Urea; D045304:Cytochromes c; D011433:Propranolol; D003545:Cysteine; D004229:Dithiothreitol; D007460:Iodoacetamide