title: Matrix-assisted laser desorption/ionization directed nano-electrospray ionization tandem mass spectrometric analysis for protein identification.
abstract: In those cases where the information obtained by peptide mass fingerprinting or matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-MS/MS) is not sufficient for unambiguous protein identification, nano-electrospray ionization (nano-ESI) and/or electrospray ionization tandem mass spectrometry (ESI-MS/MS) analysis must be performed. The sensitivity of nano-ESI/MS, however, is lower than that of MALDI-MS, especially at very low analyte concentrations and/or in the presence of contaminants, such as salt and detergents. Moreover, to perform ESI-MS/MS, the peptide masses of the precursor ions must be known. The approach described in this paper, MALDI-directed nano-ESI-MS/MS, makes use of information obtained from the more sensitive MALDI-MS experiments in order to direct subsequent nano-ESI-MS/MS experiments. Peptide molecular ions found in the MALDI-MS analysis are then selected, as their (+2) precursor ions, for nano-ESI-MS/MS sequencing, even though these ions cannot be detected in the ESI-MS spectra. This method, originally proposed by Tempst et al. (Anal. Chem. 2000, 72: 777-790), has been extended to provide better sensitivity and shorter analysis times; also, a comparison with liquid chromatography/tandem mass spectrometry (LC/MS/MS) has been performed. These experiments, performed using quadrupole time-of-flight instruments equipped with commercially available nano-ESI sources, have allowed the unambiguous identification of in-gel digested proteins at levels below their ESI-MS detection limits, even in the presence of salts and detergents.
authors.affiliation: Biomedical Research Centre, University of British Columbia, Vancouver, BC, V6T 1Z3, Canada.
authors.forename: Juergen
authors.initials: J
authors.lastname: Kast
authors.forename: Carol E
authors.initials: CE
authors.lastname: Parker
authors.forename: Koen
authors.initials: K
authors.lastname: van der Drift
authors.forename: J Michael
authors.initials: JM
authors.lastname: Dial
authors.forename: Sharon L
authors.initials: SL
authors.lastname: Milgram
authors.forename: Matthias
authors.initials: M
authors.lastname: Wilm
authors.forename: Michael
authors.initials: M
authors.lastname: Howell
authors.forename: Christoph H
authors.initials: CH
authors.lastname: Borchers
mesh_terms: D036103:Nanotechnology / Q000379:methods*; D011506:Proteins / Q000032:analysis*; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D011506:Proteins