Closely spaced external standard

A universal method of achieving 5 ppm mass accuracy over the entire MALDI plate in axial matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Eugene Moskovets, Hsuan Shen Chen, Anna Pashkova, Tomas Rejtar, Viktor Andreev, Barry L. Karger

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

Close deposition of the sample and external standard was used in axial matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) to achieve mass accuracy equivalent to that obtained with an internal standard across the entire MALDI plate. In this work, the sample and external standard were deposited by continuous deposition in separate traces, each approximately 200 μm wide. The dependence of the mass accuracy on the distance between the sample and standard traces was determined across a MALDI target plate with dimensions of 57.5 mm × 57.0 mm by varying the gap between the traces from 100 μm to 4 mm. During acquisition, two adjacent traces were alternately irradiated with a 200-Hz laser, such that the peaks in the resulting mass spectra combined the sample and external standard. Ion suppression was not observed even when the peptide concentrations in the two traces differed by more than two orders of magnitude. The five peaks from the external standard trace were used in a four-term mass calibration of the masses of the sample trace. The average accuracy across the whole plate with this method was 5 ppm when peaks of the sample trace had signal-to-noise ratios of at least 30 and the gap between the traces was approximately 100 μm. This approach was applied to determining peptide masses of a reversed-phase liquid chromatographic (LC) separation of a tryptic digest of β-galactosidase deposited as a long serpentine trace across the MALDI plate, with accuracy comparable to that obtainable using internal calibration. In addition, the eluent from reversed-phase LC separation of a strong cation-exchange fraction containing tryptic peptides from a yeast lysate along with the closely placed external standard was deposited on the MALDI plate. The data obtained in the MS and MS/MS modes on a MALDI-TOF/TOF mass spectrometer were combined and used in database searching with MASCOT. Since the significant score is a function of mass accuracy in the MS mode, database searching with high mass accuracy reduced the number of false positives and also added peptides which otherwise would have been eliminated at lower mass accuracy (false negatives).

Original languageEnglish
Pages (from-to)2177-2187
Number of pages11
JournalRapid Communications in Mass Spectrometry
Volume17
Issue number19
StatePublished - Oct 13 2003

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Ionization
Mass spectrometry
Desorption
Lasers
Peptides
Galactosidases
Calibration
Liquids
Mass spectrometers
Yeast
Cations
Signal to noise ratio
Ion exchange
Ions

ASJC Scopus subject areas

  • Analytical Chemistry
  • Spectroscopy

Cite this

Closely spaced external standard : A universal method of achieving 5 ppm mass accuracy over the entire MALDI plate in axial matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. / Moskovets, Eugene; Chen, Hsuan Shen; Pashkova, Anna; Rejtar, Tomas; Andreev, Viktor; Karger, Barry L.

In: Rapid Communications in Mass Spectrometry, Vol. 17, No. 19, 13.10.2003, p. 2177-2187.

Research output: Contribution to journalArticle

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