Optimization of a MALDI TOF-TOF mass spectrometer for intact protein analysis



A MALDI TOF-TOF instrument was optimized and evaluated for intact protein analysis by tandem mass spectrometry. Ion source voltages and delay times were adjusted to affect an up to a 10-fold improvement in fragment ion yield compared to data obtained using default settings employed in peptide analysis. For large peptides (3–4.5 kDa), up to 90% of all possible b- and y-fragment ions were observed, which provides sufficient information for de novo sequencing and unambiguous protein identification. Product ion signals associated with preferential cleavages C-terminal to aspartic acid and glutamic acid residues and N-terminal to proline residues became dominant with increased protein molecular weight. Matrix effects were also evaluated and, among the eight matrices examined, α-cyano-4-hydroxycinnamic acid (CHCA) was found to produce the best intact protein tandem mass spectra for proteins up to 12 kDa. Optimized performance yielded detection limits of 50–125 fmol for proteins of 4 and 12 kDa, respectively. This improved performance has yielded an instrument with potential to be a useful tool in proteomic investigations via analysis of intact proteins.


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Copyright information

© American Society for Mass Spectrometry 2005

Authors and Affiliations

  1. 1.Department of Cell and Molecular PharmacologyMedical University of South CarolinaCharlestonUSA

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