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A Calibration Method That Simplifies and Improves Accurate Determination of Peptide Molecular Masses by MALDI-TOF MS

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http://pubman.mpdl.mpg.de/cone/persons/resource/persons50168

Gobom,  Johan
Dept. of Vertebrate Genomics (Head: Hans Lehrach), Max Planck Institute for Molecular Genetics, Max Planck Society;

Egelhofer,  Volker
Max Planck Society;

Theiss,  Dorothea
Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons50409

Lehrach,  Hans
Dept. of Vertebrate Genomics (Head: Hans Lehrach), Max Planck Institute for Molecular Genetics, Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons50448

Nordhoff,  Eckhard
Dept. of Vertebrate Genomics (Head: Hans Lehrach), Max Planck Institute for Molecular Genetics, Max Planck Society;

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Citation

Gobom, J., Mueller, M., Egelhofer, V., Theiss, D., Lehrach, H., & Nordhoff, E. (2002). A Calibration Method That Simplifies and Improves Accurate Determination of Peptide Molecular Masses by MALDI-TOF MS. Analytical Chemistry, 74(15), 3915-3923.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0010-8BDE-7
Abstract
The use of delayed ion extraction in MALDI time-of-flight mass spectrometry distorts the linear relationship between m/z and the square of the ion flight time (t2) with the consequence that, if a mass accuracy of 10 ppm or better is to be obtained, the calibrant signals have to fall close to the analyte signals. If this is not possible, systematic errors arise. To eliminate these, a higher-order calibration function and thus several calibrant signals are required. For internal calibration, however, this approach is limited by signal suppression effects and the increasing chance of the calibrant signals overlapping with analyte signals. If instead the calibrants are prepared separately, this problem is replaced by an other; i.e., the ion flight times are dependent on the sample plate position. For this reason, even if the calibrants are placed close to the sample, the mass accuracy is not improved when a higher-order calibration function is applied. We have studied this phenomenon and found that the relative errors, which result when moving from one sample to the next, are directly proportional to m/z. Based on this observation, we developed a two-step calibration method, that overcomes said limitations. The first step is an external calibration with a high-order polynomial function used for the determination of the relation between m/z and t2, and the second step is a first-order internal correction for sample position-dependent errors. Applying this method, for instance, to a mass spectrum of a mixture of 18 peptides from a tryptic digest of a recombinant protein resulted in an average mass error of 1.0 ppm with a standard deviation of 3.5 ppm. When instead using a conventional two-point internal calibration, the average relative error was 2.2 ppm with a standard deviation of 15 ppm. The new method is described and its performance is demonstrated with examples relevant to proteome research.