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Clinical Chemistry 50: 545-551, 2004. First published January 15, 2004; 10.1373/clinchem.2003.028209
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(Clinical Chemistry. 2004;50:545-551.)
© 2004 American Association for Clinical Chemistry, Inc.

Proteomics and Protein Markers

Mass Spectrometric Phenotyping of Val34Leu Polymorphism of Blood Coagulation Factor XIII by Differential Peptide Display

Harald Tammen1,a, Thomas Möhring1, Markus Kellmann1, Andreas Pich2, Hans H. Kreipe2 and Rüdiger Hess1

1 BioVisioN AG, Hannover, Germany.2 Institute of Pathology, Medical School Hannover, Hannover, Germany.

aAddress correspondence to this author at: BioVisioN AG, Feodor Lynen Strasse 5, 30625 Hannover, Germany. Fax 49-511-53889666; e-mail h.tammen{at}biovision-discovery.de.

Background: The Val34Leu mutation in the activation peptide of factor XIII (FXIIIA) correlates with a lower incidence of myocardial infarction and ischemic stroke but an increased risk for hemorrhagic stroke. We describe mass spectrometric detection of the activation peptide variants in human serum.

Methods: We used differential peptide display (DPD) to compare comprehensive peptide maps from pairs of serum samples from healthy volunteers. Peptides were separated by liquid chromatography, and fractions were subjected to mass spectrometry. Mass spectra of all fractions were combined, giving a peptide map representing a two-dimensional display of peptide masses. After comparison of peptide mass maps, peptides that differentiated FXIIIA phenotypes were identified by mass spectrometry.

Results: Val34Leu polymorphisms of the activation peptide of FXIIIA were identified in 20 serum samples from 10 volunteers by DPD, and their sequences were confirmed by nanoelectrospray-ionization quadrupole time-of-flight mass spectrometry. Analysis of three (V34V, V34L, and L34L) phenotypes was confirmed by allele-specific genotypic analysis in all (n = 10) volunteers.

Conclusion: DPD provides a simple and easy-to-use phenotype assay with advantages over PCR-based assays in being faster and directly analyzing the compound of interest.




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