On-line Abstract Book

Mapping of the Human Blood Plasma Peptidome for Biomarkers Search

Agata Malinowska¹, Magdalena Bakun¹, Jaroslaw Poznanski¹, Dorota Sands², Michal Dadlez¹

Correspondence address: Agata Malinowska, Insitute of Biochemistry and Biophysics PAS, Mass Spectrometry Laboratory, Pawinskiego 5A, Warsaw, 02-106 Poland.

Keywords: Biomarkers; Fourier Transform ICR; Plasma, Human; Proteomic.

Novel aspect: Approach to data acquisition and analysis.

 

The low molecular weight fraction of human blood plasma or serum (peptidome) is believed to be a rich repository of important biological information. Mass spectrometric mapping of the human blood plasma peptidome has been suggested recently as a possible new diagnostic tool for early stage ovarian and lung cancer detection. The diagnostic value of this approach is currently the matter of intense discussion.

We have worked out a protocol for mapping of the human blood plasma peptidome by liquid chromatography and mass spectrometry using an LTQ FTICR mass spectrometer. Samples are first filtered on a 5 or 10 kDa membrane in the presence of a chaotropic agent to obtain the peptidome fraction, then peptide mixtures are resolved on RP-18 HPLC and analysed on a mass spectrometer. As a result two dimensional maps (molecular mass vs. HPLC retention time) of peptides present in the sample are obtained. Different parameters of sample preparation steps have been tested (filtration cutoff and temp., filter type, chaotropic agent, acetonitrile gradient, sample storage conditions and sample stability during analysis) and optimized. We have developed new bioinformatics tools for datasets analysis wich allow for data smoothing, peak picking, deisotoping and m/z-to-mass conversion. As a pilot study 45 human blood plasma samples from healthy donors were mapped and some of the peptides sequenced and assigned to plasma proteins. This set is the basis for a statistical analysis of the contents of peptidome maps. In cooperation with dr. Dorota Sands (Institute of Mother and Child, Warsaw) also 45 samples from patients affected by cystic fibrosis and their family members were analysed in search for disease prognostic markers. The comparison of obtained datasets was made in cooperation with a group of prof. J. Tiuryn (University of Warsaw) using existing statistical tools.