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Open Access Technical Note

Label-free peptide profiling of Orbitrap™ full mass spectra

Mark K Titulaer12*, Dominique de Costa3, Christoph Stingl1, Lennard J Dekker1, Peter AE Sillevis Smitt1 and Theo M Luider1*

Author Affiliations

1 Laboratory of Neuro-Oncology and Clinical and Cancer Proteomics, Department of Neurology, Erasmus University Medical Center, Dr. Molewaterplein 50, P.O. Box 2040, 3000 CA Rotterdam, The Netherlands

2 Academic Medical Center, University of Amsterdam, Meibergdreef 9, P.O. Box 22660, 1100 DD Amsterdam, The Netherlands

3 Department of Pulmonology, Erasmus University Medical Center, Dr. Molewaterplein 50, P.O. Box 2040, 3000 CA Rotterdam, The Netherlands

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BMC Research Notes 2011, 4:21  doi:10.1186/1756-0500-4-21

Published: 27 January 2011

Abstract

Background

We developed a new version of the open source software package Peptrix that can yet compare large numbers of Orbitrap™ LC-MS data. The peptide profiling results for Peptrix on MS1 spectra were compared with those obtained from a small selection of open source and commercial software packages: msInspect, Sieve™ and Progenesis™. The properties compared in these packages were speed, total number of detected masses, redundancy of masses, reproducibility in numbers and CV of intensity, overlap of masses, and differences in peptide peak intensities. Reproducibility measurements were taken for the different MS1 software applications by measuring in triplicate a complex peptide mixture of immunoglobulin on the Orbitrap™ mass spectrometer. Values of peptide masses detected from the high intensity peaks of the MS1 spectra by peptide profiling were verified with values of the MS2 fragmented and sequenced masses that resulted in protein identifications with a significant score.

Findings

Peptrix finds about the same number of peptide features as the other packages, but peptide masses are in some cases approximately 5 to 10 times less redundant present in the peptide profile matrix. The Peptrix profile matrix displays the largest overlap when comparing the number of masses in a pair between two software applications. The overlap of peptide masses between software packages of low intensity peaks in the spectra is remarkably low with about 50% of the detected masses in the individual packages. Peptrix does not differ from the other packages in detecting 96% of the masses that relate to highly abundant sequenced proteins. MS1 peak intensities vary between the applications in a non linear way as they are not processed using the same method.

Conclusions

Peptrix is capable of peptide profiling using Orbitrap™ files and finding differential expressed peptides in body fluid and tissue samples. The number of peptide masses detected in Orbitrap™ files can be increased by using more MS1 peptide profiling applications, including Peptrix, since it appears from the comparison of Peptrix with the other applications that all software packages have likely a high false negative rate of low intensity peptide peaks (missing peptides).