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Identification of free phosphopeptides in different biological fluids by a mass spectrometry approach

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Abstract

Human body fluids have been rediscovered in the post-genomic era as a great source of biological markers and perhaps as source of potential biomarkers of disease. Recently, it has been found that not only proteins but also peptides and their modifications can be indicators of early pathogenic processes. This paper reports the identification of free phosphopeptides in human fluids using an improved IMAC strategy coupled to iterative mass spectrometry-based scanning techniques (neutral loss, precursor ion, multiple reaction monitoring). Many peptides were detected in the enriched extract samples when submitted to the MS-integrated strategy, whereas they were not detected in the initial extract samples. The combination of the IMAC-modified protocol with selective “precursor ion” and constant “neutral loss” triple quadrupole scan modes confers a high sensitivity on the analysis, allowing rapid phosphopeptide identification and characterization, even at low concentrations. To the best of our knowledge this work represents the first report exclusively focused on the detection of free phosphorylated peptides in biological fluids.

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Acknowledgments

This work was supported by grants from the INBB, the Ministero dell’Università e della Ricerca Scientifica (Progetti di Rilevante Interesse Nazionale 2002, 2003, 2005, 2006; FIRB 2001). Support from the National Center of Excellence in Molecular Medicine (MIUR, Rome) and from the Regional Center of Competence (CRdC ATIBB, Regione Campania, Naples) is gratefully acknowledged.

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Authors and Affiliations

  1. Department of Organic Chemistry and Biochemistry, Federico II University of Naples, 80126, Naples, Italy

    Claudia Cirulli & Giovanni Chiappetta

  2. Department of Organic Chemistry and Biochemistry, School of Biotechnological Sciences, Federico II University of Naples, 80126, Naples, Italy

    Gennaro Marino & Angela Amoresano

  3. Institute of Biomedical Technologies, National Research Council, 20090 Segrate, Milan, Italy

    Pierluigi Mauri

  4. Dipartimento di Chimica Organica e Biochimica, Università degli Studi Federico II, Complesso Universitario Monte S. Angelo, via Cynthia 4, 80126, Napoli, Italy

    Angela Amoresano

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  1. Claudia Cirulli
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  2. Giovanni Chiappetta
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  3. Gennaro Marino
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  4. Pierluigi Mauri
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  5. Angela Amoresano
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Correspondence to Angela Amoresano.

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Cirulli, C., Chiappetta, G., Marino, G. et al. Identification of free phosphopeptides in different biological fluids by a mass spectrometry approach. Anal Bioanal Chem 392, 147–159 (2008). https://doi.org/10.1007/s00216-008-2266-7

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  • DOI: https://doi.org/10.1007/s00216-008-2266-7

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