Pre-analytical factors in clinical proteomics investigations: Impact of ex vivo protein modifications for multiple sclerosis biomarker discovery

Damiana Pieragostino, Francesca Petrucci, Piero Del Boccio, Dante Mantini, Alessandra Lugaresi, Sara Tiberio, Marco Onofrj, Domenico Gambi, Paolo Sacchetta, Carmine Di Ilio, Giorgio Federici, Andrea Urbani

Research output: Contribution to journalArticlepeer-review


Serum proteome investigations have raised an incredible interest in the research of novel molecular biomarker, nevertheless few of the proposed evidences have been translated to the clinical practice. One of the limiting factors has been the lack of generally accepted guidelines for clinical proteomics studies and the lack of a robust analytical and pre-analytical ground for the proposed classification models. Pre-analytical issues may results in a deep impact for biomarker discovery campaign. In this study we present a systematic evaluation of sample storage and sampling conditions for clinical proteomics investigations. We have developed and validated a linear MALDI-TOF-MS protein profiling method to explore the low protein molecular weight region (5-20 kDa) of serum samples. Data normalization and processing was performed using optimise peak detection routine (LIMPIC) able to describe each group under investigation. Data were acquired either from healthy volunteers and from multiple sclerosis patients in order to highlight ex vivo protein profile alteration related to different physio-pathological conditions. Our data showed critical conditions for serum protein profiles depending on storage times and temperatures: 23 °C, 4 °C, - 20 °C and - 80 °C. We demonstrated that upon a - 20 °C short term storage, characteristic degradation profiles are associated with different clinical groups. Protein signals were further identified after preparative HPLC separation by peptide sequencing on a nanoLC-Q-TOF TANDEM mass spectrometer. Apolipoprotein A-IV and complement C3 protein fragments, transthyretin and the oxidized isoforms in different apolipoprotein species represent the major molecular features of such a degradation pattern.

Original languageEnglish
Pages (from-to)579-592
Number of pages14
JournalJournal of Proteomics
Issue number3
Publication statusPublished - Jan 3 2010


  • Clinical Proteomics
  • Mass spectrometry
  • Neurodegenerative disease
  • Pre-analytical validation

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics


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