title: Pre-analytical factors in clinical proteomics investigations: impact of ex vivo protein modifications for multiple sclerosis biomarker discovery.
abstract: 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 degrees C, 4 degrees C, -20 degrees C and -80 degrees C. We demonstrated that upon a -20 degrees 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.
authors: Centro Studi sull'Invecchiamento (Ce.S.I.), Fondazione G. d'Annunizio, Chieti, Italy.
authors: Damiana
authors: D
authors: Pieragostino
authors: Francesca
authors: F
authors: Petrucci
authors: Piero
authors: P
authors: Del Boccio
authors: Dante
authors: D
authors: Mantini
authors: Alessandra
authors: A
authors: Lugaresi
authors: Sara
authors: S
authors: Tiberio
authors: Marco
authors: M
authors: Onofrj
authors: Domenico
authors: D
authors: Gambi
authors: Paolo
authors: P
authors: Sacchetta
authors: Carmine
authors: C
authors: Di Ilio
authors: Giorgio
authors: G
authors: Federici
authors: Andrea
authors: A
authors: Urbani
mesh_terms: D000293:Adolescent; D000328:Adult; D000465:Algorithms; D015415:Biomarkers / Q000032:analysis* / Q000097:blood* / Q000378:metabolism; D001793:Blood Preservation / Q000009:adverse effects / Q000379:methods; D001798:Blood Proteins / Q000032:analysis / Q000378:metabolism; D015986:Confounding Factors, Epidemiologic; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D008954:Models, Biological; D009103:Multiple Sclerosis / Q000097:blood* / Q000175:diagnosis / Q000378:metabolism; D010865:Pilot Projects; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D055550:Protein Stability; D040901:Proteomics / Q000379:methods / Q000592:standards* / Q000706:statistics & numerical data; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods / Q000592:standards; D055815:Young Adult
chemical_list: D015415:Biomarkers; D001798:Blood Proteins