Blood is a complex liquid tissue including a corpuscular and a non-corpuscular fractions and represent the major connection between cells and tissues of an organism, allowing transport of oxygen and many molecules (i. e. hormones, amino acids, carbohydrates, lipids, vitamins, mineral salts and water), and the reject of waste substances.
The proteome/peptidome of a blood sample is directly linked to the metabolic state and utilizable to provide information on the physiological and/or pathological processes occurring in the body. The proteins and peptides present in serum and plasma, the non-corpuscular blood fractions, are a combination of those playing circulatory functions and those secreted or released into the circulation from cells both during normal biological events and in diseases .
Because serum and plasma are the most available clinical samples, they represent one of the most convenient choices for minimal-invasive detection of peptides potentially relevant as specific disease-biomarkers. Naturally occurring peptides typically derive from cleavages of the most abundant full-length proteins as a result of numerous proteases activity contained in these fluids or produced by cells .
As reported, a high number of peptides are only present in serum and not detectable in plasma. In fact, more than 40% of the peptides detected in serum are generated by ex vivo processes during specimen collection and preparation, introducing peptide signals which have to be interpreted with prudence . Serum generation is associated with the coagulation cascade and the activation of complement system, processes leading cell lysis and multiproteinase activation, which cleave various proteins producing a lot of peptides. Moreover, peptides are released from the blood clot during the specimen collection [i. e. thymosin β4 (Tβ4), zyxin]. In plasma preparation the addition of chelating agents deeply stabilized the protein/peptide composition [8, 9]. However, for several clinical analyses serum is preferred because the anticoagulants can sometimes interfere with the results. Whatever the fluid, peptidomic analysis of serum and/or plasma is critical due to the wide range of protein and peptide concentrations that spans over ten orders of magnitude . For this reason the first goal of the Plasma Proteome Project of the Human Proteome Organization is to analyze serum and plasma specimens to elucidate their characteristics and to create a global, open-source knowledge base/data repository [10, 11], as well as to standardize the procedures of collection, handling and storage of blood specimens [8, 9].
Pre-treatment of serum and plasma samples for top-down peptidomic analysis is particularly critical due to the high complexity of these biological samples, due to the large span of molecular weight distribution of peptides and the biological variability (i. e. gender, age, genetic, environmental, dietary, and psychological factors) . Several pre-treatment...