title: Cibacron Blue and proteomics: the mystery of the platoon missing in action.
abstract: The use of Cibacron Blue columns (HiTrapBlue) in proteome analysis for removal of plasma albumin, for facilitating biomarker discovery, has not borne any fruit. In fact, the visibility of low-abundance proteins was obscured. It is here reported that, upon albumin sequestering from plasma, there is adsorption, via hydrophobic interaction, of a substantial number of plasma proteins, which are lost for subsequent analysis if the blue resin is eluted via an ion shock (2 M NaCl) or with a somewhat more robust eluant (5 M urea, 2 M thiourea, 2% CHAPS, 2% sulphobetain 3-10) as recommended by manufacturers. Such treatments, in fact, release at most 25 to 30 unique gene products, including albumin. If, however, the Affigel-Blue resin, after elution with either of the two above eluants, is further eluted with boiling 4% SDS in 25 mM DTT, all the missing proteins (amounting to at least 112 unique species) are desorbed and biomarker analysis can be conducted in a correct way. It is also suggested that such blue-resin treatment could be coupled to ProteoMiner adsorption, this coupled treatment further enhancing the chances of success for discovery of low-abundance proteins.
authors.affiliation: Department of Chemistry, Materials and Chemical Engineering Giulio Natta, Politecnico di Milano, Via Mancinelli 7, 20131 Milano, Italy.
authors.forename: Francesco
authors.initials: F
authors.lastname: Di Girolamo
authors.forename: Pier Giorgio
authors.initials: PG
authors.lastname: Righetti
authors.forename: Alfonsina
authors.initials: A
authors.lastname: D'Amato
authors.forename: Maxey C M
authors.initials: MC
authors.lastname: Chung
mesh_terms: D001798:Blood Proteins / Q000032:analysis*; D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D040901:Proteomics / Q000379:methods*; D012685:Sepharose / Q000031:analogs & derivatives* / Q000737:chemistry
chemical_list: D001798:Blood Proteins; C017842:cibacron blue-sepharose; D012685:Sepharose