Cibacron Blue and proteomics: The mystery of the platoon missing in action

Francesco Di Girolamo, Pier Giorgio Righetti, Alfonsina D'Amato, Maxey C M Chung

Research output: Contribution to journalArticlepeer-review

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.

Original languageEnglish
Pages (from-to)2856-2865
Number of pages10
JournalJournal of Proteomics
Volume74
Issue number12
DOIs
Publication statusPublished - Nov 18 2011

Keywords

  • Biomarkers
  • Combinatorial peptide ligand libraries
  • Dye ligand affinity chromatography
  • Plasma

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics

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