title: Anion-exchange chromatography using short monolithic columns as a complementary technique for human serum albumin depletion prior to human plasma proteome analysis.
abstract: In order to enable the detection of low abundance proteins from human plasma, it is necessary to remove high abundance proteins. Among them, human serum albumin and immunoglobulin G represent more than 75% of all such proteins. In this paper, the characterization of short monolithic columns was performed followed by the optimization of a multidimensional approach, known as conjoint liquid chromatography, to deplete human serum albumin and immunoglobulin G from a human plasma sample. Two different chromatographic modes were used: ion-exchange chromatography and affinity chromatography. A monolithic stationary phase (convective interaction media disk) bearing strong anion-exchange groups and another immobilized with protein G were placed in series into one housing. The optimal binding conditions were found that removed a majority of human serum albumin and immunoglobulin G from the human plasma sample. This method was compared to the depletion using a combination of pseudo-affinity and affinity columns. The results of the human serum albumin and immunoglobulin G depletion were confirmed by 2D electrophoresis. It has been shown that anion-exchange and affinity chromatography using convective interaction media monolithic columns can represent an efficient complementary technique for human serum albumin and immunoglobulin G removal from human plasma.
authors.affiliation: Lek Pharmaceuticals d.d., Verovskova 57, SI-1526 Ljubljana, Slovenia.
authors.forename: Tatjana Cerk
authors.initials: TC
authors.lastname: Petric
authors.forename: Peter
authors.initials: P
authors.lastname: Brne
authors.forename: Bostjan
authors.initials: B
authors.lastname: Gabor
authors.forename: Lidija
authors.initials: L
authors.lastname: Govednik
authors.forename: Milos
authors.initials: M
authors.lastname: Barut
authors.forename: Ales
authors.initials: A
authors.lastname: Strancar
authors.forename: Lucija Zupancic
authors.initials: LZ
authors.lastname: Kralj
mesh_terms: D015415:Biomarkers; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000737:chemistry; D007525:Isoelectric Focusing; D008024:Ligands; D010949:Plasma / Q000737:chemistry; D020543:Proteome / Q000737:chemistry*; D012709:Serum Albumin / Q000737:chemistry*
chemical_list: D015415:Biomarkers; D007074:Immunoglobulin G; D008024:Ligands; D020543:Proteome; D012709:Serum Albumin