title: Selectivity of monolithic supports under overloading conditions and their use for separation of human plasma and isolation of low abundance proteins.
abstract: Human serum albumin (HSA) and immunoglobulin G (IgG) represent over 75% of all proteins present in human plasma. These two proteins frequently interfere with detection, determination and purification of low abundance proteins that can be potential biomarkers and biomarker candidates for various diseases. Some low abundance plasma proteins such as clotting factors and inhibitors are also important therapeutic agents. In this paper, the characterization of ion-exchange monolithic supports under overloading conditions was performed by use of sample displacement chromatography (SDC). If these supports were used for separation of human plasma, the composition of bound and eluted proteins in both anion- and cation-exchange mode is dependent on column loading. Under overloading conditions, the weakly bound proteins such as HSA in anion-exchange and IgG in cation-exchange mode are displaced by stronger binding proteins, and this phenomenon was not dependent on column size. Consequently, small monolithic columns with a column volume of 100 and 200 μL are ideal supports for high-throughput screening in order to develop new methods for separation of complex mixtures, and for sample preparation in proteomic technology.
authors.affiliation: Institute of Immunology, Zagreb, Croatia.
authors.forename: Marija
authors.initials: M
authors.lastname: Brgles
authors.forename: James
authors.initials: J
authors.lastname: Clifton
authors.forename: Robert
authors.initials: R
authors.lastname: Walsh
authors.forename: Feilei
authors.initials: F
authors.lastname: Huang
authors.forename: Marijana
authors.initials: M
authors.lastname: Rucevic
authors.forename: Lulu
authors.initials: L
authors.lastname: Cao
authors.forename: Douglas
authors.initials: D
authors.lastname: Hixson
authors.forename: Egbert
authors.initials: E
authors.lastname: Müller
authors.forename: Dj
authors.initials: Dj
authors.lastname: Josic
mesh_terms: D001798:Blood Proteins / Q000302:isolation & purification*; D002852:Chromatography, Ion Exchange / Q000295:instrumentation / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry; D012709:Serum Albumin / Q000737:chemistry; D012965:Sodium Chloride / Q000737:chemistry; D053719:Tandem Mass Spectrometry
chemical_list: D001798:Blood Proteins; D007074:Immunoglobulin G; D012709:Serum Albumin; D012965:Sodium Chloride