title: [PHEMA/PEI]-Cu(II) based immobilized metal affinity chromatography cryogels: Application on the separation of IgG from human plasma.
abstract: The immobilized metal-affinity chromatography (IMAC) has gained significant interest as a widespread separation and purification tool for therapeutic proteins, nucleic acids and other biological molecules. The enormous potential of IMAC for proteins with natural surface exposed-histidine residues and for recombinant proteins with histidine clusters. Cryogels as monolithic materials have recently been proposed as promising chromatographic adsorbents for the separation of biomolecules in downstream processing. In the present study, IMAC cryogels have been synthesized and utilized for the adsorption and separation of immunoglobulin G (IgG) from IgG solution and whole human plasma. For this purpose, Cu(II)-ions were coupled to poly(hydroxyethyl methacrylate) PHEMA using poly(ethylene imine) (PEI) as the chelating ligand. In this study the cryogels formation optimized by the varied proportion of PEI from 1% to 15% along with different amounts of Cu (II) as chelating metal. The prepared cryogels were characterized by scanning electron microscopy, Fourier transform infrared spectroscopy, and thermogravimetric analysis. The [PHEMA/PEI]-Cu(II) cryogels were assayed for their capability to bind the human IgG from aqueous solutions. The IMAC cryogels were found to have high affinity toward human IgG. The adsorption of human IgG was investigated onto the PHEMA/PEI cryogels with (10% PEI) and the concentration of Cu (II) varied as 10, 50, 100 and 150 mg/L. The separation of human IgG was achieved in one purification step at pH7.4. The maximum adsorption capacity was observed at the [PHEMA/PEI]-Cu(II) (10% PEI) with 72.28 mg/g of human IgG. The purification efficiency and human IgG purity were investigated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).
authors.affiliation: Department of Chemistry, Biochemistry Division, Hacettepe University, Ankara, Turkey.
authors.forename: Monireh
authors.initials: M
authors.lastname: Bakhshpour
authors.affiliation: Department of Chemistry, Biochemistry Division, Hacettepe University, Ankara, Turkey.
authors.forename: Ali
authors.initials: A
authors.lastname: Derazshamshir
authors.affiliation: Department of Chemistry, Biochemistry Division, Hacettepe University, Ankara, Turkey.
authors.forename: Nilay
authors.initials: N
authors.lastname: Bereli
authors.affiliation: Laboratory of "Valorisation des Ressources Naturelles et Produits de Santé (VRNPS)", Doctoral School of Sciences and Technology, Lebanese University, Rafic Hariri University Campus, Hadath, Lebanon.
authors.forename: Assem
authors.initials: A
authors.lastname: Elkak
authors.affiliation: Department of Chemistry, Biochemistry Division, Hacettepe University, Ankara, Turkey. Electronic address: denizli@hacettepe.edu.
authors.forename: Adil
authors.initials: A
authors.lastname: Denizli
mesh_terms: D000327:Adsorption; D002614:Chelating Agents / Q000737:chemistry; D002846:Chromatography, Affinity; D003300:Copper / Q000737:chemistry*; D059985:Cryogels / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007097:Imines / Q000737:chemistry*; D007074:Immunoglobulin G / Q000097:blood / Q000302:isolation & purification*; D009994:Osmolar Concentration; D011095:Polyethylenes / Q000737:chemistry*; D011102:Polyhydroxyethyl Methacrylate / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared; D013696:Temperature; D013818:Thermogravimetry
chemical_list: D002614:Chelating Agents; D059985:Cryogels; D007097:Imines; D007074:Immunoglobulin G; D011095:Polyethylenes; C505405:poly(ethylene imine); D011102:Polyhydroxyethyl Methacrylate; D003300:Copper
keywords: Antibody purification; Cryogels; Human IgG; IMAC