Magnetic IDA-modified hydrophilic methacrylate-based polymer microspheres for IMAC protein separation.
Preparation of a new type of magnetic non-porous poly(2-hydroxyethyl methacrylate-co-glycidyl methacrylate) microspheres with hydrophilic properties containing coupled iminodiacetic acid (IDA) is described. The prepared microspheres were used for the immobilization of Ni(II) or Fe(III) ions to show their application in protein binding studies. Human IgG was bound to magnetic Ni(II)-IDA-modified microspheres and conditions of its adsorption and elution were optimized. Non-specific binding of the protein to magnetic microspheres in the absence of Ni(II) ions was low. Fe(III) ions immobilized on magnetic IDA-modified microspheres were used for the specific binding of porcine pepsin, as a model phosphoprotein. The ability of phosphate buffer to release the adsorbed enzyme from the microspheres and a low adsorption of the dephosphorylated protein indicate the participation of phosphate groups in the pepsin interaction. The elaborated method represents a rapid technique that can be used not only for the separation of proteins but also for analytical purposes.
Prikryl
Petr
P
Institute of Pathophysiology, 1st Faculty of Medicine and Centre of Experimental Hematology, Charles University, Prague, Czech Republic. petr.prikryl@lf1.cuni.cz
Horák
Daniel
D
Tichá
Marie
M
Kucerová
Zdenka
Z
D000327:Adsorption; D000818:Animals; D002623:Chemistry Techniques, Analytical / Q000295:instrumentation / Q000379:methods*; D002846:Chromatography, Affinity / Q000379:methods*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007098:Imino Acids / Q000032:analysis* / Q000737:chemistry; D007074:Immunoglobulin G / Q000737:chemistry; D007477:Ions; D008280:Magnetics; D008689:Methacrylates / Q000737:chemistry*; D008863:Microspheres; D011108:Polymers / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D013552:Swine
D007098:Imino Acids; D007074:Immunoglobulin G; D007477:Ions; D008689:Methacrylates; D011108:Polymers; D011506:Proteins; C008109:iminodiacetic acid