PEG-Modified Macroporous Poly(Glycidyl Methacrylate) and Poly(2-Hydroxyethyl Methacrylate) Microspheres to Reduce Non-Specific Protein Adsorption Helena Hlı ´dkova ´, Daniel Hora ´k,* Vladimı ´r Proks, Zdenka Kuc ˇerova ´, Michal Peka ´rek, Jan Kuc ˇka 1. Introduction Non-specific adsorption of biomolecules such as proteins and peptides from biological media on the surface of polymer microspheres is a serious problem in many bioapplications. Biofouling decreases efficiency of func- tional proteins in peptide and polynucleotide synthesis, stationary phases for HPLC [1] and affinity chromatography, biological catalysts, [2] separation of bioactive substances, [3] drug delivery systems, [4] cell cultivation in tissue engineer- ing, [5] etc. Main types of bonds responsible for protein adsorption include hydrophobic [6] and electrostatic interactions, [7] or hydrogen [8] and affinity binding. [9] In order to reduce non- specific interactions in chromatography, binding mobile phase should flow at a relatively moderate ionic strength. At the same time, the packing material has to be hydrophilic and non-ionogenic. Here, site-directed immobilization of a hydrophilic biomolecule can be recommended using reactive functional groups of the support matrix. [10] Protein biofouling on a particle surface can be suppressed by treatment of the microspheres with natural polymers such as albumin, casein, and dextran. However, such coatings may have undesirable effects, e.g. contamination with bacteria, viruses or other residues. Therefore, water-soluble synthetic polymers, including poly(ethylene glycol) (PEG), have been suggested for surface coating. [11] PEGylation has been demonstrated to minimize the non-specific interaction and aggregation of particles in physiological media. [12–15] Although both PEG chain length and density are important factors to prevent protein adsorption, Full Paper H. Hlı ´dkova ´, Dr. D. Hora ´k, Dr. V. Proks, M. Peka ´rek, Dr. J. Kuc ˇka Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic, Heyrovsky ´ Sq. 2, 162 06 Prague 6, Czech Republic E-mail: horak@imc.cas.cz Dr. Z. Kuc ˇerova ´ First Faculty of Medicine, Institute of Pathophysiology, Charles University, U Nemocnice 5, 128 53 Prague 2, Czech Republic To minimize non-specific protein adsorption on macroporous poly(glycidyl methacrylate) and poly(2-hydroxyethyl methacrylate) microspheres containing amino and/or carboxyl groups, the microspheres are coated with a,v-bis-carboxy poly(ethylene glycol) and amino-terminated poly(ethylene glycol-co-propylene glycol) or a-methoxy-v-amino poly(ethylene glycol). Adsorption of bovine serum albumin (BSA), g-globulin, 125 I-BSA, pepsin, and chymotrypsin on neat and PEGylated microspheres is deter- mined by UV–VIS spectroscopy of supernatants and eluates or by measurement of radioactivity in an ionization chamber. Neat and PEGylated microspheres adsorb 0.8–70% and 0.02–44% of protein, respectively. Macromol. Biosci. 2013, 13, 503–511 ß 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim wileyonlinelibrary.com DOI: 10.1002/mabi.201200446 503