Hindawi Publishing Corporation
Scienti�ca
Volume 2012, Article ID 126170, 28 pages
http://dx.doi.org/10.6064/2012/126170
Research Article
Mechanisms for Covalent Immobilization of Horseradish
Peroxidase on Ion-Beam-Treated Polyethylene
Alexey V. Kondyurin, Pourandokht Naseri, Jennifer M. R. Tilley, Neil J. Nosworthy,
Marcela M. M. Bilek, and David R. McKenzie
Applied and Plasma Physics, School of Physics, University of Sydney, A28, Sydney, NSW 2006, Australia
Correspondence should be addressed to Alexey V. Kondyurin; a.kondyurin@physics.usyd.edu.au
Received 21 October 2012; Accepted 20 November 2012
Academic Editors: E. A. Abou Neel, S. S. Banerjee, and P. Ferruti
Copyright © 2012 Alexey V. Kondyurin et al. is is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly
cited.
e surface of polyethylene was modi�ed by plasma immersion ion implantation. Structure changes including carbonization and
oxidation were observed. High surface energy of the modi�ed polyethylene was attributed to the presence of free radicals on the
surface. e surface energy decay with storage time aer treatment was explained by a decay of the free radical concentration
while the concentration of oxygen-containing groups increased with storage time. Horseradish peroxidase was covalently attached
onto the modi�ed surface by the reaction with free radicals. Appropriate blocking agents can block this reaction. All aminoacid
residues can take part in the covalent attachment process, providing a universal mechanism of attachment for all proteins. e
native conformation of attached protein is retained due to hydrophilic interactions in the interface region. e enzymatic activity of
covalently attached protein remained high. e long-term activity of the modi�ed layer to attach protein is explained by stabilisation
of unpaired electrons in sp
2
carbon structures. A high concentration of free radicals can give multiple covalent bonds to the protein
molecule and destroy the native conformation and with it the catalytic activity. e universal mechanism of protein attachment to
free radicals could be extended to various methods of radiation damage of polymers.
1. Introduction
e attachment of proteins to polymer surfaces provides a
means of modifying the response of an organism to the
surface and is therefore an important step for improving
the biocompatibility and functionality of medical implant
table devices [1]. In medicine, the immune response can
cause adverse reactions to implanted prosthetics or during
operations in which blood is exposed to the surfaces of
medical devices such as heart-lung machines. In biosensors,
attached proteins may be used to detect the presence of
molecules in the environment. e attached protein must be
strongly bound to the surface to prevent it being washed off
under operational conditions including a high rate of �ow of
liquid across the surface [2]. Additionally the surface must
allow the protein to preserve its bioactivity [3].
Given the rigorous protocols that must be undertaken
to obtain approvals for the use of new polymer materials in
medical applications, it is preferable to modify the surface of
an existing polymer than to develop an entirely new polymer
material. e preparation of polymer surfaces for protein
binding can be done using a number of chemical and physical
modi�cations, such as the attachment of linker molecules
to provide covalent binding through speci�c active groups
[4, 5] plasma treatment [6–12] UV treatment [13–16], and
ion beam implantation [17–20].
Despite its importance in applications, the attachment
mechanism of proteins on polymer surfaces is not yet well
understood [21–27]. e uncertainty of the mechanism could
make the behaviour of the surface unpredictable in a given
application. In a limited number of applications, the physical
adsorption on a polymer surface is acceptable. However,
in some applications, the stronger attachment provided by
covalent bonding is desirable and linker groups have been
employed to achieve this [28]. e mechanism of attachment
provided by linker groups is usually assumed to be covalent
bonding via the chemistry that is expected for the active
group of the linker and the protein, based on a general