Immobilization of Proteins on the Surface of Silanized Hydroxyapatite
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K9.18.1
Immobilization of proteins on the surface of silanized hydroxyapatite Akira Monkawa1, Toshiyuki Ikoma1,2, Yuri Kumagai1,2, Syunji Yunoki1, Junzo Tanaka1,2 1 Biomaterials Center, National Institute for Material Science, Tsukuba, Japan. 2 CREST, Japan Science and Technology Corporation, Saitama, Japan.
Abstract Hydroxyapatite (HAp) is widely used as bioceramics for bone and dental tissue reconstructions due to its excellent biocompatibility with hard tissues and high osteoconductivity. Although it is well known that HAp has the high adsorbed ability, its ability is decreased in high ionic solution. To immobilize protein on the HAp surface, this study demonstrates that Collagen (Col) and fibronectin (Fnt) were immobilized on the surface of hydroxyapatite (HAp) sintered body with and without the silane coupling agent of aminopropyltriethoxysilane (APS). The proteins immobilized on the HAp and APS/HAp surfaces were investigated by atomic force microprobe (AFM, Simadzu; SPM 9500) analyses and ζ potential measurements. The stability of protein/APS/HAp and protein/HAp composites was evaluated after immersion in phosphate buffer and NaCl solutions with various concentrations. AFM analyses and ζ potential measurements revealed that proteins immobilized on the APS/HAp are more stable than those immobilized on the HAp in high ionic solutions. Introduction Surface modification is of great importance for improving the initial cell attachment and proliferation of cells. The interfacial interaction between biomaterials and cells can be controlled by proteins on the surface of materials. Hydroxyapatite (HAp: Ca10(PO4)6(OH)2) has an interesting property as an adsorbent for proteins. The absorption ability of HAp is useful for the preparation of protein/HAp composites; however the proteins are easily removed from the HAp surface depended on the ionic strengths and pH values in aqueous systems. Therefore, it is difficult to hold proteins on the HAp surface in vivo. To overcome these problems, the introduction of chemical bonds between protein and HAp is a useful technique. Silane coupling agent is a binding material with the ability to bond inorganic materials such as glass, mineral fillers, metals and metallic oxides to organic resins. The previous studies show that the silane coupling agent is useful material for coating on the HAp surface [1], however the previous experimental studies using silane coupling agent to bond protein to HAp are limited [2]. This study focused on the coupling agent as a binding material between HAp and two different proteins such as acidic proteins of fibronectin (Fnt) and basic proteins of collagen (Col). We attempt to immobilize Fnt and Col on the HAp sintered body by using the coupling agent. The presence of silane was confirmed by ζ potential measurements, and that of adsorbed proteins was confirmed by AFM images and ζ potential measurements.
K9.18.2
Materials and Methods HAp sintered body (HAp disk) was prepared by the reported method [3]. HAp disks were polished with diamond pastes of 1µ
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