Surface Modification of Siliceous Materials Using Maleimidation and Various Functional Polymers Synthesized by Reversible Addition–Fragmentation Chain Transfer Polymerization

A novel surface modification method was investigated. The surface of siliceous materials was modified using polystyrene, poly­(acrylic acid), poly­(<i>N</i>-isopropylacrylamide), and poly­(<i>p</i>-acrylamidophenyl-α-mannoside) synthesized by reversible addition–fragmentation chain transfer polymerization. Thiol-terminated polymers were obtained by reduction of the thiocarbonate group using sodium borohydride. The polymers were immobilized on the surface via the thiol–ene click reaction, known as the Michael addition reaction. Immobilization of the polymers on the maleimidated surface was confirmed by X-ray photoelectron spectroscopy, infrared spectroscopy, and contact angle measurements. The polymer-immobilized surfaces were observed by atomic force microscopy, and the thickness of the polymer layers was determined by ellipsometry. The thickness of the polymer immobilized by the maleimide–thiol reaction was less than that formed by spin coating, except for polystyrene. Moreover, the polymer-immobilized surfaces were relatively smooth with a roughness of less than 1 nm. The amounts of amine, maleimide, and polymer immobilized on the surface were determined by quartz crystal microbalance measurements. The area occupied by the amine-containing silane coupling reagent was significantly less than the theoretical value, suggesting that a multilayer of the silane coupling reagent was formed on the surface. The polymer with low molecular weight had the tendency to efficiently immobilize on the maleimidated surface. When poly­(<i>p</i>-acrylamidophenyl-α-mannoside)-immobilized surfaces were used as a platform for protein microarrays, strong interactions were detected with the mannose-binding lectin concanavalin A. The specificity of poly­(<i>p</i>-acrylamidophenyl-α-mannoside)-immobilized surfaces for concanavalin A was compared with poly-l-lysine-coated surfaces. The poly-l-lysine-coated surfaces nonspecifically adsorbed both concanavalin A and bovine serum albumin, while the poly­(<i>p</i>-acrylamidophenyl-α-mannoside)-immobilized surface preferentially adsorbed concanavalin A. Moreover, the poly­(<i>p</i>-acrylamidophenyl-α-mannoside)-immobilized surface was applied to micropatterning with photolithography. When the micropattern was formed on the poly­(<i>p</i>-acrylamidophenyl-α-mannoside)-spin-coated surface by irradiation with ultraviolet light, the pattern of the masking design was not observed on the surface adsorbed with fluorophore-labeled concanavalin A using a fluorescent microscope because of elution of poly­(<i>p</i>-acrylamidophenyl-α-mannoside) from the surface. In contrast, fluorophore-labeled concanavalin A was only adsorbed on the shaded region of the poly­(<i>p</i>-acrylamidophenyl-α-mannoside)-immobilized surface, resulting in a distinctive fluorescent pattern. The surface modification method using maleimidation and reversible addition–fragmentation chain transfer polymerization can be used for preparing platforms for microarrays and micropatterning of proteins

Biotinylation of Silicon and Nickel Surfaces and Detection of Streptavidin as Biosensor

The availability of metal mesh device sensors has been investigated using surface-modified nickel mesh. Biotin was immobilized on the sensor surfaces consisting of silicon and nickel via a thiol–ene click reaction, known as the Michael addition reaction. Biotinylation on the maleimidated surface was confirmed by X-ray photoelectron spectroscopy. The binding of streptavidin to the biotinylated surfaces was evaluated using a quartz crystal microbalance and a metal mesh device sensor, with both techniques providing similar binding constant value. The recognition ability of the biotin immobilized using the thiol-maleimide method for streptavidin was comparable to that of biotin immobilized via several other methods. The adsorption of a biotin conjugate onto the streptavidin-immobilized surface via the biotin–streptavidin–biotin sandwich method was evaluated using a fluorescent microarray, with the results demonstrating that the biological activity of the streptavidin remained