Analytical applications of surface plasmon resonance techniques

In situ studies of the adsorption of alkyl thiols and concanavalin A (Con A) to gold were investigated using surface plasmon resonance spectroscopy (SPR) and surface plasmon microscopy (SPM). In addition, the use of silver island films in the design and construction of fluorescently based sensors was examined. SPR experiments indicated that hexane (C{dollar}sb6),{dollar} dodecane(C{dollar}sb{lcub}12{rcub}){dollar} and hexadecane (C{dollar}sb{lcub}16{rcub}){dollar} thiol adsorption to gold surfaces occurred by a two step process, involving fast adsorption to the surface to give 80% (C{dollar}sb{lcub}12{rcub},{dollar} C{dollar}sb{lcub}16{rcub}){dollar} or 50% (C{dollar}sb6){dollar} coverage followed by a slow (100 fold slower) rearrangement of the adsorbed thiol. SPM shows these surfaces to be smooth and homogenous on the scale larger than 4 {dollar}mu{dollar}m. SPR experiments indicated that Con A adsorbed to gold surfaces initially by a diffusion limited process until a critical coverage was achieved. The time dependence of subsequent protein adsorption suggested the existence of two distinct binding site populations. Subsequent polysaccharide binding involved a two step process. This was based on the adsorption and subsequent dehydration of the polysaccharide layer leading to compaction. SPM showed these surfaces to be smooth and homogeneous on the 4 {dollar}mu{dollar}m scale except for the Au/glycogen/Con A system which was allowed to react for periods greater than 10 hours. Behenic acid (BA) monolayers, containing 0.1 to 1 mol%, of the fluorophores nitrobenzoxadiazole (NBD) and 6-tetramethylrhodamine-thiocarbamoyl (TRITC) covalently attached to the head group of the lipid dipalmitoyl-L-phosphatidyl-ethanolamine (PE), were deposited by Langmuir-Blodgett (LB) techniques onto glass and 40 A thick silver island films. Fluorescence was observed for both the NBD and TRITC films deposited on silver island films with average island diameters greater than 200 A. The fluorescence responses resulting from structural changes in a 1 mol% NBD-PE/BA monolayer on a glass surface and a bilayer on a silver island surface induced by exposure to water vapour and a mixture of water and acetic acid vapour were found to be reproducible and reversible upon hydration. An amplified fluorescence response was observed for the silver island surface. Partitioning of acetic acid into the bilayer on the silver island surface further amplified the fluorescence response to subsequent hydration.