Chapter 11
Biosensor Design and Application Downloaded from pubs.acs.org by UNIV OF CALIFORNIA SANTA BARBARA on 09/23/18. For personal use only.
Concanavalin A and Polysaccharide on Gold Surfaces Study Using Surface Plasmon Resonance Techniques 1
1
2
R. F. DeBono , U. J. Krull , and Gh. Rounaghi 1
Chemical Sensors Group, Erindale Campus, University of Toronto, Mississauga, Ontario L5L 1C6, Canada Department of Chemistry, Faculty of Science, Mashhad University, Mashhad, Iran
2
Dynamic studies involving deposition and subsequent denaturation of selective binding proteins, or of complexes formed by selective interactions, provide insight into problems of reversibility and useful lifetimes of biosensors. The adsorption of concanavalin A (Con A) in the presence and absence of Ca and Mn onto polycrystalline gold surfaces and subsequent interactions of the protein with the polysaccharides glycogen and dextran have been investigated in situ using surface plasmon resonance (SPR) spectroscopy and surface plasmon microscopy (SPM). The adsorption of concanavalin A and subsequent reactions of the protein with the polysaccharides were followed by observing changes of reflectivity at a fixed angle of incidence as a function of time. Our results suggest that Con A exhibits non-Langmuirianadsorption kinetics. The polysaccharide binding involves two Langmuirian processes based on adhesion and subsequent dehydration of the polysaccharide layer leading to compaction. The negligible effect of ethylenediaminetetraacetic acid (EDTA) on the reversal of the complexation reaction between dextran and Con A at the surface provides further evidence of the dense structure of this layer. Characterization by SPM shows these surfaces to be smooth and homogeneous on the 4 μm scale except for the Au/glycogen/Con A system which was allowed to react for periods greater than 10 hrs. 2+
0097-6156/92/0511-0121$06.00/0 © 1992 American Chemical Society
2+
122
BIOSENSOR DESIGN AND APPLICATION
Recent interest in the areas of biocompatibility and development of biosensors has prompted studies of the adsorption or covalent deposition of proteins and polymers onto solid surfaces (1-4). Changes in the conformation or orientation of proteins or protein-ligand complexes at solid/liquid interfaces during and after the adsorption process may occur and can result in the denaturation of the protein or complexes (5,6). Such processes can determine the useful lifetime and reversibility of biosensors. Applications involving the close contact of biological materials with a foreign surface must account for surface induced changes. The optical techniques of surface plasmon resonance (SPR) spectroscopy and microscopy (SPM) can be used for the physical characterization and real time observation of dynamic events at an organic film located in close proximity to a metal surface. One system that is representative of a selective interaction is the adsorption of concanavalin A (Con A ) to a polycrystalline gold surface, and the subsequent selective interaction of the protein with a polysaccharide. Con A is a well characterized lectin, found in jack bean, which reacts to form a precipitate with a restricted group of branched polysaccharides (a-Dmannopyranosyl) in a manner analogous to an antibody-antigen reaction (8). A t p H 7.0 Con A exists as a tetramer( if ionic strength > 0.3) with a net negative charge (9). The isoelectric point is approximately 5. A t low ionic strength (
