ABSTRACT
Porous silicon dioxide surfaces have been used for monitoring the specific affinity binding of low molecular weight molecules to streptavidin. Streptavidin was immobilized to the porous silicon dioxide surface by spontaneous adsorption at pH 7.4. Binding of biotin and an oligopeptide synthesized by means of combinatorial chemistry were monitored with an in situ null ellipsometer. Measurements were also done with hydroxy-azobenzene-2-carboxylic acid and DL-6-8-thioctic acid amide. The performance of porous silicon dioxide as a potential surface in biosensor applications was compared with a planar silicon dioxide surface. Porous silicon dioxide showed a 10-fold amplification of the response compared to planar silicon dioxide. It was possible to monitor the binding of biotin and the oligopeptide in the concentration range 2-40 microM. A response time as low as 30 s was obtained for the oligopeptide at 40 microM.
Subject(s)
Biosensing Techniques , Silicon , Streptavidin , Adsorption , Amino Acid Sequence , Biotin , Lasers , Molecular Sequence Data , Molecular Weight , Oxidation-Reduction , Porosity , Protein ConformationABSTRACT
A technique using an optical instrument, a reflectometer, for quantitative determination of proteinase activity in microliter samples of complex serum-like fluids, e.g., crevicular exudate from single sites, was developed. The technique allowed the use of various proteins as enzyme substrate. The reflectometer measures the mass of a layer, such as protein, adsorbed to a reflecting surface. This is done by measuring the reflected light intensity of the p-polarized light beam on a surface. We used methylized silicon surfaces that were coated with fibrinogen, alpha 2-macroglobulin, or hemoglobin as enzyme substrates. The test solution was incubated overnight in a basin made in an agar gel applied on the top of the protein-coated surface. In 82 exudates from periodontitis sites, with pocket depths greater than or equal to 6 mm, fibrinogenolytic activity corresponding to 1 microgram ml-1 of trypsin and pronase P was found in 20% of the samples.
