A new fluorescence-based biosensor is presented. The biosensing scheme is based on the fact that a fluorophore in close proximity to a metal film (<100 A) experiences strong quenching of fluorescence and a dramatic reduction in the lifetime of the excited state. By immobilizing the analyte of interest (or a structural analog of the analyte) to a metal surface and exposing it to a labeled receptor (e.g. antibody), the fluorescence of the labeled receptor becomes quenched upon binding because of the close proximity to the metal. Upon exposure to free analyte, the labeled receptor dissociates from the surface and diffuses into the bulk of the solution. This increases its separation from the metal and an increase of fluorescence intensity and/or lifetime of the excited state is observed that indicates the presence of the soluble analyte. By enclosing this system within a small volume with a semipermeable membrane, a reversible device is obtained. We demonstrate this scheme using a biotinylated self-assembled monolayer (SAM) on gold as our surface immobilized analyte analog, fluorescently labeled anti-biotin as a receptor, and a solution of biotin in PBS as a model analyte. This scheme could easily be extended to transduce a wide variety of protein-ligand interactions and other biorecognition phenomena (e.g. DNA hybridization) that result in changes in the architecture of surface immobilized biomolecules such that a change in the separation distance between fluorophores and the metal film is obtained.

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A glucose biosensor based on an amperometric oxygen electrode has been developed. Polycarbonate and Silastic membranes were assembled (glued together) to form a multilayer sandwich glucose diffusion barrier. The effects of the glue layer composition and thickness of the Silastic membrane on sensor response parameters have been investigated in order to optimize the sensor. The parameters measured were the sensitivity, the concentration range of the linear dependence of the sensor response to glucose, and the long-term operation time. The sensors with the sandwich-type glucose diffusion membrane (Silastic membrane prepared from 20% Silastic suspension, glue layer prepared from polyurethane, 0.5 w/v % in ThF solution and standard polycarbonate membrane) demonstrated linearity of response up to 520 mg/dl glucose at 25 degrees C and up to 400 mg/dl at 37 degrees C. These sensor showed good reproducibility of response without significant interference effects (from 1 to 5% of the background current value). The long-term continuous operational time of the sensors was over 40 days at 37 degrees C, and over 60 days at 25 degrees C.

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