Total internal reflection fluorescence (TIRF) is an optical effect particularly well-suited to the study of molecular and cellular phenomena at liquid/solid interfaces. Such interfaces are central to a wide range of biochemical and biophysical processes: binding to and triggering of ceils by hormones, neurotransmitters, and antigens; blood coagulation at foreign surfaces; electron transport in the mitochondrial membrane; adherence and mo­bility of bacteria, algae, and cultured animal cells to surfaces; and possible enhancement of reaction rates with cell surface receptors upon nonspecific adsorption and surface diffusion of an agonist. Liquid/solid interfaces also have important medical and industrial applications: e.g. detection of serum antibodies by surface immobilized antigens; and the manufacture of biochemical products by surface-immobilized enzymes. Total internal reflection spectroscopy for optical absorption studies (called attenuated total reflection or ATR) was developed somewhat earlier than the fluorescence applications and has been widely used in surface

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Total internal reflection fluorescence

A specifically-reactive sample-collecting and testing device possessing a cavity or cavities each having a dimension small enough to enable sample liquid to be drawn into the cavity by capillary action, wherein said cavity includes an electrode structure for making measurements of one or more electrically measurable characteristics of the sample, and wherein a surface of a wall of the cavity optionally also carries a coating of a material appropriate to the test to be carried out in the device.

Devices for use in chemical test procedures

Immunoassays are now very widely used in the clinical laboratory, either because no other type of assay system is feasible or because they are often the most effective and suitable of the possible analytical methods. The last decade has seen the development and refinement of many new immunoassay reagents and systems. The major trend has been away from liquid-phase assays involving radioisotopic labels, towards fast homogeneous or solid-phase assays capable of operation anywhere; and towards precise and reliable nonisotopic, automated or semi-automated laboratory assays, often with detection limits measured in pico- or attomoles. The use of monoclonal antibodies is now widespread, and the methodologies of labels and of solid-phase components are much more sophisticated. New assay formulations, novel homogeneous systems, immunosensors, free-analyte assays, the importance of thorough validation and of interfering substances, and future trends are discussed.

A Decade of Development in Immunoassay Methodology

Biosensors incorporate a biological sensing element that converts a change in an immediate environment to signals conducive for processing. Biosensors have been implemented for a number of applications ranging from environmental pollutant detection to defense monitoring. Biosensors have two intriguing characteristics: (1) they have a naturally evolved selectivity to biological or biologically active analytes; and (2) biosensors have the capacity to respond to analytes in a physiologically relevant manner. In this paper, molecular biosensors, based on antibodies, enzymes, ion channels, or nucleic acids, are briefly reviewed. Moreover, cell-based biosensors are reviewed and discussed. Cell-based biosensors have been implemented using microorganisms, particularly for environmental monitoring of pollutants. Biosensors incorporating mammalian cells have a distinct advantage of responding in a manner that can offer insight into the physiological effect of an analyte. Several approaches for transduction of cellular signals are discussed: these approaches include measures of cell metabolism, impedance, intracellular potentials, and extracellular potentials. Among these approaches, networks of excitable cells cultured on microelectrode arrays are uniquely poised to provide rapid, functional classification of an analyte and ultimately constitute a potentially effective cell-based biosensor technology. Three challenges that constitute barriers to increased cell-based biosensor applications are presented: analytical methods, reproducibility, and cell sources. Possible future solutions to these challenges are discussed.

Development and application of cell-based biosensors.

Abstract The Resonant Mirror biosensor is a new design of optical sensor, aimed at combining the improved sensitivity of waveguide sensors with the simplicity of fabrication and ease of use associated with surface plasmon sensors. It uses the evanescent wave associated with a dielectric resonant structure to probe reactions occurring in a sensing layer, deposited within a few hundred nanometers of the device surface and is aimed at the real time monitoring of biological assays, without use of labelled molecules. This paper describes the operation of the device and performance features relevant to its use as a biosensor.

The Resonant Mirror: a novel optical biosensor for direct sensing of biomolecular interactions

An analyte in solution is made to react with a spectific reactant coated on the wave-guide thus modifying the optical properties thereof. The index of refraction of the wave-guide material is higher than that of the reaction medium which ensures that a light signal injected into said guide be carried by multiple total reflection, the distance of penetration of the evanescent wave associated with the totally reflected signal being of the same order of magnitude or greater than the thickness of the analyte-reactant product layer.

Method for the determination of species in solution with an optical wave-guide

Opto-electronic immunosensors: a review of optical immunoassay at continuous surfaces.

Immunoassays at a quartz-liquid interface: Theory, instrumentation and preliminary application to the fluorescent immunoassay of human immunoglobulin G

An analyte in solution is made to react with a specific reactant coated on the wave-guide thus modifying the optical properties thereof. The index of refraction of the wave-guide material is higher than that of the reaction medium which ensures that a light signal injected into said guide be carried by multiple total reflection, the distance of penetration of the evanescent wave associated with the totally reflected signal being of the same order of magnitude or greater than the thickness of the analyte-reactant product layer.

Method and apparatus for the determination of species in solution with an optical wave-guide

A novel optical immunoassay system was developed and tested using human IgG as a model antigen. Following adsorption of antiserum to the surface of an optical waveguide, the immobilised antibody was then reacted with a solution containing antigen. The reaction was detected utilising the evanescent wave component of a light beam totally internally reflected within the waveguide. The growing antigeh-antibody layer resulted in an increase of scattered light which was monitored kinetically. Individual measurements were completed within 5 minutes and the resultant dose response curve had a sensitivity limit of approximately 10 nmol/L.

John F. Place

Papers

Method for the determination of species in solution with an optical wave-guide

Opto-electronic immunosensors: a review of optical immunoassay at continuous surfaces.

Immunoassays at a quartz-liquid interface: Theory, instrumentation and preliminary application to the fluorescent immunoassay of human immunoglobulin G

Method and apparatus for the determination of species in solution with an optical wave-guide

Preliminary results obtained with a No-Label, Homogeneous, Optical Immunoassay for Human Immunoglobulin G