Biosensors absorption-based fluorescence

While planar optical sensors exist in various forms, the focus of this chapter has been on planar waveguide-based platforms that employ evanescent wave effects as the basis for sensing. The advantages of evanescent wave interrogation of thin film optical sensors have been discussed for both optical absorption and fluorescence-based sensors. These include the ability to increase device sensitivity without adversely affecting response time in the case of absorption-based platforms and the surface-specific excitation of fluorescence for optical biosensors, the latter being made possible by the tuneable nature of the evanescent field penetration depth. 

As opposed to conventional analytical techniques, optical sensors and biosensors, particularly those employing absorption and fluorescence-based sensing materials potentially allow for measurement through transparent or semi-transparent materials in a non-destructive fashion4, 5> 9 10. Chemical sensor technology has developed rapidly over the past years and a number of systems for food applications have been introduced and evaluated with foods. 

Fiber-optic biosensors are analytical devices in which a fiber optic device serves as a transduction element. The usual aim of fiber-optic biosensors is to produce a signal proportional to the concentration of target analyte to which the biological element reacts. Fiber-optic biosensors are based on the transmission of light along silica glass fiber, or POF to the site of analysis. They can be used in combination with different types of spectroscopic technique, e.g. absorption, fluorescence, phosphorescence, or surface plasmon resonance (SPR) (14). 

The majority of the biosensors that have been reported are based on the deposition of biologically active species such as enzymes and antibodies at the surface of an electrochemical or optical transducer. The most common principle is to identify the analyte by use of a chemically selective enzyme. The enzyme-substrate reaction produces a secondary chemical signal by means of catalysis, e.g. H" or H2O2. This signal is then recognized and quantitatively converted to an electrical signal, e.g. a potential, a current, or a change of absorption or fluorescence, by a suitable transducer. 

Most optical detection methods for biosensors are based on ultra-violet (UV) absorption spectrometry, emission spectroscopic measurement of fluorescence and luminescence, and Raman spectroscopy. However, surface plasmon resonance (SPR) has quickly been widely adopted as a nonlabeling technique that provides attractive advantages. Fueled by numerous new nanomateiials, their unique, SPR-based or related detection techniques are increasingly being investigated [28-31]. 

Biosensor in Table 1-8 is a gel entrapping biomolecules, such as proteins, which react with molecules to be analyzed, resulting in a change in spectra of optical absorption or fluorescence (Dave, 1994). Phycobiliprotein-doped gel (Chen, 1996), for instance, presents a sensor based on the change in fluorescence spectrum. Availability of carbon-enzyme hybrid electrodes is discussed with glucose biosensor as a test case (Sampath, 1996). 

Optical nucleic acid biosensors and microarrays based on fluorescence detection make use of fluorescent dyes to provide a measurable signal for target/probe hybridization. Ideally, fluorophores used for detecting nucleic acid hybridization should exhibit large molar absorptivity, resistance to pho-tobleaching, quantum yields that approach unity and the abibty to produce a resolvable signal at low concentrations both quickly and reproducibly [40]. 

The field of applications for optical biosensors is wide, covering clinical, industrial control processes, veterinary, food, environmental monitoring, among others [1]. For all these applications, it is desirable to have a compact sensor of high sensitivity, fast response time and which is able to perform real-time measurements. These requirements can be achieved mainly with optical sensors, due to the intrinsic nature of optical measurements that accommodate a great number of different techniques based on emission, absorption, fluorescence, refractometry or polarimetry. 

Biosenscx are usually based on extrinsic sensors. The biocatalyst is immobilized on the tip of an optical fiber, and the biosensor measures the absorption, fluorescence, or chemi/bioluminescence produced. 

The creation of an optical sensor that detects the cofactors on which many enzymes dqiend is a much massociated with many enzymes, particularly dehydrogenases. This cofactor has a maximal absorption at 340 nm and a maximal fluorescence emission at 400 nm, which is easily detectable with a photomultiplier. NADH also has the advantage that it can be immobilized on the same support as the enzyme (see 3.3.1.e). nber-optic biosensors based on the fluorimetric detection of NADH have been constructed for the determination of lactate and pyruvate [208]. These sensors use inunobilized lactate dehydrogenase AD) to catalyse the following equilibrium reaction ... 

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