Biosensor devices enzyme electrode

Since the report of carbon nanotubes (CNT) in 1991, much attention has been focused on their unique electrical and mechanical properties. The integration of bioactive molecules (enzymes, proteins, antigens, antibodies, DNA, etc.) with CNT enables the use of the hybrid systems as biosensor devices (enzyme electrodes, immunosensors, or DNA sensors), but the hydrophobicity of the CNT limits their application in biology. Thus, the functionalization of CNT with biopolymers promises to be one of the most successful methods to improve the hydrophilicity of CNT. These biopolymers can bring a hydrophilic surface of the CNT for covalent, absorptive, or ionic bindings with bioactive molecules. 

Enzyme sensors are based primarily on the immobilization of an enzyme onto an electrode, either a metallic electrode used in amperometry (e.g., detection of the enzyme-catalyzed oxidation of glucose) or an ISE employed in potentiometry (e.g., detection of the enzyme-catalyzed liberation of hydronium or ammonium ions). The first potentiometric enzyme electrode, which appeared in 1969 due to Guilbault and Montalvo [140], was a probe for urea with immobilized urease on a glass electrode. Hill and co-workers [141] described in 1986 the second-generation biosensor using ferrocene as a mediator. This device was later marketed as the glucose pen . The development of enzyme-based sensors for the detection of glucose in blood represents a major area of biosensor research. 

Besides the broad applications of electrically contacted enzyme electrodes as amperometric biosensors, substantial recent research efforts are directed to the integration of these functional electrodes as biofuel cell devices. The biofuel cell consists of an electrically contacted enzyme electrode acting as anode, where the oxidation of the fuel occurs, and an electrically wired cathode, where the biocatalyzed reduction of the oxidizer proceeds (Fig. 12.4a). The biocatalytic transformations occurring at the anode and the cathode lead to the oxidation of the fuel substrate and the reduction of the oxidizer, with the concomitant generation of a current through the external circuit. Such biofuel cells can, in principle, transform chemical energy stored in biomass into electrical energy. Also, the use... 

Types of biosensors can be named either by the biological components, physical transducing devices, or the measured analytes. Researchers were originally using biological components to define types of biosensors (Table 2). Types of transducers had also been included in the name to identify the physical transducing device, i.e., enzyme electrodes, acoustic-immunosensors, optical biosensors, piezoelectric-immunosensors, and biochips. 

Redox enzymes are the active component in many electrochemical enzyme electrode biosensor devices.1821 The integration of two different redox enzymes with an electrode support, in which one of the biocatalysts is photoswitchable between ON and OFF states, can establish a composite multisensor array. The biomaterial interface that includes the photoswitchable enzyme in the OFF state electrochemi-cally transduces the sensing event of the substrate corresponding to the nonphoto-switchable enzyme. Photochemical activation of the light-active enzyme leads to the full electrochemical response, corresponding to the analysis of the substrates of the two enzymes. As a result, the processing of the signals transduced by the composite biomaterial interface in the presence of the two substrates permits the assay of the... 

The hydrophobias are a case where protein nanofibers can play a dual role in creating a biosensor. They can aid in the immobilization of bioactive components within a biosensor and also add further functionality to the transducing element of a biosensor device. Hydrophobins are self-assembling [3-sheet structures observed on the hyphae of filamentous fungi. They are surface active and aid the adhesion of hyphae to hydrophobic surfaces (Corvis et al., 2005). These properties can be used to create hydrophobia layers on glass electrodes. These layers can then facilitate the adsorption of two model enzymes glucose oxidase (GOX) and hydrogen peroxidase (HRP) to the electrode surface. The hydrophobin layer also enhances the electrochemical properties of the electrodes. 

In the field of biosensor technology, immobilized enzyme electrode development occupies a place of prominence due to the attractive performance of this hybrid device. Coupling an immobilized enzyme layer with an electrochemical sensor combines the advantages of using an insolubilized enzyme system (see below) with the sensitivity of readily available potentiometric and amperometric electrodes. The resulting biosensor enables direct, reliable, and reproducible... 

Several implanted biosensors have been developed and evaluated in both animals and humans (see Chapter 4). Detection systems are based on enzymes, electrodes, or fluorescence. The most widely studied method is an electrochemical sensor that uses glucose oxidase. This sensor can be implanted intravenously or subcutaneously. Intravenous implantation in dogs for up to 3 months has demonstrated the feasibility of this approach. Alternatives to enzymes are being developed, including artificial glucose receptors. Less success has been achieved with subcutaneous implants. Implantation of a needle type of sensor into the subcutaneous tissue induces a host of inflammatory responses that alters the sensitivity of the device. Microdialysis with hoUow fibers or ultrafiltration with biologically inert material can decrease this problem. 

Garry A. Rechnitz together with S. Katz introduced one of the first papers in the field of biosensors with the direct potenfiometric determination of urea after urease hydrolysis. At that fime the term biosensor had not yet been coined. Thus, these types of devices were called enzyme electrodes or biocatalyfic membrane electrodes [100[. 

The need to improve the electrical communication between redox proteins and electrodes, and the understanding that the structural orientation at the molecular level of redox proteins and electroactive relay units on the conductive surfaces is a key element to facilitate ET, introduced tremendous research efforts to nano-engineer enzyme electrodes with improved ET functionalities. The present chapter addresses recent advances in the assembly of structurally aligned enzyme layers on electrodes by means of surface reconstitution and surface crosslinking of structurally oriented enzyme/cofactor complexes on electrodes. The ET properties of the nano-structured interfaces is discussed, as well as the possible application of the systems in bioelectronic devices such as biosensors, biofuel cell elements or optical and electrical switches. 

Clearly, electrochemical indication prevails over all other methods of transduction. Potentiometric and amperometric enzyme electrodes are at the leading edge of biosensor technology with respect to the body of scientific literature as well as to commercially available devices (Schindler and Schindler, 1983). Only a few conductometric biosensors have been described, but the relevance of this sensor type may increase because of the relative ease of their preparation and use. Furthermore, the development of biochemically sensitized field effect transistors, being at present only at an initial stage, offers new prospects (Pinkerton and Lawson, 1982). 

In this chapter, we focus on the use of the continuous resonance QCM and EQCM devices to study enzymatic polymerization reactions, biochemical and biomimetic processes and to create thin polymer films electrochemically on the QCM electrode surface. We describe some QCM applications of thin polymer films in enzyme electrode and other biosensors and briefly describe specific cell binding systems to create whole cell biosensors. 

Enzymes-based biosensors are well reported in the literature for chemical toxicity screening. The sensor devices produced using enzymes are usually simple and easy to fabricate, inexpensive, and sensitive to low levels of toxicants. Immobilization of enzymes on the electrode surface can include adsorption, covalent attachment, or film deposition using a range of procedures [68-70]. The sensor system relies primarily on two enzyme mechanisms catalytic transformation of a pollutant and detection of pollutants that inhibit or mediate the enzyme s activity. In catalytic enzyme biosensor, the enzyme specific for the substrate of interest (toxin in this case)... 

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