Enzyme biosensors covalent bonding

Pn enzyme electrode may be envisaged as a self-contained analytical biosensor comprising a thin enzyme layer overlying, for example, a carbon or plat intro anode. The enzymes need to be immobilized and this is best realised by direct covalent bonding to an insoluble matrix rather than by physical means. Several viable polymers are suitable for this purpose. 

When constructing biosensors, which are to be used continuously in vivo or in situ, maintaining sensor efficiency while increasing sensor lifetime are major issues to be addressed. Researchers have attempted various methods to prevent enzyme inactivation and maintain a high density of redox mediators at the sensor surface. Use of hydrogels, sol-gel systems, PEI and carbon paste matrices to stabilize enzymes and redox polymers was mentioned in previous sections. Another alternative is to use conductive polymers such as polypyrrole [123-127], polythiophene [78,79] or polyaniline [128] to immobilize enzymes and mediators through either covalent bonding or entrapment in the polymer matrix. Application to various enzyme biosensors has been tested. 

Proteins such as antibodies and enzymes can be deliberately anchored on microfluidic device surfaces by covalent bonds or molecular recognition in order to fabricate array biosensors. Nraispecific adsorption is the (usually) undesirable adsorption of molecules on the surface, and it ends up with... 

Proteins such as antibodies and enzymes can be deliberately anchored on microfiuidic device surfaces by covalent bonds or molecular recognition in order to fabricate aiTay biosensors. Nonspecific adsorption is the (usually) undesirable adsorption of molecules on the surface, and it ends up with loss of analyte. In nonspecific adsorption, the molecule-surface interactions are initially weaker, so the adsorption process is both slower and more difficult to control. Subsequent denaturation leads to stronger adhesion [7], as well as to changes in the surface hydrophilicity and roughness [8]. It is important to understand the mech-... 

Technologies for immobilizing proteins on surfaces that have been developed over the last 25 years are particularly critical to the continued development of biosensors. There a dozens of techniques for activating surfaces so as to be able to covalently bond proteins while maintaining their biological activity. These methods have been used routinely for manufacturing adsorbents for affinity chromatography, dip-stick type analytical methods, e.g., immunoassays, and immobilized enzymes for commercial catalysts. 

In order to make a useful biosensor, enzyme has to be properly attached to the transducer with maintained enzyme activity. This process is known as enzyme immobilization. The choice of immobilization method depends on many factors such as the nature of the enzyme, the type of transducer used, the physiochemical properties of analyte, and the operating conditions [73]. The major requirement out of all these is its maximum activity in immobilized microenvironment. Enzyme-based electrodes provide a tool to combine selectivity of enzyme toward particular analyte and the analytical power of electrochemical devices. The amperometric transducers are highly compatible when enzymes such as urease, generating electro-oxidizable ions, are used [74]. The effective fabrication of enzyme biosensor based on how well the enzyme bounds to the transducer surface and remains there during use. The enzyme molecules dispersed in solutions will have a freedom of their movement randomly. Enzyme immobilization is a technique that prohibits this freedom of movement of enzyme molecules. There are four basic methods of immobilizing enzymes on support materials [75] and they are physical adsorption, entrapment, covalent bonding, and cross-linking, as shown in the Fig. 36. 

Chitosan films can also be readily functionalized with proteins, enzymes, antibodies, and DNA, for selective coatings for biosensors. Different methods have been demonstrated for chitosan modification physical interaction (eg, surface absorption, entrapment), chemical bonding, or covalent crosslink with each other (Koev et al., 2010). 

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