Immobilization techniques

The next generation of amperomethc enzyme electrodes may weU be based on immobilization techniques that are compatible with microelectronic mass-production processes and are easy to miniaturize (42). Integration of enzymes and mediators simultaneously should improve the electron-transfer pathway from the active site of the enzyme to the electrode. 

Choice of Method. Numerous enzyme immobilization techniques have been described in the Hterature comprehensive books on this and related subjects, including industrial appHcations, are available (33—36). The more general techniques and some selection criteria are included herein. 

Dispersion Characteristics The chief characteristics of gas-in-liquid dispersions, like those of hquid-in-gas suspensions, are heterogeneity and instabihty. The composition and structure of an unstable dispersion must be obsei ved in the dynamic situation by looking at the mixture, with or without the aid of optical devices, or by photographing it, preferably in nominal steady state photographs usually are required for quantitative treatment. Stable foams may be examined after the fact of their creation if they are sufficiently robust or if an immobilizing technique such as freezing is employed [Chang et al., Ind. Eng Chem., 48, 2035 (1956)]. 

The combination of ionic liquids and compressed CO2 - at opposite extremes of the volatility and polarity scales - offers a new and intriguing immobilization technique for homogeneous catalysis. 

All these immobilization techniques run the risk of altering activity compared with the native enzyme. Improved activity is occasionally reported, but this is the exception. The immobilization techniques listed above are in approximate order of loss in activity. Physical entrapment normally causes no change. Adsorption will distort the shape of the molecule compared with the native... 

Lichlyter D, Haidekker MA (2009) Immobilization techniques for molecular rotors -towards a solid-state viscosity sensor platform. Sens Actuators B Chem 139 648-656... 

Wu, K. Y. A., and Wisecarver, K. D., Biological Phenol Degradation in a Countercurrent Three-Phase Fluidized Bed Using a Novel Cell Immobilization Technique, AIChE Symp. Series, 86(276) 113 (1989)... 

In view of the conductive and electrocatalytic features of carbon nanotubes (CNTs), AChE and choline oxidases (COx) have been covalently coimmobilized on multiwall carbon nanotubes (MWNTs) for the preparation of an organophosphorus pesticide (OP) biosensor [40, 41], Another OP biosensor has also been constructed by adsorption of AChE on MWNTs modified thick film [8], More recently AChE has been covalently linked with MWNTs doped glutaraldehyde cross-linked chitosan composite film [11], in which biopolymer chitosan provides biocompatible nature to the enzyme and MWNTs improve the conductive nature of chitosan. Even though these enzyme immobilization techniques have been reported in the last three decades, no method can be commonly used for all the enzymes by retaining their complete activity. 

W.H. Scouten, J.H. Luong, and R.S. Brown, Enzyme or protein immobilization techniques for applications in biosensor design. Trends Biotechnol. 13,178—185 (1995). 

Katzbauer B, Narodoslawsky B, Moser A (1995) Classification system for immobilization techniques. Bioprocess Eng 12 173-179... 

Advances have been achieved in recent years, such as the use of CL reagents as labels to derivatize and sensitively determine analytes containing amine, carboxyl, hydroxy, thiol, and other functional groups and their application in HPLC and CE [35, 36], the synthesis and application of new acridinium esters [37], the development of enhanced CL detection of horseradish peroxidase (HRP) labels [38], the use of immobilization techniques for developing CL-based sensors [39-42], some developments of luminol-based CL in relation to its application to time-resolved or solid-surface analysis [43], and the analytical application of electrogenerated CL (ECL) [44-47], among others. 

By immobilizing Mn(III)-tetrakis(4-sulfonatophenyl)-porphyrin on dioctadecyl-dimethyl ammonium chloride bilayer membranes incorporated into a PVC film, Kuniyoshi et al. [65] developed an epinephrine CL sensor, which allowed determination of epinephrine down to 3 pM with an RSD of 1.0% for 50 pM of this biological compound. Compared with the previously reported epinephrine CL sensor [66], the present authors noted that the alkaline carrier solution, at high concentration levels, caused gradual deterioration of the immobilized catalyst, and this problem could be solved by the use of immobilization techniques other than ion exchange, e.g., solubilization of the catalyst that has octadecyl groups in the bilayer molecules. 

Immobilization techniques have been applied in the preparation of immobilized CL reagents, with specific advantages such as reusability, improved stability, and increased efficiency. These strategies have been applied in the development of CL sensors, which today constitute the most important tools in analytical chemistry because of the high sensitivity offered. Optical fibers have been used to transfer light in order to improve the quality of detection, and new types of flow-through cells have been introduced in the construction of CL sensors. Also, selectivity has been considerably improved by the utilization of enzymatic or antigen-antibody reactions. 

Immobilized cells are widely used for both practical and academic purposes. Immobilization techniques have been developed for cell stabilization and easy operation. The methods for immobilizing of the whole cells may be categorized as follows ... 

Finally - and perhaps most importantly - the fluorous tagging of the catalyst that introduces affinity for the fluorous phase can be a very mild immobilization technique, as there is no direct covalent link with a support and the sepa-... 

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