Enzyme Biosensor Assembly

To prepare the biosensor, the enzyme has to be immobilized in a thin layer at the surface 

Physical entrapment within an inert polymeric membrane (in this case the enzyme is mixed with a monomer solution, which is then polymerized to a gel - polyacrylamide gel, starch, agar gel, and so on, thus trapping the enzyme) or behind a membrane (in this case the enzyme solution is simply confined by an analyte permeable membrane, such as a dialysis membrane, as a thin film covering the indicator electrode this method is also called microencapsulation)  

Adsorption is the simplest and fastest way to prepare immobilized enzymes. Adsorption can roughly be divided into two classes physical adsorption and chemical adsorption. Physical adsorption is weak and occurs mainly via van der Waals interactions. Chemical adsorption is stronger and involves the formation of covalent bonds. 

Several enzymes can be immobilized within the same reaction layer to increase the range of possible biosensor analytes, or to provide efficient regeneration of enzyme co-subsfiates, or to improve the biosensor selectivity by decreasing the local concentration of electrochemical interfering substances. 

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A combination of a biosensor assembly such as that described in the previous paragraph and several enzyme reactors allows simultaneous... 

The same problem was also present when other amperometric biosensors, based on different class of enzymes, were assembled. For example, the amperometric detection of nicotinamide adenine dinucleotide (NADH) has been a matter of investigation for many years in the biosensor field [1,2]. 

Figure 18 Schematic of a glucose biosensor assembled from diphenylalanine peptide nanotubes. The enzyme GOX has been cross-linked to these nanotubes, which are further linked to the gold (Au) electrode and immobilized in a polyethyleneimine (PEI) matrix. The nanofibers act in two ways they immobilize the sensing enzyme and enhance the transducer. Reprinted in part with permission from Yemini et al. (2005a) (copyright 2005 American Chemical Society).

X. Luo, G. D. Vidal, A. J. Killard, A. Morrin, and M. R. Sm3dh, Nanocauliflowers A nanostructured polyaniUne-modified screen-printed electrode with a self-assembled polystyrene template and its application in an amperometric enzyme biosensor, Electroanalysis, 19, 876-883 (2007). 

Two examples of this layer-by-layer strategy are illustrated. In the first, electrochemical enzyme biosensors for glucose were prepared. Indium tin oxide (ITO) glass electrodes were modified by self-assembly using an immersion technique with up to three bUayers of polyamidoamine (PAMAM) dendrimers containing gold nanoparticles of diameter 3 nm and poly(vinyl sulfonate) (PVS). The gold... 

Biosensors ai e widely used to the detection of hazardous contaminants in foodstuffs, soil and fresh waters. Due to high sensitivity, simple design, low cost and real-time measurement mode biosensors ai e considered as an alternative to conventional analytical techniques, e.g. GC or HPLC. Although the sensitivity and selectivity of contaminant detection is mainly determined by a biological component, i.e. enzyme or antibodies, the biosensor performance can be efficiently controlled by the optimization of its assembly and working conditions. In this report, the prospects to the improvement of pesticide detection with cholinesterase sensors based on modified screen-printed electrodes are summarized. The following opportunities for the controlled improvement of analytical characteristics of anticholinesterase pesticides ai e discussed ... 

Moreover, it has been demonstrated that CNTs promote the direct electrochemistry of enzymes. Dong and coworkers have reported the direct electrochemistry of microperoxidase 11 (MP-11) using CNT-modified GC electrodes [101] and layer-by-layer self-assembled films of chitosan and CNTs [102], The immobilized MP-11 has retained its bioelectrocatalytic activity for the reduction of H202 and 02, which can be used in biosensors or biofuel cells. The direct electrochemistry of catalase at the CNT-modified gold and GC electrodes has also been reported [103-104], The electron transfer rate involving the heme Fe(III)/Fe(II) redox couple for catalase on the CNT-modified electrode is much faster than that on an unmodified electrode or other... 

Besides catalyzing styrene and benzaldehyde, CYP enzymes play an important role in the metabolism of endogenous compounds as well as in pharmacokinetics and toxicokinetics. Joseph [228] developed a biosensor with human CYP3A4 as a novel drugscreening tool. It was constructed by assembling enzyme films on Au electrodes by alternate adsorption of a layer of CYP3A4 on top of a layer of PDDA. The biosensor was applied to detect verapamil, midazolam, quinidine, and progesterone. 

Several protein assemblies have successfully been fabricated on the solid surfaces sifter the bioinformation transduction. These include the following molecular systems molecularly interfaced redox enzymes on the electrode surfaces, calmodulin / protein hybrides, and ordered antibody array on protein A. These protein assemblies find a wider application in various fields such as biosensors, bioreactors, and intelligent materials. 

M. Badea, A. Curulli and G. Palleschi, Oxidase enzyme immobilisation through electropolymerised films to assemble biosensors for batch and flow injection analysis, Biosens. Bioelectron., 18(5-6) (2003) 689-698. 

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