Genosensors enzymes

Amperometric or voltammetric biosensors typically rely on an enzyme system that catalyt-ically converts electrochemically non-active analytes into products that can be oxidized or reduced at a working electrode. Although these devices are the most commonly reported class of biosensors, they tend to have a small dynamic range due to saturation kinetics of the enzyme, and a large overpotential is required for oxidation of the analyte this may lead to oxidation of interfering compounds as well (e.g., ascorbate in the detection of hydrogen peroxide). In addition to the use in enzyme-based biosensors, amperometric transducers have also been used to measure enzyme-labelled tracers for affinity-based biosensor (mainly immunosensors and genosensors). Enzymes which are commonly used for this purpose include horseradish peroxidase (HRP) [17] and alkaline phosphatase (AP) [18,19,21]. 

A disposable electrochemical enzyme-amplified genosensor was described for specific detection of Salmonella (Del Giallo et al., 2005). A DNA probe specific for Salmonella was immobilized onto screen-printed carbon electrodes and allowed to hybridize with a biotinylated PCR-amplified product of Salmonella. The hybridization reaction was detected using streptavidin conjugated-AP where the enzyme catalyzed the conversion of electroinactive a-naphthyl phosphate to electroactive a-naphthol, which was detected by differential pulse voltammetry. 

Chapters 1 to 5 deal with ionophore-based potentiometric sensors or ion-selective electrodes (ISEs). Chapters 6 to 11 cover voltammetric sensors and biosensors and their various applications. The third section (Chapter 12) is dedicated to gas analysis. Chapters 13 to 17 deal with enzyme based sensors. Chapters 18 to 22 are dedicated to immuno-sensors and genosensors. Chapters 23 to 29 cover thick and thin film based sensors and the final section (Chapters 30 to 38) is focused on novel trends in electrochemical sensor technologies based on electronic tongues, micro and nanotechnologies, nanomaterials, etc. 

Fig. 30.4. Evaluation of 2.9 pmol of IS200 amplicon using a genosensor based on dry-adsorption of the target on GEC transducer 68.2 pmol of biotinylated probe 9.00 /ig of enzyme conjugate. The nonspecific adsorption signal is shown in black (more details in Ref. [2]) (N — 3).

For enzyme-based biosensors the mode of detection is based on the catalytic activity and/or binding capacity. Because of the protein nature of almost all enzymes, the catalytic activity depends on the conformation. Exceptions are catalytic ribonucleic acids called DNA biosensors or genosensors. DNA fragments are used as probes for detecting low concentrations of DNA in large samples (see also Part I, Chapters 2 and 3). Because of the highly diluted DNA concentration, microelectromechanical systems which are able of performing PCRs are employed. 

Similar to the labeling of enzymes with redox mediators, genosensors were developed for sequence-specific DNA detection by making use of the electrochemieal properties of organometallic compounds, mainly ferrocene and its derivatives. " Eleetroehemical genosensors are partieularly attractive because they are highly sensitive and robust, cheap compared to other detection modes sueh as fluoreseence, and they can be easily miniaturized. The potential in this technique has attraeted commereial interest. ... 

M. Giallo, D. Ariksoysal, G. Marrazza, and M. Mascini, Disposable electrochemical enzyme-amplified genosensor for Salmonella bacteria detection. Ana/. Lett. 38(15), 2509-2523 (2005). 

Methods for the Detection and Identification of Microorganism Utilizing Enzyme-Based Genosensors on Screen-Printed Chips... 

Advantages of the Enzyme-Based Electrochemical Genosensors in Detecting Bacteria on Screen-Printed Carbon Chips... 

C. Y. Yean, B. Kamarudin, D. A. Ozkan, L. S. Yin, P. Lalitha, A. Ismail, and M. Ozsoz, Enzyme-linked amperometric electrochemical genosensor assay for the detection of PCR amplicons on a streptavidin-treated screen-printed carbon electrode, Ano/. Chem. 80,2774-2779 (2008). 

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