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Zeptomole-detecting biosensor for alkaline phosphatase in an electrochemical immunoassay for 2,4-dichlorophenoxyacetic acid

Item Type:Article
Title:Zeptomole-detecting biosensor for alkaline phosphatase in an electrochemical immunoassay for 2,4-dichlorophenoxyacetic acid
Creators Name:Bauer, C.G. and Eremenko, A.V. and Ehrentreich-Foerster, E. and Bier, F.F. and Makower, A. and Halsall, H.B. and Heineman, W.R. and Scheller, F.W.
Abstract:A bienzyme substrate-recycling biosensor in a flow injection analysis system is described for the sensitive measurement of alkaline phosphatase (ALP) and applied to the fast readout of a competitive immunoassay for the widely used pesticide 2,4-dichlorophenoxyacetic acid (2,4-D). The phenol-indicating biosensor consists of a Clark-type electrode covered by a membrane with coentrapped tyrosinase and quinoprotein glucose dehydrogenase. ALP dephosphorylates phenyl phosphate to phenol (K(m) = 36 microM) outside the flow system. Phenol is oxidized in the sensor membrane by the oxygen-consuming tyrosinase via catechol to o-quinone. The quinone is reconverted to catechol by glucose dehydrogenase. This substrate cycling results in a 350-fold amplified sensor response to phenol. The oxygen consumption of the enzyme couple in the presence of phenol is monitored as a decrease in current. A total of 3.2 fM ALP (320 zmol/ 100 microL) has been detected after a 57.5 min incubation with phenyl phosphate. All involved reagents are stable over the time of measurement. The sensor does not produce any measurable blank signals. The immunoassay detects 0.1 microgram/L 2,4-D, the maximum concentration for pesticides allowed in drinking water by European Community regulations. The applicability of this biosensor for fast immunoassay readout is demonstrated by a 2 min incubation. By comparison, a standard photometric method (p-nitrophenyl phosphate) requires overnight incubation.
Keywords:2,4-Dichlorophenoxyacetic Acid, Alkaline Phosphatase, Biosensing Techniques, Calibration, Electrochemistry, Herbicides, Immunoassay
Source:Analytical Chemistry
Page Range:2453-2458
Date:1 August 1996
Official Publication:https://doi.org/10.1021/ac960218x
PubMed:View item in PubMed

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