Biomedical Engineering Reference
In-Depth Information
oxidoreductase, HRP, and the hydrolytic enzyme AP are frequently used because of
their ability to yield an electroactive product following the catalytic conversion of a
substrate. The magnitude of the current arising from the redox reaction of the product
can then be quantitatively related to the amount of analyte present. This signal-generation
scheme is further illustrated by the example of the enzymatic reaction of AP on its sub-
strate 4-aminophenyl phosphate (4-APP) to yield 4-aminophenol (4-AP) in Scheme 1. This
is followed by the oxidation of 4-AP to 4-quinone imine (4-QI) to produce a Faradaic
current that is proportional to the amount of analyte. Notably, a major advantage of
enzyme labels is, as a result of the catalytic effect, the amplifi cation of the signal that
can be detected even when a minute quantity of enzyme is used.
O
O
O
O
P
O
200 mV
vs Ag/AgCI
2H 2e
O
OH
O
AP
NH 2
NH 2
NH
SCHEME 1
Due to the broad substrate specifi city of AP, and the drive for higher effi ciency, several
studies have recently investigated the suitability of alternative substrates to the com-
mon 4-APP [47, 48]. For example, Pemberton et al. compared 4-APP and 1-naphthyl
phosphate (1-NP) as AP substrates in an amperometric immunosensor for progester-
one [47]. The signal generation scheme when 1-NP is used as a substrate is illustrated
in Scheme 2.
O
O
O
O
P
270 mV
vs Ag/ AgCI
H e
O
OH
O
AP
SCHEME 2
They found that the hydrolysis products of 4-AP and 1-naphthol produced well-
defi ned anodic responses at low potentials at a bare SPCE. However, the presence of
antibody immobilized on the electrode surface slowed the diffusion of 4-AP towards
the electrode surface. In addition, 4-AP may interact with polyphenols on the electrode
surface, thus reducing the electroactive working area of the electrode by fouling. In
contrast, diffusion of 1-naphthol to the electrode surface was not hindered by immobi-
lized antibody. This feature, along with its low cost, ease of availability, and high solu-
bility, resulted in 1-NP being the preferred AP substrate in their work.
 
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