Page 108 - Academic Press Encyclopedia of Physical Science and Technology 3rd BioTechnology

P1: GNB Final Pages
Encyclopedia of Physical Science and Technology EN005F-954 June 15, 2001 20:48

Fiber-Optic Chemical Sensors 821

a. Biocatalysts as the recognition element in fiber- bioaffinity fiber-optic biosensors are based on transducing
optic biosensor. Enzymes are the most commonly em- antibody–antigen (analyte) interactions into an optical
ployed biocatalysts for fiber-optic biosensor fabrication. signal that is proportional to the antigen concentration.
These proteins selectively catalyze the conversion of a Several detection schemes are employed; the simplest one
substrate to product. In fiber-optic biosensors, enzymes involves the detection of intrinsically fluorescent analytes
are used to catalyze the conversion of a non-optically de- such as polynuclear aromatic hydrocarbons (PAHs).
tectable analyte (the analyte is the enzyme’s substrate) Antibodies are immobilized on the fiber surface and a
into an optically detectable product. The optical signal ob- fluorescence signal is obtained when the analyte (antigen)
tained, for example, absorbance or fluorescence, is propor- is present, as shown in Fig. 16a. A more generalized de-
tional to the product concentration and thereby to the ana- tection scheme, often called a competition assay, is based
lyte’s concentration. The enzymatic reaction products are on competition for the antibody-binding site between
measured either directly, if they are optically detectable, an externally added fluorescent-labeled antigen and the
or indirectly by using indicators as described in Section antigen present in the sample (analyte), as shown in
III.B.1. Indicators are used to measure common reaction Fig. 16b. The antibody is immobilized on the optical fiber
+
products such as H , ammonia, oxygen, carbon dioxide, surface, a known concentration of fluorescent-labeled
and hydrogen peroxide. An example of this approach is the antigen is added, and the fluorescence signal obtained is
glucose biosensor. The enzyme glucose oxidase catalyzes set as the initial signal. To perform an analysis, the same
the oxidation of glucose with oxygen (the substrates) to fluorescent-labeled antigen concentration is mixed with
produce gluconolactone and H 2 O 2 . The glucose concen- a sample containing an unknown antigen concentration.
tration is determined by using an indicator to measure
either the amount of oxygen consumed or the amount of
H 2 O 2 produced using an appropriate indicator,

glucose oxidase
Glucose + O 2 −−−−−−−−→ Gluconic acid + H 2 O 2 .
The use of enzymes as sensing materials for fiber-optic
biosensors is relatively limited since the stability and the
activity of most enzymes significantly decrease when they
are removed from their natural environment inside the
cells, although immobilization techniques may help. To
overcome this stability problem, whole cells may be used
as biocatalysts. Although some of the fiber-optic biosen-
sor specificity is reduced with whole-cells, the enzymes
within the cells function more efficiently because they are
in an optimum environment containing all the necessary
cofactors. Whole-cell biocatalysts are particularly advan-
tageous when the detection is based on a sequence of mul-
tiple enzymatic reactions. These enzymatic cascade reac-
tions are very difficult and complicated to accomplish by
coimmobilizing the enzymes but are relatively straightfor-
ward by employing whole cells. The methods for detecting
the products in cell-based fiber-optic biosensors are simi-
lar to those employed in enzyme fiber-optic biosensors.

b. Bioaffinity as the recognition mechanism in
fiber-optic biosensors. Antibodies, receptors, and
nucleic acids are highly selective and can recognize their
binding partners. When the affinity binding reaction is in
equilibrium and readily reversible, a sensor results. If the FIGURE 16 Schematic principles of bioaffinity fiber-optic biosen-
sors. (a) Detection of intrinsically fluorescent molecule using im-
reaction is essentially irreversible, the resulting sensor
mobilized antibody. (b) Competition assay using a fluorescent-
is called a probe, as it must be regenerated or recharged labeled antigen. (c) Sandwich immunoassay using an immobilized
before it can be used to make another measurement. Most antibody and a fluorescent-labeled antibody.