Biomedical Engineering Reference
In-Depth Information
The use of aptamer-based biosensors would offer a number of advantages in this
respect. Indeed, the function of immobilized aptamers could be regenerated using
extremes of heat, salt concentration, or chelating agents, without loss of activity, result-
ing in a reusable transducer (52). Aptamers can also be easily modified for immobi-
lization without affecting their function (52). Furthermore, they can be selected for
under conditions resembling those of the real matrix, which is particularly useful for
environmental and food applications (51). Lastly, aptamers can be easily labeled with
a wide range of reporter molecules, allowing the design of a variety of detection
formats (52).
Recent work on aptamers in affinity sensors includes the use of an RNA aptamer spe-
cific for
L
-adenosine (63) and a DNA aptamer for human thrombin (64). Results in both
cases were promising, with the aptamers showing high specificity and the ability to be
regenerated (52).
20.3.4.4 Molecular Beacons
Molecular beacons are a class of fluorogenic probe in which a fluorophore is attached
to one terminus and a quencher is attached to the other. In the absence of the target
molecule, the probe forms a hairpin loop, bringing the fluorophore and the quencher
into close proximity, resulting in the absence of a fluorescent signal. In the presence of
a target molecule, however, the hairpin loop structure is opened (the target is
complementary to the loop region), the fluorophore and quencher are distanced from
each other, and a fluorescent signal is produced, as illustrated in Figure 20.7. Until
recently, these probes have only been available for the detection of nucleic acid targets,
due to the inability of these probes to interact with other classes of targets, such as
proteins (52).
Aptamers, however, have the ability to interact with a very wide range of targets and
have, therefore, been exploited to extend this technique to include targets other than
nucleic acids. For this purpose, molecular beacons, termed ligand beacons when used in
this way, are designed to be complementary to a nucleotide region in the aptamer. In the
absence of the target (Figure 20.8A), the ligand beacon binds the aptamer, resulting in the
separation of the fluorophore and the quencher and the production of a fluorescent signal.
In the presence of the target (Figure 20.8B), however, the binding site on the aptamer is no
longer available to the ligand beacon because it is bound by the target. This results in the
ligand beacon adopting an internal hairpin loop structure and in the absence of a fluores-
cent signal (52).
+
Molecular
beacon
Target
Hybrid
FIGURE 20.7
Functioning of a molecular beacon.