Biomedical Engineering Reference
In-Depth Information
4.7.1 Optical Detection Methods
4.7.1.1 Absorption and Emission Methods
Conventional colorimetric methods have been in existence for a long time. These
have been used for DNA, proteins, bacteria, virus, cells, and other biomolecules
(REFS). These have been coupled with polymerase chain reaction (PCR), high-
performance liquid chromatography, ELISA, and other assays (REFS). Colori-
metric assays coupled with molecular fluorophore assays offer high sensitivity of
detection, but they suffer from several drawbacks that include complex handling
procedures, easy contamination, high cost, and lack of portability.
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In this manner, the use of NMs along with simple detection alternatives such as
those based on light absorption measurement is very attractive. One of the most
exploited NM for optical absorption detection is the AuNP. It has been used for
various analytes including pregnancy tests, HIV, and other clinical applications.
A biosensor based upon AuNPs that were chemically modified with 5′and
3′ (alkanethiol) capped ONTs that exhibited extraordinary selectivity for DNA
detection was reported.
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This NP-based simple colorimetric detection of a tar-
get ONT in the presence of a mixture of ONTs that differed by one nucleotide in
the target region. The same AuNPs were also used by the same group for a rapid,
highly selective, and sensitive colorimetric assay for detecting cysteine.
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4.7.1.2 Fluorescence
Fluorescence-based medical biosensors involving various types of NMs have
also dominated the literature. QDs are extensively used in fluorescent biosensors
eventhough they are toxic. QDs, which are typically synthesized as hydrophobic
molecules, are surface modified to enable hydrophilicity and biocompatibility.
The strategy developed for enabling biocompatibility is to modify the surface
of QDs with more biologically friendly coatings such as 3-mercaptopropanoic
acid, dihydrolipoic acid, HS-poly(ethylene glycol)-carboxyl acid (HS-PEG-
COOH), dextran, chitosan, or other hydrophilic and bifunctional ligands.
A QD-based fluorescence whole-cell imaging sensor that offer rapid, repro-
ducible, accurate, and long-term cell staining system was recently reported.
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The
QD-based imaging sensor involved capture of whole cells with highly specific anti-
bodies against the cell membrane proteins. The capture antibodies were covalently
immobilized on silanized glass slides. The whole cells were exposed to a second
antibody against EpCAM proteins that are found on the surface of the cells result-
ing in a sandwich-type assay. To complete the sensor, QD-labeled antibodies were
exposed to the assay complex. QDs emitting at 620 nm giving a red color rendered
the cells as bright red circular objects under UV-illuminated microscope (
Figure
4.7
). The results of the whole-cell sensor indicated that the QD-based imaging
sensor exhibited brighter signals compared with those using the organic dye Texas
red. The QD-based whole-cell imaging sensor was still brightly visible under the
microscope after 1 week. The assay involved only 1 pmol of QD that was used to
detect as low as 5000 cells in an unoptimized system. The whole-cell biosensor
