Low-Cost Microdevices for Point-of-Care Testing - Point-of-Care Diagnostics on a Chip - page 13

cycling [ 74 ]. Both methods have low power consumption and are amenable for use
at the point-of-care.
By contrast, isothermal techniques have been developed to perform nucleic
acid amplification without thermal cycling. This feature removes the need of
using different temperatures by making use of enzymes to perform reaction at a
single temperature which translates into less complex and cheaper instrumentation
that is amenable for point-of-care use [ 25 , 69 ]. Current commercially available
platforms [ 25 ], for example, helicase-dependent amplification (HDA; BioHelix),
transcription-mediated amplification (TMA; Gen-Probe), nucleic acid sequence-
based amplification (NASBA; BioMerieux), strand displacement amplification
(SDA; Becton Dickinson), loop-mediated amplification (LAMP; Eiken), and recom-
binase polymerase amplification (RPA; TwistDx) are implemented in laboratory-
based NAT and microdevices for point-of-care.

1.4.3.3 Product Detection

Amplified product can be examined at the end of reaction (end-point detection)
or during the reaction (real-time detection). There are many techniques to detect
nucleic acids, which one of the primary methods is quantifying nucleic acids
by ultraviolet light (260 nm) absorption. Nonetheless, fluorescence-based tech-
niques remain the most commonly employed due to its high level of sensitivity
and low background noise [ 64 ]. Fluorescent dyes can either bind to nonspecific
locations (general interactions) or specific locations of molecule, depending on
the application. Cepheid Inc., for example, has developed a real-time fluorescent
PCR detection (fluorescently labeled probes), requiring instruments that may be
used in some but not all point-of-care settings. Cepheid's GeneXpert test platform
(Fig. 1.5 a) has been tested for clinical trials in four developing countries and showed
promising results for detecting tuberculosis [ 69 ]. However, this system requires
uninterrupted and stable electrical power supply and annual validation of the system,
and generates considerable more waste than microscopy technique [ 75 ]whichmay
lead to problems of waste management in resource-limited settings.
At the point-of-care, electrochemical methods may also be suitable due to their
compatibility with low-cost and portable analyzers [ 60 ]. For example, Nanosphere
is building a scanner-based detector to detect DNA via nanoparticle probes [ 60 ].
Gold nanoparticles functionalized with oligonucleotides are used as probes for DNA
sequences complementary to the sequences of those oligonucleotides (Fig. 1.5 b).
Visualization of the gold nanoparticle, and hence DNA content, is performed using
a signal amplification method in which silver is reduced at the surface of the gold,
and a scanner is used to measure amount of light scattered.
Lateral flow devices for end-point detection have been used for simple and
low-cost method. A DNA strip using specific probes functionalized on membrane
to capture target sequences from amplification reaction can be used for identi-
fication of target strains. GenoType (Hain Lifescience GmbH) and INNO-LiPA
(Innogenetics NV) have developed assays to detect infectious diseases. BioHelix
has developed a strip cassette for the isothermal amplification assay (Fig. 1.6 ).