3 Aptamer-Based Point of Care Testing Schemes
59
Humanosteopontin (OPN) protein, a potential cancer biomarker, has substantial
promise for POCT. Biotinylated aptamer was recently used to precapture OPN from
samples, an OPN antibody was immobilized on the test line for second specific target
recognition, and streptavidin-modified GNPs sprayed on the color detection pad. This
achieved a sensitivity of as low as 0.1 ng. mL
−1 OPN with strong dynamic detection
between 10 and 500 ng. mL
−1 within 5 min. The LFA allowed the qualitative and
semiquantitative detection of OPN in serum at standard levels, as in cancer patients,
and can discriminate against OPN from high-specific protein interference [39].
Another example of aptamer-based competitive LFA includes one step determination of ochratoxin A (OTA) in corn samples. Briefly, biotin-cDNA was immobilized on a nitrocellulose filter on the test line. Without OTA, Cy5-labeled aptamer
combined with complementary strands formed a stable double helix. In the presence
of OTA, the Cy5-aptamer/OTA complexes were generated, and the free aptamer was
captured in the test zone, with prominent decrease in fluorescence on the test line.
The test strip showed linear relationship in the range from 1 to 1000 ng · mL
−1 with
the LOD of 0.40 ng · mL
−1 , IC15 value of 3.46 ng · mL
−1 and recoveries from 96.4 to
104.67 % in spiked corn samples [40]. These findings demonstrated the superiority
of an aptamer-based LFA over conventional antibody-based strip assays and comparable performance to ELISA. In a similar approach, a simple, rapid, highly sensitive,
and cost-effective nucleic acid LFA was developed for the detection of HCV core
antigen, showing a lower detection limit of 100 pg.mL
−1 and 10 pg.mL
−1 with a
scanner and the naked eye, respectively [41].
A simple and sensitive aptamer-based biosensor for detection of Escherichia coli
O157: H7 (E. coli O157:H7) was developed [42]. In this assay, two different aptamers
specific for the outer membrane of E. coli O157:H7 were used. One of the aptamers
was used for magnetic bead enrichment, and the other was used as a signal reporter
for this pathogen, which was amplified by isothermal strand displacement amplification (SDA), which was further detected by a LFA. Only the captured aptamers on cell
membrane were amplified. The generated signals can be read out by the naked eye. As
low as 10 colony forming units (CFU) of E. coli O157:H7 were detected in this study.
Without DNA extraction, the reduced handling requirement and simplified equipment make this assay a simple and quick alternative to conventional methods. Other
important LFA sensors for bacterial detections include aptamer cocktail (mixture of
three aptamers for detection of E.coli) [43]. LFA has demonstrated its potential for
simple biosensors that could be used in POCT, and clinical diagnostics.
3.3.2 Interferometry
Interferometry is the superimposition of electromagnetic waves with similar or
different phases. Superimposition of electromagnetic waves which are similar
in phase produce constructive interference, while superimposition of those with
different phases yield destructive interference. The interaction between biomolecules
can affect the interference, and the information obtained from this interference change
59
Humanosteopontin (OPN) protein, a potential cancer biomarker, has substantial
promise for POCT. Biotinylated aptamer was recently used to precapture OPN from
samples, an OPN antibody was immobilized on the test line for second specific target
recognition, and streptavidin-modified GNPs sprayed on the color detection pad. This
achieved a sensitivity of as low as 0.1 ng. mL
−1 OPN with strong dynamic detection
between 10 and 500 ng. mL
−1 within 5 min. The LFA allowed the qualitative and
semiquantitative detection of OPN in serum at standard levels, as in cancer patients,
and can discriminate against OPN from high-specific protein interference [39].
Another example of aptamer-based competitive LFA includes one step determination of ochratoxin A (OTA) in corn samples. Briefly, biotin-cDNA was immobilized on a nitrocellulose filter on the test line. Without OTA, Cy5-labeled aptamer
combined with complementary strands formed a stable double helix. In the presence
of OTA, the Cy5-aptamer/OTA complexes were generated, and the free aptamer was
captured in the test zone, with prominent decrease in fluorescence on the test line.
The test strip showed linear relationship in the range from 1 to 1000 ng · mL
−1 with
the LOD of 0.40 ng · mL
−1 , IC15 value of 3.46 ng · mL
−1 and recoveries from 96.4 to
104.67 % in spiked corn samples [40]. These findings demonstrated the superiority
of an aptamer-based LFA over conventional antibody-based strip assays and comparable performance to ELISA. In a similar approach, a simple, rapid, highly sensitive,
and cost-effective nucleic acid LFA was developed for the detection of HCV core
antigen, showing a lower detection limit of 100 pg.mL
−1 and 10 pg.mL
−1 with a
scanner and the naked eye, respectively [41].
A simple and sensitive aptamer-based biosensor for detection of Escherichia coli
O157: H7 (E. coli O157:H7) was developed [42]. In this assay, two different aptamers
specific for the outer membrane of E. coli O157:H7 were used. One of the aptamers
was used for magnetic bead enrichment, and the other was used as a signal reporter
for this pathogen, which was amplified by isothermal strand displacement amplification (SDA), which was further detected by a LFA. Only the captured aptamers on cell
membrane were amplified. The generated signals can be read out by the naked eye. As
low as 10 colony forming units (CFU) of E. coli O157:H7 were detected in this study.
Without DNA extraction, the reduced handling requirement and simplified equipment make this assay a simple and quick alternative to conventional methods. Other
important LFA sensors for bacterial detections include aptamer cocktail (mixture of
three aptamers for detection of E.coli) [43]. LFA has demonstrated its potential for
simple biosensors that could be used in POCT, and clinical diagnostics.
3.3.2 Interferometry
Interferometry is the superimposition of electromagnetic waves with similar or
different phases. Superimposition of electromagnetic waves which are similar
in phase produce constructive interference, while superimposition of those with
different phases yield destructive interference. The interaction between biomolecules
can affect the interference, and the information obtained from this interference change