2 research outputs found
Lateral Flow Aptasensor for Small Molecule Targets Exploiting Adsorption and Desorption Interactions on Gold Nanoparticles
A lateral flow assay
(LFA) can provide a rapid and cost-effective
means to detect targets in situ; however, existing LFA formats (predominantly
sandwich assays) are not suitable for small molecule targets. We present
a new LFA design that probes the dissociation of aptamers from the
surface of gold nanoparticles upon recognition of small targets. The
target-induced removal of aptamer molecules from the surface of the
colored particles results in the particles being captured on a test
line comprised of the protein bovine serum albumin immobilized on
nitrocellulose. On the other hand, in the absence of target, aptamer
coated particles are protected from capture on the test line and are
instead captured at a control line comprised of the protein lysozyme.
This protein is strongly positively charged under measurement conditions
and therefore captures all gold nanoparticles regardless of the presence
of aptamers. The effectiveness and operation mechanism of this simply
fabricated sensor was demonstrated by using a previously reported
35-mer aptamer for a small molecule, 17β-estradiol. The sensor
exhibited nanomolar level of detection, excellent selectivity against
potential interfering molecules, and robust operation in natural river
water samples. The simplicity and performance of the sensor platform
renders it applicable to a wide range of other aptamers targeting
small molecules, as we demonstrated with a novel bisphenol A aptamer.
Additionally, we show that our LFA design is not confined to the specific
proteins used as test and control lines, provided that their charge
is appropriate to modulate the interaction with aptamer-coated or
bare nanoparticles
Ultrasensitive Colorimetric Detection of 17β-Estradiol: The Effect of Shortening DNA Aptamer Sequences
We report a strategy enabling ultrasensitive
colorimetric detection
of 17β-estradiol (E2) in water and urine samples using DNA aptamer-coated
gold nanoparticles (AuNPs). Starting from an established sensor format
where aggregation is triggered when target-bound aptamers dissociate
from AuNP surfaces, we demonstrated that step-change improvements
are easily accessible through deletion of excess flanking nucleotides
from aptamer sequences. After evaluating the lowest energy two-dimensional
configuration of the previously isolated E2 binding 75-mer aptamer
(<i>K</i><sub>D</sub> ∼25 nM), new 35-mer and 22-mer
aptamers were generated with <i>K</i><sub>D</sub>’s
of 14 and 11 nM by simply removing flanking nucleotides on either
side of the inner core. The shorter aptamers were found to improve
discrimination against other steroidal molecules and to improve colorimetric
sensitivity for E2 detection by 25-fold compared with the 75-mer to
200 pM. In comparing the response of all sequences, we find that the
excess flanking nucleotides suppress signal transduction by causing
target-bound aptamers to remain adhered to AuNPs, which we confirm
via surface sensitive electrochemical measurements. However, comparison
between the 22-mer and 35-mer systems show that retaining a small
number of excess bases is optimal. The performance advances we achieved
by specifically considering the signal transduction mechanism ultimately
resulted in facile detection of E2 in urine, as well as enabling environmental
detection of E2 at levels approaching biological relevance