Abstract
Small molecules are important
targets with the potential of clinical or commercial applications such as
medical diagnostics, environmental monitoring, and forensic science. Thus,
efforts to develop methods for portable, low-cost, and quantitative on-site
detection of a broad range of small molecules are gaining momentum.Various immunoassays have also been
developed for the detection of small molecules such as cocaine and/or its major
metabolite benzoylecgonine in biofluids, including the enzyme-linked
immunosorbent assay (ELISA).
Unfortunately, the use of immunoassays for the detection of designer
drugs is often limited because of the high cost of generating new antibodies
and issues with narrow target binding-spectrum and poor specificity, and most
of these assays offer only limited capabilities for naked-eye detection,
because the resulting absorbance changes can only be detected by instruments. FIU inventors have developed aptamer
sensors that report the presence of small-molecule target such as cocaine and
synthetic cathinones via a sensitive colorimetric signal for naked-eye
detection. The aptamer sensors are CBSAzyme-based sensors having both target-mediated cooperative behavior of the
CBSA and peroxidase-mimicking catalytic activity of DNAzyme. The CBSAzyme-based sensors comprise a long
fragment and a short fragment, the long fragment comprising a first segment of
a split DNAzyme and a long fragment of a CBSA, the short fragment comprising a
second segment of the split DNAzyme and a short fragment of the CBSA. The two fragments of the CBSAzyme remain
separate in the absence of the small-molecule target, but effectively assemble
in the presence of the small-molecule target. The assembly of the two fragments of the
CBSAzyme activates the DNAzyme that subsequently catalyzes the oxidation of
2,2'-azinobis(3-ethylbenzthiazo-line)-6-sulfonic acid (ABTS), producing a
visible color change from colorless to dark green that reveals the presence of
the target within minutes.In an initial
demonstration, the inventors have generated a cocaine-binding CBSAzyme that
enables naked-eye detection of this drug at concentrations as low as 10 uM
within 15 minutes. They subsequently demonstrated the generality of this assay
strategy by coupling the same split DNAzyme into a CBSAzyme that responds to
the designer drug methylenedioxypyrovalerone (MDPV). This CBSAzyme enabled
visual detection of MDPV and 11 other synthetic cathinones at concentrations as
low as 30 uM within 15 minutes, but did not respond to five common cutting
agents.
Benefit
Naked-eye detection of small molecules within minutesHigh sensitivity and specificityCompatible for both clinical and field settingsLabel-free and instrument-free
Market Application
Colorimetric assays in both clinical and field settings for small-molecule targets including illicit drugs, toxins, disease biomarkers, and pharmaceuticals for applications such as law enforcement, environmental monitoring, and clinical diagnostics.
Publications
Luo Y, Yu H, Alkhamis O, et al.
Label-Free, Visual Detection of Small Molecules Using Highly Target-Responsive
Multimodule Split Aptamer Constructs. Anal Chem. 2019;91(11):7199-7207.
doi:10.1021/acs.analchem.9b00507
Brochure
