A Nucleic Acid Nanostructure Built through On-Electrode Ligation for Electrochemical Detection of a Broad Range of Analytes

Abstract

For an assay to be most effective in point-of-care clinical analysis, it needs to be economical, simple, generalizable, and free from tedious workflows. While electrochemistry-based DNA sensors reduce instrumental costs and eliminate complicated procedures, there remains a need to address probe costs and generalizability, as numerous probes with multiple conjugations are needed to quantify a wide range of biomarkers. In this work, we have opened a route to circumvent complicated multi-conjugation schemes using enzyme-catalyzed probe construction directly on the surface of the electrode. With this we have created a versatile DNA nanostructure probe and validated its effectiveness by quantification of proteins (streptavidin, anti-digoxigenin, anti-tacrolimus) and small molecules (biotin, digoxigenin, tacrolimus) using the same platform. Tacrolimus, a widely prescribed immunosuppressant drug for organ transplant patients, was directly quantified with electrochemistry for the first time, with the assay range matching the therapeutic index range. Finally, the stability and sensitivity of the probe was confirmed in a background of minimally diluted human serum.

Accelerated Electron Transfer in Nanostructured Electrodes Improves the Sensitivity of Electrochemical Biosensors

“…[ 5 , 6 , 7 , 8 , 9 , 10 , 11 ] Sensitivity is a key consideration for many biomedical applications, because many clinical biomarkers are present at nanomolar to picomolar concentrations, and the biosensor must achieve sufficient sensitivity in a complex background of interferent molecules. [ 12 , 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 ] Unfortunately, due to noise limitations in existing electronic measurement systems, the signal‐to‐noise ratio of conventional electrochemical biosensors degrades precipitously when they are miniaturized to the micron scale, [ 21 ] reducing their sensitivity and making meaningful measurements of analyte concentrations challenging or even impossible in many cases. [ 22 , 23 ]…”

Detection and beyond: challenges and advances in aptamer-based biosensors

“…This approach has been used to realize electrochemical detection of a broad range of analytes by employing the concept of on-electrode ligation. 163 The molecular sensor consisted of three DNA modules. The first was an anchor module that was immobilized on a gold electrode by a thiol group.…”

Reagentless biomolecular analysis using a molecular pendulum

“…Indeed, electrochemical sensors for glucose and lactate are the dominant sensors used for the development of dynamic detection systems for monitoring in physiological systems because redox-active enzymes can be used to report on the presence of the target analytes in the absence of added reporters. Reagentless electrochemical sensors that are instead affinitybased and compatible with in vivo monitoring applications have been generated based on DNA aptamers that serve as recognition elements [13][14][15][16][17][18] . While powerful for the collection of pharmacokinetic data in living systems where analyte concentrations are high, aptamer-based sensors typically have low binding affinities that render them incompatible with many sensing applications.…”