Effect of signaling probe conformation on sensor performance of a displacement-based electrochemical DNA sensor

Here we report the effect of the signaling probe conformation on sensor performance of a "signal-on" folding-based electrochemical DNA sensor. The sensor is comprised of a methylene blue (MB)-modified signaling probe and an unlabeled capture probe that partially hybridize to each other at the distal end. In presence of the full-complement target which binds to the unlabeled capture probe, the labeled signaling probe is released. Two different signaling probes were used in this study, in which one is capable of assuming a stem-loop conformation (SLP-MB), whereas the other probe adopts a flexible linear conformation (LP-MB). In the presence of the full complement target DNA, both sensors showed a large increase in MB current when interrogated using alternating current (ac) voltammetry, verifying the release of the signaling probe. Overall, the SLP-MB sensor showed higher % signal enhancement; the LP-MB sensor, however, showed distinctly faster binding kinetics when interrogated under the same experimental conditions. The SLP-MB sensor displayed a wider usable ac frequency range when compared to the LP-MB sensor. Despite these differences, the detection limit and dynamic range were found to be similar among the two sensors. In addition to 6-mercapto-1-hexanol, longer chain hydroxyl-terminated alkanethiols were used to construct these sensors. Our results showed that sensors fabricated with longer chain diluents, independent of the sensor architecture, were not only functional, the signaling capability was significantly enhanced.