A Novel Electrochemical DNA Sensor Based on Redox Modulated Fluorescence Intensity

In clinical applications, a highly sensitive and specific biosensor is required for detecting the trace amount of genetic analyte1. There have been numerous techniques developed for improving the sensitivity and selectivity of the DNA biosensor. Many PCR-based techniques including gene sequencing and fluorescence quantitative polymerase chain reaction have shown advantages such as high accuracy and sensitivity2. However, these techniques are complicated to use and require professionally trained personal. On the other hand, electrochemistry-based measurements such as Square-wave Voltammetry and Cyclic Voltammetry usually assume the electron transfer kinetics of the redox-active monolayer assemblies are uniform. The signal generated is an average and the detailed information about the modified surface is difficult to obtain. Here we show a novel detection method using electrochemistry coupled with fluorescence microscopy. A mixed SAM composed of Methylene Blue (MB) or AlexaFluor488 (AF488) labeled single stranded DNA were electro-deposited on a single crystal gold bead electrode3. We observed a unique profile of fluorescence signal (AF488) modulated by the redox probe (MB) upon DNA hybridization events. In addition, we have found that different surface crystallographies exhibit different sensitivities to the complementary strand (e.g. target) which can be explained by differences in surface coverages4. We demonstrate a coupled electrochemical and fluorescence DNA sensor. References: Pheeney, C. G.; Barton, J. K. DNA Electrochemistry with Tethered Methylene Blue. Langmuir 2012, 28(17), 7063–7070. Santhanam, M.; Algov, I.; Alfonta, L. DNA/RNA Electrochemical Biosensing Devices a Future Replacement of PCR Methods for a Fast Epidemic Containment. Sensors (Switzerland) 2020, 20 (16), 1–15. Ma, T.; Bizzotto, D. Improved Thermal Stability and Homogeneity of Low Probe Density DNA SAMs Using Potential-Assisted Thiol-Exchange Assembly Methods. Chem. 2021. 93 (48), 15973–15981. Mirmomtaz, E.; M. Castronovo.; L. Casalis. Quantitative study of the effect of coverage on the hybridization efficiency of surface- bound DNA nanostructures. Nano Lett. 2008, 8 (12), 4134–4139. Figure 1