Understanding Changes of Both Faradaic and Nonfaradaic Currents with Temperature in DNA Monolayer-Based Sensors on Gold Electrodes

Abstract

Electrochemical sensors are well studied for detecting DNA, RNA, small molecules, inorganic ions, etc. Using the latest DNA-based sensors, a change in the location of a redox reporter attached to an electrode-bound nucleic acid gives rise to faradaic current changes, which can be monitored by square wave voltammetry (SWV) and used for sensing of various analytes. However, the nonfaradaic current generated along with the faradaic current has often been viewed as a limiting factor in sensor performance and either subtracted or ignored. Nonfaradaic current can be suppressed somewhat by surface blocking and reduced electrode surface area, and our group recently introduced a differential potentiostat (DiffStat) that provided efficient, in-hardware subtraction. On the other hand, removal of nonfaradaic current may lead to loss of important information embedded within. Moreover, a change in temperature can affect both faradaic and nonfaradaic currents by altering the movement of DNA strands and other ions present in the solution. Usually, faradaic current is proportional to the temperature. However, temperature-dependent changes in nonfaradaic current have not been well studied in DNA monolayer-based sensors. In this study, the changes in both faradaic and nonfaradaic currents were monitored in an electrochemical DNA hybridization assay. A thiolated DNA (thio-DNA) monolayer was assembled onto gold electrodes, and current was measured at different square wave frequencies (1 to 1000 Hz). The DNA analyte and the methylene blue tagged DNA (MB-DNA) were hybridized with the thio-DNA separately to compare the signals. Using an in-house built Peltier controller, the effects of temperature were monitored during operation of an analyte-induced proximity assay with SWV readout. In-house written MATLAB code was developed to automatically analyze each set of raw SWV data. Within this code, a polynomial baseline was calculated in the vicinity of the redox potential of MB-DNA (around -200 mV) to allow faradaic and nonfaradaic currents to be separated, post assay. Two-dimensional heatmaps were generated to study the variation of these currents with both temperature and square wave frequency. The faradaic current increased up to a certain temperature and then decreased due to dissociation (or melting) of the DNA from the surface. The nonfaradaic current, on the other hand, increased with temperature in a viscosity-dependent manner and also increased with the square wave frequencies as expected from a capacitive, single exponential time decay. Current changes exhibited by the DNA sensors upon analyte binding and temperature fluctuation suggested that both faradaic and nonfaradaic signals could be used for multidimensional analysis of sensor responses in the future. Figure 1