Bioaffinity Detection of Biomolecules in Complex Matrices: Real Time Detection of DNA/RNA-Hybridization in Cell Extract Via Electrochemical Impedance Spectroscopy Measurements

Abstract

The quantification of significant amounts of disease biomarkers circulating in the bloodstream represents one of the challenging frontiers in biomedicine. The complexity of blood composition has opened the quest for novel detection technologies, capable of discerning small amount of specific biomarkers from other blood proteins/oligonucleotides and of reliably measuring them. In this context, we have developed a device based on differential double-layer capacitance readout at microfabricated gold electrodes and demonstrated its detection performance in real bio-sample volumes. By means of electrochemical impedance spectroscopy (EIS) measurements in a three electrodes setup, we first calibrated the system following the in situ hybridization of a self-assembled monolayer of single-stranded, short oligonucleotides on the gold microelectrode through the measurement of differential capacitance changes as a function of time, for different concentrations of complementary DNA in a saline buffer. Based on this calibration we used the device to quantify the presence of microRNAs (miRNAs) in cellular extract. We demonstrated that our device is fast, sensitive, reusable, reproducible and perfectly suited to measure binding affinities in complex matrices, as cell lysate, serum and blood. We put forward the possibility to apply this platform to the bioaffinity detection of protein biomarkers as well as circulating drugs in blood, for therapeutic drug monitoring applications.