Electrokinetic Enrichment and Label-Free Electrochemical Detection of Nucleic Acids by Conduction of Ions Along the Surface of Bioconjugated Beads for Point-of-Care Testing

In this paper, we report a method to integrate electrokinetic pre-enrichment of nucleic acids within a bed of probe-modified microbeads with their label-free electrochemical detection. In this detection scheme, hybridization of locally enriched target nucleic acids to the beads modulates the conduction of ions along the bead surfaces. This is a fundamental advancement in that this mechanism is similar to that observed in nanopore sensors, yet occurs in a bed of microbeads with microscale interstices. In application, this approach has several distinct advantages. First, electrokinetic enrichment requires only a simple DC power supply, and in combination with non-optical detection, makes this method amenable to point-of-care application. Second, the sensor is easy to fabricate, comprised of a packed bed of commercially-available microbeads, which can be readily modified with a wide range of probe types, thereby making this a versatile platform. Finally, the sensor is highly sensitive (picomolar) despite the modest 100-fold pre-enrichment we employ here by faradaic ion concentration polarization (fICP). Further gains are anticipated under conditions for fICP focusing that are known to yield higher enrichment factors (up to 100,000-fold enrichment). Here, we demonstrate detection of 3.7 pM single-stranded DNA complementary to the bead-bound oligoprobe, following a 30-min single step of enrichment and hybridization. Our results indicate that a shift in the slope of a current-voltage curve (CVC) occurs upon hybridization, and that this shift is proportional to the logarithm of the concentration of target DNA. Finally, we investigate the proposed mechanism of sensing by developing a numerical simulation that shows an increase in ion flux through the bed of insulating beads given changes in surface charge and zeta potential consistent with our experimental conditions.