(166d) A New Quantitative Molecular Detection Platform for Field Applications

Despite claims to the contrary, commercially viable field-use genetic or immuno-assays have yet to appear on the market, even though upstream units like PCR chips, cell-lysing units and bead/membrane filtration technologies have matured sufficiently to be integrated into a hand-held portable device. The key technology bottle neck is detection. Current technologies, like real-time PCR, rely on fluorescent optical detection, which requires bulky and expensive equipment unsuitable for field applications. Label-free electrochemical sensors for DNA, RNA and peptide biomarkers, with functionalized molecular probes, are believed to be the most promising detection platform for such portable units. However, classical electrochemical sensing technologies based on conductance, capacitance or reactance measurements remain very insensitive, even with reporter redox pairs, voltage-gating and surface monolayer (SAM) electrode modification. Also, like fluorescent sensing, their diffusion-limited assay time is in excess of one hour for a typical field sample volume. We report here several field-use enabling breakthroughs in our laboratory for electrochemical sensing. A flow format is designed to effect convection-enhanced mass transfer and target concentration. The high flow rate also produces a shear-enhanced selectivity unavailable in standard batch optical platforms. A low-frequency AC field is applied across a nanoporous structure/membrane to deionize the flowing sample near the electrode sensor to enhance sensitivity by orders of magnitude. The high-field in the deionized region allows dielectrophoretic trapping of large target molecules from the flowing solution. The same AC field produces a Warburg spectrum at the electrode sensor to allow sensitive and quantitative detection of local conductivity/reactivity changes due to the presence of only a few thousand captured molecules. The result is a high throughput (10 microliter/min), high selectivity (SNP discrimination), low detection limit (picoMolar), calibrated (picoM to microM dynamic range) and rapid (10 minutes) detection platform that is currently being integrated with upstream units to produce a turn-key and portable commercial device that includes debris filtration, lysing and PCR amplification.