Faradaic-free electrokinetic nucleic acid amplification (E-NAAMP) using localized on-chip high frequency Joule heating.

We present a novel Faradaic reaction-free nucleic acid amplification (NAA) method for use with microscale liquid samples. Unlike previous Joule heating methods where the electrodes produce electrolysis gaseous by-products and require both the electrodes be isolated from a sample and the venting of produced electrolysis gas, our electrokinetic Nucleic Acid Amplification (E-NAAMP) method alleviates these issues using a radio frequency (RF) alternating current electric field. In this approach, a pair of microscale thin film gold electrodes are placed directly in contact with a nucleic acid reaction mixture. A high frequency (10-40 MHz) RF potential is then applied across the electrode pair to induce a local Ohmic current within the sample and drive the sample temperature to increase by Joule heating. The temperature increase is sustainable in that it can be generated for several hours of constant use without generating any pH change to the buffer or any microscopically observable gaseous electrolysis by-products. Using this RF Joule heating approach, we demonstrate successful direct thermal amplification using two popular NAA biochemical reactions: loop-mediated isothermal amplification and polymerase chain reaction. Our results demonstrate that a simple microscale electrode structure can be used for thermal regulation for NAA reactions without observable electrolytic reactions, minimal enzyme activity loss and sustained (>50 h use per device) continuous operations without electrode delamination. As such, E-NAAMP offers substantial miniaturization of the heating elements for use in microfluidic or miniaturized NAA reaction systems.