Instrumentation-Compact Digital Microfluidic Reaction Interface-Extended Loop-Mediated Isothermal Amplification for Sample-to-Answer Testing of <i>Vibrio parahaemolyticus</i>

Abstract

<i>Vibrio parahaemolyticus</i> is usually spread via consumption of contaminated seafood and causes vibriosis. By combination of digital microfluidic (DMF) and loop-mediated isothermal amplification (LAMP), we provided an automated instrumentation-compact DMF-LAMP device for sample-to-answer detection of <i>V. parahaemolyticus</i>. For the first time, how much the proper mixing might facilitate the DMF-LAMP process is explored. The results illustrated that increasing the number of flow configurations and decreasing the fluid-reversibility will extend the interfacial surface available for diffusion-based mass transfer within a droplet microreactor, thus contributing to the overall amplification reaction rate. Noticeably, the DMF-LAMP amplification plateau time is shortened by proper mixing, from 60 min in static mixing and traditional bulk LAMP to 30 min in 2-electrode mixing and 15 min in 3-electrode mixing. The device achieved much higher detection sensitivity (two copies per reaction) than previously reported devices. <i>V. parahaemolyticus</i> from spiked shrimps is detected by Q-tip sampling associated with 3-electrode mixing DMF-LAMPs. The detectable signal occurs within only 3 min at a higher concentration and, at most, is delayed to 18 min, with a detection limit of <0.23 × 10³ CFU/g. Thus, the developed DMF-LAMP device demonstrates potential for being used as a sample-to-answer system with a quick analysis time, high sensitivity, and sample-to-answer format.