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

Vibrio parahaemolyticus 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 V. parahaemolyticus. 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. V. parahaemolyticus 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 x 10(3) 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.

Automated summary

By combining digital microfluidic (DMF) with loop-mediated isothermal amplification (LAMP), an automated instrument-compact DMF-LAMP device was developed for rapid and sensitive detection of Vibrio parahaemolyticus. Proper mixing was found to enhance the amplification reaction rate, shorten the amplification time, and improve detection sensitivity of the device. The device showed high sensitivity with a detection limit of <0.23 x 10(3) CFU/g, making it a promising sample-to-answer system for quick and sensitive analysis.