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A digital microfluidic (DMF) system has been developed for loop-mediated isothermal amplification (LAMP)-based pathogen nucleic acid detection using specific low melting temperature (Tm) Molecular Beacon DNA probes. A positive-temperature-coefficient heater with a temperature sensor for real-time thermal regulation was integrated into the control unit, which generated actuation signals for droplet manipulation. To enhance the specificity of the LAMP reaction, low-Tm Molecular Beacon probes were designed within the single-stranded loop structures on the LAMP reaction products. In the experiments, only 1 μL of LAMP reaction samples containing purified Trypanosoma brucei DNA were required, which represented over a 10x reduction of reagent consumption when comparing with the conventional off-chip LAMP. On-chip LAMP for unknown sample detection could be accomplished in 40 min with a detection limit of 10 copies/reaction. Also, we accomplished an on-chip melting curve analysis of the Molecular Beacon probe from 30 to 75 °C within 5 min, which was 3x faster than using a commercial qPCR machine. Discrimination of non-specific amplification and lower risk of aerosol contamination for on-chip LAMP also highlight the potential utilization of this system in clinical applications. The entire platform is open for further integration with sample preparation and fluorescence detection towards a total-micro-analysis system.

The complexity of droplet hydrodynamics on a digital microfluidic (DMF) system eventually weakens its potential for application in large-scale chemical/biological micro-reactors. We describe here an intelligent DMF technology to address that intricacy. A wide variety of control-engaged droplet manageability is proposed and demonstrated through the operation of our modular DMF prototype, which comprises: (i) rigid profiling ability of different droplet's hydrodynamics under a real-time trajectory track of dropletderived capacitance, permitting accurate and autonomous multi-droplet positioning without visual setup and heavy image signal processing; (ii) fuzzy-enhanced controllability saving up to 21% charging time when compared with the classical approach, enhancing the throughput, fidelity and lifetime of the DMF chip, while identifying and renouncing those weakened electrodes deteriorated over time, and (iii) expert manipulability of multi-droplet routings under countermeasure decisions in real time, preventing dropletto-droplet or task-to-task interference. Altogether, this work exhibits the first modular DMF system with built-in electronic-control software-defined intelligence to enhance the fidelity and reliability of each droplet operation, allowing future manufacturability of a wide range of life science analyses and combinatorial chemical screening applications.

We developed a thermal digital microfluidic (T-DMF) device enabling ultrafast DNA melting curve analysis (MCA). Within 7 seconds, the T-DMF device succeeded in differentiating a melting point difference down to 1.6 °C with a variation of 0.3 °C in a tiny droplet sample (1.2 μL), which was 300 times faster and with 20 times less sample spending than the standard MCA (35 minutes, 25 μL) run in a commercial qPCR machine. Such a performance makes it possible for a rapid discrimination of single-nucleotide mutation relevant to prompt clinical decision-making. Also, aided by electronic intelligent control, the T-DMF device facilitates sample handling and pipelining in an automatic serial manner. An optimized oval-shaped thermal electrode is introduced to achieve high thermal uniformity. A device-sealing technique averts sample contamination and permits uninterrupted chemical/biological reactions. Simple fabrication using a single chromium layer fulfills both the thermal and typical transport electrode requirements. Capable of thermally modulating DNA samples with ultrafast MCA, this T-DMF device has the potential for a wide variety of life science analyses, especially for disease diagnosis and prognosis.

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