Sub-5-Minute Ultrafast PCR using Digital Microfluidics.

The development of a rapid and reliable polymerase chain reaction (PCR) method for point-of-care (POC) diagnosis is crucial for the timely identification of pathogens. Microfluidics, which involves the manipulation of small volumes of fluidic samples, has been shown to be an ideal approach for POC analysis. Among the various microfluidic platforms available, digital microfluidics (DMF) offers high degree of configurability in manipulating μL/nL-scale liquid and achieving automation. However, the successful implementation of ultrafast PCR on DMF platforms presents challenges due to inherent system instability. In this study, we developed a robust and ultrafast PCR in 3.7-5 min with a detection sensitivity comparable to conventional PCR. Specifically, the implementation of the pincer heating scheme homogenises the temperature within a drop. The utilization of a μm-scale porous hydrophobic membrane suppresses the formation of bubbles under high temperatures. The design of a groove around the high-temperature zone effectively mitigates the temperature interference. The integration of a soluble sensor into the droplets provides an accurate and instant in-drop temperature sensing. We envision that the fast, robust, sensitive, and automatic DMF system will empower the POC testing for infectious diseases.