ABSTRACT
The latest developments in thin-film-transistor digital-microfluidics (TFT-DMF, also known by the commercial name aQdrop™) are reported, and proof of concept application to molecular diagnostics (e.g. for coronavirus disease, COVID-19) at the point-of-need demonstrated. The TFT-DMF array has 41 thousand independently addressable electrodes that are capable of manipulating large numbers of droplets of any size and shape, along any pathway to perform multiple parallel reactions. Droplets are continually tracked and adjusted through closed-loop feedback enabled by TFT based sensors at each array element. The sample-to-answer molecular in vitro diagnostic (IVD) test for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) includes nucleic acid extractions from saliva, removal of dsDNA and quantitative reverse transcription polymerase chain reaction (RT-PCR). This proof of concept illustrates how the highly configurable TFT-DMF technology can perform many reactions in parallel and thus support the processing of a range of sample types followed by multiple complex multi-step assays.
Subject(s)
COVID-19/diagnosis , Microfluidics/methods , Transistors, Electronic , COVID-19/virology , Humans , Microfluidics/instrumentation , RNA, Viral/genetics , RNA, Viral/isolation & purification , Reverse Transcriptase Polymerase Chain Reaction , SARS-CoV-2/genetics , SARS-CoV-2/isolation & purification , Saliva/virologyABSTRACT
The widespread dissemination of CTX-M extended spectrum ß-lactamases among Escherichia coli bacteria, both in nosocomial and community environments, is a challenge for diagnostic bacteriology laboratories. We describe a rapid and sensitive detection system for analysis of DNA containing the blaCTX-M-15 gene using isothermal DNA amplification by recombinase polymerase amplification (RPA) on a digital microfluidic platform; active matrix electrowetting-on-dielectric (AM-EWOD). The devices have 16,800 electrodes that can be independently controlled to perform multiple and simultaneous droplet operations. The device includes an in-built impedance sensor for real time droplet position and size detection, an on-chip thermistor for temperature sensing and an integrated heater for regulating the droplet temperature. Automatic dispensing of droplets (45 nL) from reservoir electrodes is demonstrated with a coefficient of variation (CV) in volume of approximately 2%. The RPA reaction is monitored in real-time using exonuclease fluorescent probes. Continuous mixing of droplets during DNA amplification significantly improves target DNA detection by at least 100 times compared to a benchtop assay, enabling the detection of target DNA over four-order-of-magnitude with a limit of detection of a single copy within ~15 minutes.