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JOURNAL ARTICLE

Rapid and sensitive detection of antibiotic resistance on a programmable digital microfluidic platform

Sumit Kalsi, Martha Valiadi, Maria-Nefeli Tsaloglou, Lesley Parry-Jones, Adrian Jacobs, Rob Watson, Carrie Turner, Robert Amos, Ben Hadwen, Jonathan Buse, Chris Brown, Mark Sutton, Hywel Morgan
Lab on a Chip 2015 July 21, 15 (14): 3065-75
26086197
The widespread dissemination of CTX-M extended spectrum β-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.

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