Digital Microfluidics for Amplification Monitoring of Cancer Biomarkers ^†

Digital microfluidics (DMF) is an exciting technology for liquid handling, enabling low-volume droplet motion in electrode arrays, commonly mediated by voltage signals. Such technology allows for interesting applications in the field of molecular diagnostics, namely isothermal nucleic acid amplification for the detection of cancer biomarkers in low-volume samples. We developed a digital microfluidics platform capable of performing loop-mediated isothermal amplification (LAMP) of DNA while fitting under a fluorescence microscope, thus enabling a distinction between amplifying and non-amplifying samples, as well as monitoring the reaction progression. Our DNA amplification target is the cancer biomarker c-Myc, known as a “master regulator” due to its involvement in cell growth, proliferation and metabolism, which cumulatively intervenes in tumorigenesis. Specifically, the overexpression of c-Myc has been associated with as much as 40% of all human tumors. We successfully detected the amplification of the target DNA in under one hour with only 2 µL of total reaction volume. We also show the influence of mixing strategies when mixing LAMP reagents and DNA samples in our DMF platform, achieving uniform reagent distribution within reaction droplets, thus minimizing fluorescence signal fluctuations and maximizing reaction efficiency. Deep knowledge of mixing parameters on DMF is of utter relevance to the most recent enterprise in the group, which exploits DMF as a reagent mixing platform, a precursor to digital droplet PCR (ddPCR).