Microfluidic lab-on-a-foil for nucleic acid analysis based on isothermal recombinase polymerase amplification (RPA)

Sascha Lutz,†*^a Patrick Weber,†^a Max Focke,^b Bernd Faltin,^b Jochen Hoffmann,^b Claas Müller,^c Daniel Mark,^a Günter Roth,^b Peter Munday,^d Niall Armes,^d Olaf Piepenburg,^d Roland Zengerle^abe and Felix von Stetten^ab

Author affiliations

* Corresponding authors

^a HSG-IMIT, Wilhelm-Schickard-Straße 10, Villingen-Schwenningen, Germany
E-mail: sascha.lutz@hsg-imit.de
Fax: +49 761 203 7322
Tel: +49 761 203 7312

^b Laboratory for MEMS Applications, Department of Microsystems Engineering (IMTEK), University of Freiburg, Georges-Koehler-Allee 106, Freiburg, Germany

^c Laboratory for Process Technology, Department of Microsystems Engineering (IMTEK), University of Freiburg, Georges-Koehler-Allee 106, Freiburg, Germany

^d TwistDX, Babraham, Cambridgeshire, UK

^e Centre for Biological Signalling Studies (bioss), University of Freiburg, Germany

Abstract

For the first time we demonstrate a self-sufficient lab-on-a-foil system for the fully automated analysis of nucleic acids which is based on the recently available isothermal recombinase polymerase amplification (RPA). The system consists of a novel, foil-based centrifugal microfluidic cartridge including prestored liquid and dry reagents, and a commercially available centrifugal analyzer for incubation at 37 °C and real-time fluorescence detection. The system was characterized with an assay for the detection of the antibiotic resistance gene mecA of Staphylococcus aureus. The limit of detection was <10 copies and time-to-result was <20 min. Microfluidic unit operations comprise storage and release of liquid reagents, reconstitution of lyophilized reagents, aliquoting the sample into ≤30 independent reaction cavities, and mixing of reagents with the DNA samples. The foil-based cartridge was produced by blow-molding and sealed with a self-adhesive tape. The demonstrated system excels existing PCR based lab-on-a-chip platforms in terms of energy efficiency and time-to-result. Applications are suggested in the field of mobile point-of-care analysis, B-detection, or in combination with continuous monitoring systems.