Autonomous microfluidic multi-channel chip for real-time PCR with integrated liquid handling.
We report on a novel, polymer-based, multi-channel device for polymerase chain reaction that combines, for the first time, rapid sample processing in less than 5 min with high throughput at low costs. This is achieved by sample shuttling, during which submicroliter sample plugs (approximately 100 nl) are oscillated rapidly over three constant-temperature zones by pneumatic actuation with integrated system. The accuracy and the speed of the liquid handling have been significantly increased, while the design of the device can be kept very simple and allows for mass production using conventional low-cost polymer fabrication processes. Massive parallelization can lead to a throughput up to 100 samples in 10 min including the preparation time. The amplification can be optically monitored by means of online fluorescence detection. Successful real-time PCR and the determination of the threshold cycle, Ct, using the developed device were demonstrated with plasmid DNA in a fluorescent real-time format.
Frey
Olivier
O
Sensors, Actuators and Microsystems Laboratory, Institute of Microtechnology, University of Neuchâtel, Rue Jaquet-Droz 1, 2000, Neuchâtel, Switzerland. olivier.frey@unine.ch
Bonneick
Sonja
S
Hierlemann
Andreas
A
Lichtenberg
Jan
J
D000818:Animals; D001331:Automation; D052160:Benzothiazoles; D002021:Buffers; D002417:Cattle; D004247:DNA / Q000032:analysis / Q000235:genetics; D003959:Diamines; D004867:Equipment Design; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques; D009930:Organic Chemicals; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods*; D011804:Quinolines; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013048:Specimen Handling / Q000379:methods*; D019914:Taq Polymerase / Q000378:metabolism; D013997:Time Factors; D014325:Tromethamine / Q000737:chemistry
D052160:Benzothiazoles; D002021:Buffers; D003959:Diamines; D005456:Fluorescent Dyes; D009930:Organic Chemicals; D011804:Quinolines; D014325:Tromethamine; C098022:SYBR Green I; D012710:Serum Albumin, Bovine; D004247:DNA; D019914:Taq Polymerase; D016650:Fluorescein-5-isothiocyanate