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A novel flow-through device for performing fast PCR thermal cycling is presented. The thermal gradient thermal cycling device is comprised of layers of highly thermally conducting material separated by insulating layers. Channels etched in the conducting and insulating layers create one continuous path through the device. When the device is held between platens at different temperatures and PCR sample mix is pumped through it, every fluid particle undergoes the time-temperature protocol necessary for PCR but with a temperature change rate not possible with conventional cyclers. Ultrafast thermal cycling makes it ideal for bio-defense applications, such as the instantaneous bio-aerosol agent identification system under development for the Department of Homeland Security. Its compact size and simplicity of use make it a natural choice for diagnostics, forensics, food and water testing and other DNA testing applications. Herein we describe the design and fabrication of the device developed for IBADS and the subsequent performance with various assays using plasmid and genomic template DNA. Performance under some circumstances was exceptional: Amplification rates of up to two decades per minute were recorded and total amplification of up to eight decades in 30 cycles was seen. We discuss how to optimize the performance of a device that pushes PCR to its fundamental limits and review a wide variety of performance data.
Date of Publication: May 2008