Research Article Strategies for enhancing the speed and integration of microchip genetic amplification In this work, we explore the use of methods that allow a significant acceleration of genetic analysis within microchips fabricated from low thermal conductivity materials such as glass or polymers. Although these materials are highly suitable for integrating a number of genetic analysis techniques onto lab-on-a-chip devices, their low thermal conductivity limits the rate at which heat can be transferred and hence lowers the speed of thermal cycling. However, short thermal cycling times are the key to bringing PCR to clinical point-of-care applications. Although shrinking the PCR reaction chamber volume can increase the speed of thermal cycling, this strategy is not always suitable, particularly when dealing with clinical samples with low analyte concentrations. In the present work, we combine two alternate strategies for decreasing the time required to perform PCR: implementing a heat sink and optimizing the PCR protocol. First, the heat sink substantially reduces the thermal resistance opposing heat dissipation into the ambient environment, and eliminates the parasitic thermal capacitance of the regions in the microchip that do not require heating. The low thermal conductivity of glass is used to our advantage to design the heat-sink placement to achieve fast thermal transitions while maintaining low power consumption. Second, we explore the application of two-stage PCR to provide a further reduction in the time required to perform genetic amplification by merging the annealing and extension stages of the commonly used three-stage PCR approach. In combination, we reduce the time required to perform thermal cycling by roughly a factor of 3 while improving the temperature control. Keywords: Electrophoresis / Microfluidics / PCR / Static and dynamic temperature measurements DOI 10.1002/elps.200800351 1 Introduction Molecular biology offers many useful techniques for disease diagnostics, but in their conventional form their cost and complexity limit their practical use outside of a laboratory setting. As a result, there has been substantial effort to miniaturize these techniques onto microfluidic platforms to improve speed, automation, portability, and cost-effectiveness (as reviewed in [1, 2]), moving towards a diagnostic and/or disease monitoring tool suitable for routine use in a clinical setting or in other point-of-care applications. PCR is one such molecular biology technique, and is a key technology for lab-on-a-chip (LOC) devices since it is used to amplify small amounts of DNA to levels that are readily detectable for analysis via electrophoresis. PCR involves manipulating DNA and enzymatic activity by repeatedly cycling a reaction mixture through distinct temperature stages. For high-yield PCR, temperature uniformity, accurate temperature control, and rapid transitions between temperature stages are all critical [3, 4]. Broadly, there are two major approaches to on-chip PCR [2]: the continuous flow approach and the stationary approach. The continuous flow approach can allow for very rapid temperature transitions by shuttling the PCR mixture between different temperature zones. However, we believe the stationary PCR approach is more suitable for the highly portable and inexpensive instruments that we have as our goal (e.g. our past work [24] that resulted in a $1000 instrument to perform reverse transcription (RT)-PCR-CE) since significantly less area is required. The first demonstrations of microchip PCR tended to be made from silicon [5–12] to take advantage of its favourable Viet N. Hoang 1 Govind V. Kaigala 1 Alexey Atrazhev 2 Linda M. Pilarski 2 Christopher J. Backhouse 1 1 Applied Miniaturization Laboratory, Electrical and Computer Engineering, University of Alberta, Edmonton, Alberta, Canada 2 Cross Cancer Institute and Oncology, University of Alberta, Edmonton, Alberta, Canada Received June 4, 2008 Revised August 29, 2008 Accepted August 29, 2008 Abbreviations: FE, finite element; LOC, lab-on-a-chip; RT, reverse transcription Correspondence: Professor Christopher J. Backhouse, ECERF Building, 2nd Floor, University of Alberta, Edmonton, Alberta, Canada T6G2V4 E-mail: chrisb@ualberta.ca Fax: 11780-492-1811 & 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.electrophoresis-journal.com Electrophoresis 2008, 29, 4684–4694 4684