COMPACT PCR THERMAL CYCLERS FOR REAL-TIME DNA AMPLIFICATION AND ANALYSIS
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1 COMPACT PCR THERMAL CYCLERS FOR REAL-TIME DNA AMPLIFICATION AND ANALYSIS 1
2 Polymerase Chain Reaction Process (PCR) Description PCR requires continuous thermal cycling, i.e., alternately heating and cooling of the DNA sample based on a predefined series of temperature steps Thin-film etecs can enable a new generation of compact thermal cyclers in terms of form factor, function and use etec HV56 Thin-Film Thermoelectric Module Thin-film PCR Reference Design Features Microscopic size High heat flux Low thermal mass Heating & cooling in same device Solid-state Performance Benefits Rapid temperature control Greater throughput 1-2 sec response time at the Addressable temperatures system level Smaller diagnostic and fieldlevel thermal cyclers Demonstrated over 500,000 temperature cycles without significant change in AC resistance End Use Point of care Hospitals Academia Biotech/pharma Food safety
3 Overview Polymerase chain reaction (PCR) defined A scientific technique Amplifies a single or a few copies of a specific piece of DNA Generates thousands to millions of copies of a particular DNA sequence Applications for PCR DNA cloning for sequencing DNA-based analysis of genes Diagnosis of hereditary diseases Identification of genetic fingerprints Detection and diagnosis of infectious diseases PCR is becoming one of the most widely used techniques in molecular biology Rapid Inexpensive Simple process
4 Market Drivers PCR is a Growing Market Market witnessed rapid growth and technological improvements in the past three to five years Two main drivers: molecular diagnostics and the Human Genome Project (HGP) Main segments: drug discovery, DNA amplification, forensic identification, gene therapy and diagnosis Smaller format is the trend: testing bio-pathogens or DNA identification in the field rather than transporting samples to the lab
5 What is PCR? PCR stands for polymerase chain reaction Produces millions of copies of a specific DNA sequence in a short period of time. PCR uses same molecules that nature uses for copying DNA. Two short DNA sequences called primers. An enzyme called polymerase in a mixture. Mixture is placed in a thermal cycler. Alternating heating and cooling of the DNA sample/mixture based on a predefined series of temperature steps replicated the original DNA strain. During DNA replication, each strand of the original molecule acts as a template for the synthesis of a new, complementary DNA strand. Source: National Institute of General Medical Sciences, nigms.nih.gov.
6 PCR Thermal Cyclers A thermal cycler is an automated instrument specifically designed for PCR. A typical device consists of a metal block with holes where plastic vials holding the PCR reaction mixtures are inserted. The instrument has an integrated heating/cooling unit that is used to systematically raise and lower the temperature of the block. Most cyclers use thermoelectrics to control the temperature phases. Thermal cycle time is minimized and throughput maximized by minimizing the transition time between the phases.
7 Advantages of Thin-film in PCR Thermal Cycling Thin-film thermoelectric coolers (etecs) offer several key advantages that are particularly well suited for PCR Shorter throughput times for DNA amplification Fast temperature transition (10ºC per second) Small mass of thermal cycler Higher density of wells per machine Thin-film thermoelectrics can enable a new generation of compact thermal cyclers for equipment manufacturers that lower the barriers-toentry and increase opportunities for differentiation. Physical size of conventional TEC prevents this versus the microsize of etecs More precise temperature control Ultra-fast response time Uniquely high power pumping capability Enables smaller diagnostic and field-level thermal cyclers From the lab to the field Thin FIlm Conventional 6 W Module Comparison
8 Unique IP Position US Patent 7,164,077 issued in 2007 provides patent rights on use of thin-film thermoelectric cooling and heating in PCR thermal cycling. Also covers patent rights on any biological material cooling/heating with fast-acting thin-film TE devices. 8
9 Reference Design PCR subsystem consists of: Sample cartridge holder Support platform Thin-film thermoelectric module with integrated heat spreader interface Heat sink/fan assembly Plastic Platform Sample Cartridge 20W TEC Flex Interconnect COOLINOVATIONS Heat Sink Customizable design 1 to 4 samples or more at a time Form factor enables different thermal profile for each sample Hot-Side Heat Spreader Fan
10 Performance Specifications 50 ul Sample Response Using 4 HV56 Rise time: 70 C 95 C in 1s Fall time: 95 C 50 C in 2s Cycle: 29s; avg. power: 5.8W Generic PCR 50µL Using 4 x HV56 etecs TEC Power (W) Time (s) PTEC(W) ITEC(A) Tec Current (A)
11 Reliability of Thin-Film Technology in PCR Thermal Cycling Devices surpassed baseline tests in mechanical shock, thermal storage and power cycling. Subjected to over 500,000 power cycles with little change in AC resistance, a key measure of reliability and performance. Indicates stable device performance over a large number of cycles. Results strongly indicate modules are highly reliable for use in PCR thermal cycling applications.
12 Electrical Considerations for Reference Design Assembly contains four etec HV56 modules connected electrically in series. Optimal drive current is likely to be in the -0.9A to 0.6A range. Temperature range is created by switching polarity Use external thermistor to sense the load temperature Provide a control loop to adjust the etec current Recommend Laird PR-59 Programmable Controller TEC and Flex Interconnect Provides power, temp sensing, and feedback in a single system
13 Flexible Design Considerations Sample Size 5 µl to 50 µl sample volume is possible Enables low volume reaction platform Temperature Transition Rates Format Wells Temperature transitions at 20 C/s to µl; faster rates at lower volumes Designed for microtiter plates and lab-on-a-chip formats Scales from 1-4 or more wells in compact designs Addressability Microscale TECs enable variable single or multi-well temperature differentials and rates Electrical Specifications Series or parallel configurations match system voltages and drive current requirements.
14 Thin-Film TE Modules for Thermal Cycling Designed specifically for thermal cycling applications Available in variety of sizes and wattages Scales based on design requirements 20W TEC Array with Seal Ring
15 Design Services Modeling 1-D Modeling - assess the impact of thermoelectric coolers or power generation modules on a system 3-D Modeling - for complex systems, components or packages to understand heat flow and thermal gradients. Design Beneficial to organizations with limited thermal engineering expertise Engineering Prototyping - thermoelectric device and thermal rejection system integration Test & Measurement - thermal testing, evaluation and benchmarking
16 Key Design Specifications for PCR Dimensional Specifications Length Width Height Preferred Electrical Specifications Supply current Supply voltage Input power Assembly Requirements Assembly temperatures Wire size Wire bond pad requirements Solder compatibility, etc. Thermal Specifications Expected heat load Heat source target temperature Ambient temperature Sink-to-ambient thermal resistance
17 Summary Compact PCR and point-of-service applications are driving a sustained growth: Healthcare Forensics Identification Thin-film thermoelectric etecs enable: Faster thermal response time Smaller size Addressable temperatures Areas of customization and optimization Sample size Sample holder configuration System size Cost
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