D. Mark, T. van Oordt, O. Strohmeier, G. Roth, J. Drexler, M. Eberhard, M. Niedrig, P. Patel, A. Zgaga-Griesz, W. Bessler, M. Weidmann, F. Hufert, R. Zengerle, F. von Stetten
The world's growing mobility, mass tourism, and the threat of terrorism increase the risk of the fast spread of infectious
microorganisms and toxins. Today's procedures for pathogen detection involve complex stationary devices, and are often
too time consuming for a rapid and effective response. Therefore a robust and mobile diagnostic system is required. We
present a microstructured LabDisk which performs complex biochemical analyses together with a mobile centrifugal
microfluidic device which processes the LabDisk. This portable system will allow fully automated and rapid detection of
biological threats at the point-of-need.
KEYWORDS: Temperature metrology, Polymers, Microfluidics, Infrared radiation, Convection, Sensors, Reflectors, Polymer thin films, Data transmission, Process control
The direct on-disk wireless temperature measurement system [1,2] presented at μTAS 2010 was further improved in its
robustness. We apply it to an IR thermocycler as part of a centrifugal microfluidic analyzer for polymerase chain
reactions (PCR). This IR thermocycler allows the very efficient direct heating of aqueous liquids in microfluidic cavities
by an IR radiation source. The efficiency factor of this IR heating system depends on several parameters. First there is
the efficiency of the IR radiator considering the transformation of electrical energy into radiation energy. This radiation
energy needs to be focused by a reflector to the center of the cavity. Both, the reflectors shape and the quality of the
reflecting layer affect the efficiency. On the way to the center of the cavity the radiation energy will be diminished by
absorption in the surrounding air/humidity and especially in the cavity lid of the microfluidic disk. The transmission
spectrum of the lid material and its thickness is of significant impact. We chose a COC polymer film with a thickness of
150 μm. At a peak frequency of the IR radiator of ~2 μm approximately 85 % of the incoming radiation energy passes
the lid and is absorbed within the first 1.5 mm depth of liquid in the cavity. As we perform the thermocycling for a PCR,
after heating to the denaturation temperature of ~ 92 °C we need to cool down rapidly to the primer annealing
temperature of ~ 55 °C. Cooling is realized by 3 ventilators venting air of room temperature into the disk chamber. Due
to the air flow itself and an additional rotation of the centrifugal microfluidic disk the PCR reagents in the cavities are
cooled by forced air convection. Simulation studies based upon analogous electrical models enable to optimize the disk
geometry and the optical path. Both the IR heater and the ventilators are controlled by the digital PID controller HAPRO
0135 [3]. The sampling frequency is set to 2 Hz. It could be further increased up to a maximum value being permitted by
the wireless temperature data transmission system. As we are controlling a significantly non-linear process the controller
parameters need to be optimized for all temperatures relevant for the PCR thermocycling process. Such we get a
dynamic system for both, the heating and the cooling process. Heating rates up to 5 K/s with our IR heater (100 W
electrical power) could be achieved. Cooling rates of instantly 1.3 K/s at 20 Hz rotation frequency could be even further
increased by higher rotation frequencies, faster air circulation, optimization of the controller parameters or an active air
cooling unit.
Two microfluidic cartridges intended for upgrading standard laboratory instruments with automated liquid handling
capability by use of centrifugal forces are presented. The first microfluidic cartridge enables purification of DNA from
human whole blood and is operated in a standard laboratory centrifuge. The second microfluidic catridge enables
genotyping of pathogens by geometrically multiplexed real-time PCR. It is operated in a slightly modified off-the-shelf
thermal cycler. Both solutions aim at smart and cost-efficient ways to automate work flows in laboratories.
The DNA purification cartridge automates all liquid handling steps starting from a lysed blood sample to PCR ready
DNA. The cartridge contains two manually crushable glass ampoules with liquid reagents. The DNA yield extracted
from a 32 μl blood sample is 192 ± 30 ng which corresponds to 53 ± 8% of a reference extraction.
The genotyping cartridge is applied to analyse isolates of the multi-resistant Staphyloccus aureus (MRSA) by real-time
PCR. The wells contain pre-stored dry reagents such as primers and probes. Evaluation of the system with 44 genotyping
assays showed a 100% specificity and agreement with the reference assays in standard tubes. The lower limit of
detection was well below 10 copies of DNA per reaction.
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