M. M. Sheehan

Thermal Performance Analysis of a Silicon Microreactor for Rapid DNA Analysis

This paper describes the use of thermal modelling tools in the design and characterisation of a multi-function silicon microreactor polymerase chain reaction (PCR) thermocycler system for rapid DNA diagnostic assays. MEMS technologies have been successfully applied to this application and the miniaturization of devices offers several advantages; including reduced assay times, reduced amounts of expensive reagents required as well as allowing rapid heating/cooling rates due to the lower thermal mass. This technique involves repetitive cycling at three different temperatures and requires rapid and accurate temperature changes. However, the direct measurement and monitoring of the temperature distribution on the nanostructures is still a challenge. Thus, this paper describes the use of static and transient thermal modeling to analyse the thermal performance of the system. The CFD code Flotherm was used to construct a 3D model of the PCR system. The system included assisted fan cooling in the thermal cycling chamber. Static thermal analysis was undertaken to simulate the temperature profile within the overall system and thus determine the fan characteristics needed to maintain system temperatures within operating requirements. Secondly, the PCR system operation involves repetitive thermal cycling of DNA at three different temperatures and requires rapid and accurate temperature changes. The thermal modeling results were used to determine the input power levels needed to obtain this required temperature/time profile

Design and fabrication of a silicon microreactor for DNA amplification

A silicon microreactor consisting of an integrated heater, temperature sensor and thermal isolation chamber has been described. The thermal characteristics of the device have been studied by computer simulation and a rapid heating rate (20/spl deg/C - 95/spl deg/C in less than 2 s) has been achieved. The fabrication process, consisting of microelectromechanical systems (MEMS) fabrication techniques has been established. The design features of this device, in particular the integrated heater and temperature sensor and the thermal isolation chamber allows fast heating/cooling rates and therefore enables efficient thermocycling suitable for DNA amplification.

Development of PCR conditions in a silicon microreactor DNA-amplification device

A silicon microsystem was developed which functions as a miniaturised DNA-amplification device. The system represents a technology platform for performing a polymerase chain reaction (PCR) with reduced volumes of 7 µL. The silicon microreactor was fabricated using silicon bulk micromachining, and a platinum heater was fabricated on a Pyrex substrate. A miniaturised DNA-amplification system permitted rapid heating and cooling, and shorter reaction times of 30 min were achieved. In this work, biocompatibility issues are addressed; conditions for efficient PCR in a silicon-based microreactor are established for the amplification of 500 bp DNA from the Escherichia coli bacteriophage Lambda; and the conditions are verified by amplifying a 255 bp region from the Mycobacterium tuberculosis rpoB gene. This work describes the PCR volume scale down experiments that were conducted and concentrations of the reactants; Taq polymerase, oligonucleotides, MgCl2 and template DNA were determined for DNA-amplification reactions with this novel device.

A photon-counting avalanche photodiode array with fully integrated active quenching and recharging circuit

The design of a 4 × 1 photon-counting avalanche photodiode array with fully integrated active bias controlling circuit is presented in this paper. The array uses highly sensitive Geiger mode avalanche photodiodes and is capable of detecting four single-photon-level optical signals simultaneously. The photodiode pixels can work either in parallel mode or independently because of the separate gate configuration. The photodiodes are fabricated using a CMOS compatible process with the integrated active quenching and recharging circuit manufactured via 1.5μm CMOS process. The whole system is included on a 2.5mm × 2.5mm die. Simulations show that the fully functional system can detect single photons at up to 20MHz with each pixel or 80MHz with all channels.

Nanostructured gold and platinum electrodes on silicon structures for biosensing

Gold and platinum metal electrodes on Si/SiO2 having undergone anisotropic potassium hydroxide (KOH) etch treatment are considered. This treatment etches at different rates and directions in the material resulting in creation of numerous pyramid shaped holes in the silicon substrate. This surface is used to make metal electrodes with increased electrode efficiency. The electrodes can serve as the sensors or as the sensor substrates (for surface polymer modification) and because both gold and platinum are inert they have applications for food safety biosensing. Wine, an economically significant food product, was chosen as a matrix, and impedance spectroscopy (EIS) was selected as a method of investigation of electrode behaviour. Based on results of EIS, different complexity equivalent circuits were determined by applying fitting mean square root optimisation of sensor complex impedance measurements.

Hybrid CMOS compatible active/passive quenching module

This paper demonstrates the experimental results of combining new state-of-the-art Geiger mode avalanche photodiodes with an integrated hybrid active/passive quenching circuit. This creates an ultra-compact form factor for a low-light level detection module. Both devices, the photodiode and the quenching circuit, are fabricated using conventional CMOS process technology and wafer substrates. The photodiodes operate at low voltage levels (30 V to 40V). Detector active areas are of various dimensions (10μm to 50μm) and shapes (circular, cylindrical or square). The integrated active/passive quenching circuit is included on a 2.5 mm × 2.5 mm die, which has the functionalities of bias conditioning, passive/active quench, output signal generation and active recharge. The prototypes are hybrid packaged onto a PCB substrate. The module is characterised for detecting very low level optical signals such as the single photon activities. Parameters such as dark counts, timing jitter, and responsivity will be shown for the compact detection module. Our findings show that the proposed avalanche photodiode operation is considerably faster than the conventional discrete systems and the module size is greatly reduced.

Silicon photomultipliers for improved biomolecule detection

There is a need for low cost, miniature, integrated optical systems for bioassay monitoring to meet the growing in vitro and point-of-care diagnostics markets. To this end, we are investigating the use of silicon photomultipliers (SiPM) as device upon which to base our technology development. SiPMs have been used successfully in many high-energy physics applications, but their application as a fully integrated biological detection platform has not been shown. In this paper we will present a new detection platform for the measurement of fluorescent biomolecules at much lower concentrations than commercially available systems. Our results show approaches that demonstrate the use of SiPM for the detection of fluorescent proteins and fluorescent-labelled DNA sequences. The SiPM and sample platforms are integrated so that the minimum distance separating the detector from the sample is realised. In addition, direct immobilisation of the DNA sequences onto the SiPM surface is achieved. This combined approach shows improved sensitivities for both the fluorescent proteins and fluorescent-labelled DNA. We are presenting results that show the use of SiPM as a successful technology for the measurement of fluorescent biomolecules at improved lower concentrations.

Bioassay platform for fiber coupled avalanche photodiode for improved biomolecule detection

For future fully integrated sensing applications, a CMOS sensor will be required. New CMOS photon counting sensors have recently become available and these devices provide high quantum efficiency, photon counting sensitivity, low power and new devices in arrays and with on-chip electronics. In biological applications, photon counting is focused on the detection of low intensity fluorescence signals from fluorophores conjugated to proteins or nucleic acid biomarkers from fluorescent proteins. We describe the development of a novel microtitre plate reader format, or bioassay platform that incorporates arrays of photon counting detectors for multiple parallel readout and data acquisition. Using Pyrex wafers, we have designed and fabricated custom-made reaction wells using Pyrex and deep ion trench etched silicon, which produce optically clear structures to facilitate fluorescence detection in biological samples volumes of 2 nL to 2 μL. For initial verification of the system, a new photon counting detector from SensL is used to determine the effectiveness of the wells as the bioassay platform. The compact unit consists of a fibre coupled silicon photon counting sensor, thermoelectric cooler, thermoelectric controller, active quenching circuit, power supplies, and an USB interface to the operating software. Included in the module is a counter with time binning capability. Sensitivity increases of more than two orders of magnitude in fluorescence detection are expected over commercially available instruments. This system demonstrates that a miniaturized, low cost solution is possible for fluorescence bioassay detection, which can be used to meet growing demands in the in vitro diagnostics and Point of Care markets.

Analysis of specification of an electrode type sensor equivalent circuit on the base of impedance spectroscopy simulation

Simulation of electrochemical impedance spectroscopy (EIS) based on a LabVIEW model of a complex impedance measuring system in the frequency domain has been investigated to specify parameters of Randles equivalent circuit, which is ordinarily used for electrode sensors. The model was based on a standard system for EIS instrumentation and consisted of a sensor modelled by Randles equivalent circuit, a source of harmonic frequency sweep voltage applied to the sensor and a transimpedance amplifier, which transformed the sensor current to voltage. It provided impedance spectroscopy data for different levels of noise, modelled by current and voltage equivalent noise sources applied to the amplifier input. The noise influence on Randles equivalent circuit specification was analysed by considering the behaviour of the approximation error. Different metrics including absolute, relative, semilogarithmic and logarithmic based distance between complex numbers on a complex plane were considered and compared to one another for evaluating this error. It was shown that the relative and logarithmic based metrics provide more reliable results for the determination of circuit parameters.