A disposable device is described for use in High Content Screening within the drug research process; it interfaces with a standard 384 well microtiter plate providing simultaneous full kinetic reaction data for a complete row of 24 wells. The device can be used in secondary screening for lead optimisation, for ADME (Absorption Distribution Metabolism and Excretion) assessment and toxicological assessment of candidate compounds. The device is a microfluidic unit and incorporates the necessary substrate and reaction chemicals. The microfluidic circuitry includes mixing paths, a reaction chamber, a single use valve and a waste reservoir as well as electrical measurement contacts. Movement of the fluids is provided by a single vacuum line which is distributed to all 24 separate cells. The device described incorporates a self-validation technique which provides a built in test facility.

Traditional laboratory ultraviolet/visible/near-infrared spectroscopy instruments are tabletop-sized pieces of equipment that exhibit very high performance, but are generally too large and costly to be widely distributed for process control applications or used as spectroscopic sensors. Utilizing a unique, and proven, micro-optical technology platform origi-nally developed, qualified and deployed in the telecommunications industry, we have developed a new class of spectro-scopic micro-instrumentation that has laboratory quality resolution and spectral range, with superior speed and robust-ness. The fundamentally lower cost and small form factor of the technology will enable widespread use in process moni-toring and control. This disruption in the ground rules of spectroscopic analysis in these processes is enabled by the re-placement of large optics and detector arrays with a high-finesse, high-speed micro electro mechanical system (MEMS) tunable filter and a single detector, that enable the manufacture of a high performance and extremely rugged spectrome-ter in the footprint of a credit card. Specific process monitoring and control applications discussed in the paper include pharmaceutical, gas sensing and chemical processing applications.

MALDI-MS (matrix-assisted laser desorption/ionization mass spectrometry) is one of the most commonly used techniques for protein analysis. In conventional systems sample preparation is typically done in well-plates and transferred onto a MALDI target by robotic systems, which are complex, huge, expensive and slow. In this paper, we present a droplet-based microfluidic interface to transfer protein samples from a well-plate format onto a MALDI target for MS analysis. The droplets are actuated using the electrowetting phenomenon, and are immersed in silicone oil which prevents non-specific adsorption and enables the manipulation of high concentrations of proteins. Droplet transport and droplet formation were evaluated as a function of protein concentration using bovine serum albumin (BSA) as a test system. Droplet transport was possible for BSA concentrations up to 10mg/mL which is three orders of magnitude higher than previously reported results on handling proteins by electrowetting. Droplet formation from on-chip reservoirs, using only electrowetting forces and no external pressure assistance, was possible up to concentrations of 0.01mg/mL. An interface between a well-plate format and the electrowetting chip, and a scheme to passively stamp droplets onto a target substrate was then designed and tested by stamping BSA solutions. In two separate experiments 3.6fmoles and 16fmoles of BSA were stamped onto a glass slide using 0.001mg/mL and 0.01mg/mL samples respectively. A protein mixture with known constituents (ABI 4700 proteomics analyzer calibration solution) was stamped onto a MALDI plate and the individual proteins were correctly identified in the mass spectrum obtained using MALDI-TOF MS. The preliminary results establish the feasibility of using an electrowetting-based microfluidic system to handle proteins especially for protein stamping applications. The proposed system has a small footprint, is easy to control, and is very fast compared to conventional robotic systems. In addition, there are no moving parts and the associated mechanical reliability issues. Future work involves scaling to a larger number of samples and integration of sample preparation steps on-chip.

Intel's Precision Biology research effort is working to combine Intel's expertise in nanotechnology with aspects of biology and medicine to create highly sensitive instrumentation for biomolecular analysis. The ability to manipulate, detect, and identify biological molecules at ultra-low concentrations is important for applications ranging from whole-genome DNA sequencing to protein-based early disease detection. In this paper we describe our work to develop a molecular labeling system based on Surface-Enhanced Raman Spectroscopy (SERS), to enable highly sensitive protein detection. We also present a variety of techniques our team has developed for microfluidic transport and identification of single molecules in solution.

Sandia's hand-held MicroChemLab^TM system uses a micromachined preconcentrator (PC), a gas chromatography channel (GC) and a quartz surface acoustic wave array (SAW) detector for sensitive/selective detection of gas-phase chemical analytes. Requisite system size, performance, power budget and time response mandate microfabrication of the key analytical system components. In the fielded system hybrid integration has been employed, permitting optimization of the individual components. Recent improvements in the hybrid-integrated system, using plastic, metal or silicon/glass manifolds, is described, as is system performance against semivolatile compounds and toxic industrial chemicals. The design and performance of a new three-dimensional micropreconcentrator is also introduced. To further reduce system dead volume, eliminate unheated transfer lines and simplify assembly, there is an effort to monolithically integrate the silicon PC and GC with a suitable silicon-based detector, such as a magnetically-actuated flexural plate wave sensor (magFPW) or a magnetically-actuated pivot plate resonator (PPR).

Rapid identification of viral and bacterial species is dependent of the ability to manipulate biological agents into a form where they are directly analyzed. Many of these species of interest, such as bacterial spores, are inherently hearty and very difficult to lyse or solubilize. Standard protocols for spore inactivation include, chemical treatment, sonication, pressure and thermal lysis. While these protocols are effective for the inactivation of these agents they are less well suited for sample preparation for analysis using capillary electrophoresis techniques. In order to overcome this difficulty we fabricated a simple capillary device to perform thermal lysis of vegatative bacterial cells and bacterial spores. Using an ethylene glycol buffer to super heat bacterial spores we were able to perform rapid flow through lysis and solubilzation of these agents. This device was then coupled to a sample preparation station for on-line fluorescamine dye lableling and buffer exchange for direct analysis using a miniaturized capillary electrophoresis instrument. Using this integrated device were we enabled to perform sample lysis, labeling and protein fingerprint analysis of vegatative bacterial cells, bacterial spores and viruses in less than 10 minutes. The described device is simple, inexpensive and easily integratable with various microfluidic devices.

The integration of optical transducers is generally considered a key issue in the further development of lab-on-a-chip Microsystems. We present a technology for miniaturized, polymer based lasers, suitable for integration with planar waveguides and microfluidic networks. The lasers rely on the commercial laser dye Rhodamine 6G as active medium, and the laser resonator is defined in a thin film of polymer on a low refractive index substrate. Two types of devices are demonstrated: solid and microfluidic polymer based dye lasers. In the microfluidic dye lasers, the laser dye is dissolved in a suitable solvent and flushed though a microfluidic channel, which has the laser resonator embedded. For solid state dye lasers, the laser dye is dissolved in the polymer forming the laser resonator. The miniaturized dye lasers are optically pumped by a frequency doubled, pulsed Nd:YAG laser (at 532 nm), and emit at wavelengths between 560 nm and 590 nm. The lasers emit in the plane of the chip, and the emitted light is coupled into planar polymer waveguides on the chip. The feasibility of three types of polymers is demonstrated: SU-8, PMMA and a cyclo-olefin co-polymer (COC) - Topas. SU-8 is a negative tone photoresist, allowing patterning with conventional UV lithography. PMMA and Topas are thermoplasts, which are patterned by nanoimprint lithography (NIL). The lasing wavelength of the microfluidic dye lasers can be coarse tuned over 30 nm by varying the concentration of laser dye, and fine tuned by varying the refractive index of the solvent. This is utilized to realize a tunable laser, by on-chip mixing of dye, and two solvents of different index of refraction. The lasers were also integrated with waveguides and microfluidic networks.

A polymer based biochip for rapid 2D separations of peptides, proteins, and other biomedically relevant molecules was designed and fabricated. Like traditional 2D polyacrylamide gel electrophoresis (2D-PAGE) methods, the device will allow molecules to separate based on isoelectric point (pI) and molecular weight (MW). Our design, however, integrates both an initial capillary isoelectric focusing (cIEF) step followed by capillary electrophoresis (CE) in multiple parallel channels, all on a single microfluidic chip. Not only is the "lab-on-a-chip" design easier to use and less expensive, but the miniaturization of the device produces very rapid separations. Compared to traditional 2D-PAGE, which can take hours to complete, we estimate separation times on the order of seconds. Fluorescence detection will be used in the preliminary stages of testing, but the device also is equipped with integrated electrodes in the electrophoresis channels to perform multiplexed electrochemical detection for quantitative analysis. We will present preliminary results of the chip development and testing.

We present a new method to manufacture arrays of microlenses with varying diameter and/or varying focal length on the same substrate material. The method combines direct laser machining with a casting method and is based on the exposure of poly-methylmethacrylate (PMMA) to an UV Excimer Laser (248 nm). A following thermal treatment of the PMMA results in spherical caps in the PMMA which subsequently serves as a mould to replicate inverse structures in poly-dimethylsiloxane (PDMS) by casting. Lenses with a focal length of 300 µm to 4,000 µm have been realized in a PDMS replicate from the PMMA, however, this method is not limited to these materials if the soft embossing technology is applied where an elastomer such as PDMS serves as the mould.

In this paper we present a three-dimensional waveguide structure with unique optical and fluidic properties and demonstrate its application as a substrate for DNA microarrays. The structure is fabricated by thermal oxidation of a macroporous silicon membrane with a periodic pattern of discrete pores running perpendicular through the substrate. Partial oxidation generates a SiO₂ membrane, but leaves a rectangular grid of silicon walls dividing the membrane into compartments. We show that the SiO₂ walls act as optical waveguides and characterize their optical properties; modes can be launched using Koehler illumination. The silicon walls optically isolate adjacent compartments and prevent light from spreading laterally in the membrane. In a DNA hybridization experiment, the detection of 100 attomol of a Cy-3 labeled DNA fragment (17 oligonucleotides) has been achieved with a signal to noise ratio of > 3:1. We believe that even lower detection limits can be achieved by further tuning the optical parameters of the three-dimensional waveguide structure.

Lasers microprocessing is attractive for the custom fabrication of novel lab-on-a-chip designs. However, processing of glass biochips is challenging for most lasers because of the weak light interactions inherent in such transparent substrates. The F2-laser generates a high 7.9-eV photon energy that drives strong absorption in glasses, while the short 157-nm wavelength offers high-resolution patterning on the 100-nm scale. With these benefits, F2-laser ablation is well suited to the fabrication of high aspect ratio microfluidic channels and other biochip functions. F2-laser radiation also produces a strong photosensitivity response in fused silica and other glasses that enable the fabrication of buried optical waveguides, Bragg grating filters and other refractive index structures inside the glass. In this paper, we combine laser micromachining and refractive index profiling to enable single-step integration of photonic functions with microfluidic functions on a single chip. Optical waveguides were written to intercept microfluidic channels for optical sensing of cells and other bio-materials. An integrated biophotonic sensor is demonstrated for polystyrene spheres. The sensor is optically characterized for insertion loss, propagation loss, and particle sensitivity. The demonstration and analysis of this simple device offers insight into the capabilities and potential applications for laser fabricated glass lab-on-a-chip devices. Moreover, the groundwork is laid for rapid laser prototyping of custom-designed microfluidic biochips interlaced with integrated-optical circuits to define a new generation of highly functional bio-sensor and lab-on-a-chip devices.

Laser micromachining combined with digital printing allows rapid prototyping of complex bioreactors with reduced fabrication times compared to multi-mask photolithography. Microfluidic bioreactors with integrated optical waveguides for diagnostics have been fabricated via ultrashort pulse laser micromachining and digital printing. The microfluidic channels are directly laser machined into poly(dimethylsiloxane) (PDMS) silicone elastomer. Multimode optical waveguides are formed by coating the PDMS with alternating refractive index polymer layers and laser machining to define the waveguide geometry. Tapered alignment grooves are also laser machined to aid in coupling optical fibers to the waveguides. Three-dimensional (3-D) bio-scaffold matrices comprising liquid solutions that can be selectively and rapidly gelled are digitally printed inside the bioreactors and filled with nutrient rich media and cells. This paper will describe the maskless fabrication of complex 3-D bioreactors and discuss their performance characteristics.

An ideal on-site chemical/biochemical analysis system must be inexpensive, sensitive, fully automated and integrated, reliable, and compatible with a broad range of samples. The advent of digital microfluidic lab-on-a-chip (LoC) technology offers such a detection system due to the advantages in portability, reduction of the volumes of the sample and reagents, faster analysis times, increased automation, low power consumption, compatibility with mass manufacturing, and high throughput. We describe progress towards integrating sample collection onto a digital microfluidic LoC that is a component of a cascade impactor device. The sample collection is performed by impacting airborne particles directly onto the surface of the chip. After the collection phase, the surface of the chip is washed with a micro-droplet of solvent. The droplet will be digitally directed across the impaction surface, dissolving sample constituents. Because of the very small droplet volume used for extraction of the sample from a wide colection area, the resulting solution is realatively concentrated and the analytes can be detected after a very short sampling time (1 min) due to such pre-concentration. After the washing phase, the droplet is mixed with specific reagents that produce colored reaction products. The concentration of the analyte is quantitatively determined by measuring absorption at target wavelengths using a simple light emitting diode and photodiode setup. Specific applications include automatic measurements of major inorganic ions in aerosols, such as sulfate, nitrate and ammonium, with a time resolution of 1 min and a detection limit of 30 nm/m³. We have already demonstrated the detection and quantification of nitroaromatic explosives without integrating the sample collection. Other applications being developed include airborne bioagent detection.

Digital microfluidics is the second-generation lab-on-a-chip architecture based upon micromanipulation of droplets via a programmed external electric field by an individually addressable electrode array. Dielectrophoresis (DEP) and electrowetting-on-dielectric (EWOD) are of the dominant operating principles. The microfluidic mechanics of manipulating electrified droplets are complex and not entirely understood. In this article, EWOD and DEP are analyzed both analytically and numerically under a unified framework of droplet electrohydrodynamics (EHD). The numerical simulations based on droplet EHD are first validated against analytical and experimental results and have achieved a good agreement both quantitatively and qualitatively. Simulations are then used extensively in this article to illustrate device operation, to expose underlying physics, and to confirm our conclusions.

Biological applications of micro assay devices require easy implementable on-chip microfluidics for separation of plasma or serum from blood. This is achieved by a new blood separation technique based on a microchannel bend structure developed within the collaborative Micro-Tele-BioChip (μTBC) project. Different prototype polymer chips have been manufactured with an UV-LIGA process and hot embossing technology. The separation mechanisms have been identified and the separation efficiency of these chips has been determined by experimental measurements using human blood samples. Results show different separation efficiencies for cells and plasma up to 100 % depending on microchannel geometry, hematocrit, and feed velocity. This novel technique leads to an alternative blood separation method as compared to existing micro separation technologies.

A polymer microfluidic pump has been developed using electrostrictive poly(vinylidene fluoride-trifluoroethylene) based polymer, which possesses a large electrostrictive strain (5-7%) and high elastic energy density (1 J/cm³), as the driving microactuator. The microfluidic pump was realized by integrating a nozzle/diffuser type fluidic mechanical-diode structure with the polymer microactuator, which shows an actuation deflection of 80 mm for a pumping chamber of 2.2x2.2 mm². The microfluidic pump could pump methanol at a flow rate of 25 mL/min at 63 Hz with a backpressure of 350 Pa. The flow rate of this pump could be easily controlled by external electrical field. Results from both analytical and numerical analysis show that, due to the high load capability of the microactuator, the frequency response of this nozzle/diffuser pump is mainly limited by the resonance of the fluid in the fluid channel.

Valves for microfluidic systems have, for various reasons, proven to be difficult to fabricate, cumbersome to operate, and/or unreliable. We have explored the performance of a novel microfluidic valve formed by creating a flow channel past a Peltier junction. Using the Peltier junction as a thermoelectric cooler causes the fluid in the valve to freeze, forming a plug that blocks flow through the valve. Reversing the current in the Peltier junction causes the fluid to melt, reopening the valve. This type of valve is fundamentally leak-free, has no moving parts, and is electrically actuated. We have fabricated an experimental prototype capable of closing in less than one second, and of opening substantially faster. We have also developed a finite-element thermal model of the valve, and exercised it to optimize valve design. An optimized valve is predicted to have a cycle time on the order of 10 ms.

Genomic analysis tools have recently become an indispensable tool for the evaluation of gene expression in a variety of experiment protocols. Two of the main drawbacks to this technology are the labor and time intensive process for sample preparation and the relatively long times required for target/probe hybridization. In order to overcome these two technological barriers we have developed a microfluidic chip to perform on chip sample purification and labeling, integrated with a high density genearray. Sample purification was performed using a porous polymer monolithic material functionalized with an oligo dT nucleotide sequence for the isolation of high purity mRNA. These purified mRNA’s can then rapidly labeled using a covalent fluorescent molecule which forms a selective covalent bond at the N7 position of guanine residues. These labeled mRNA’s can then released from the polymer monolith to allow for direct hybridization with oligonucletide probes deposited in microfluidic channel. To allow for rapid target/probe hybridization high density microarray were printed in microchannels. The channels can accommodate array densities as high as 4000 probes. When oligonucleotide deposition is complete, these channels are sealed using a polymer film which forms a pressure tight seal to allow sample reagent flow to the arrayed probes. This process will allow for real time target to probe hybridization monitoring using a top mounted CCD fiber bundle combination. Using this process we have been able to perform a multi-step sample preparation to labeled target/probe hybridization in less than 30 minutes. These results demonstrate the capability to perform rapid genomic screening on a high density microfluidic microarray of oligonucleotides.

Gram-negative bacteria initiate a stress response in which the cells efflux potassium when electrophilic toxins are introduced into their environment. Hence, measurement of K⁺ concentration in the surrounding water using a fluorescence-based potassium-selective optode has been proposed for environmental and homeland security applications. Unfortunately, the fluorophore commonly used in such an optode is inefficient. Surface enhanced fluorescence (SEF) can be used to increase its fluorescence efficiency, which will improve the sensor's performance. To understand this phenomenon before applying it to the optode system, Rose Bengal (RB), an inexpensive and well characterized dye, in solution with gold and silver nanoparticles was studied. As expected, fluorescence from RB-gold solutions was low since alignment of gold's surface plasmon resonance (SPR) peak and absorption and fluorescence energies in RB favored energy transfer from RB to the gold nanoparticles. The alignment of the silver's SPR peak and the RB transitions favored transfer from silver to RB. SEF was observed in solutions with large dye-to-silver separation. However, little fluorescence was observed when the solution was pumped at the silver's SPR peak. Fluorescence from the dye decreased as dye-to-silver separation decreased. An explanation for these observations is presented; additional research is needed to develop a complete understanding.

Nonlinear spectroscopy based on multi-photon laser wave mixing is presented as a sensitive analytical technique for rapid detection and imaging of various analytes in microfluidic and microarray devices. Capillary electrophoresis microchips and DNA microarrays offer improved speed of analysis over conventional methods. Laser wave-mixing optical methods offer sensitive detection of small changes in chemical and physical properties. Wave-mixing spectroscopy is especially effective in using small optical path lengths available in microchips. This unusually sensitive optical absorption detection method can keep up with fast changing environments in microfluidic systems. In wave mixing, two laser beams are focused and mixed inside the analyte. The analyte probe volume, i.e., the beam overlap volume, is very small. Hence, it is inherently suitable for interfacing to microchips and microarrays for high spatial resolution analysis. Signal collection is very efficient since the signal is a coherent laser-like beam. The signal has a quadratic dependence on analyte concentration and a cubic dependence on laser power. Hence, one can monitor small changes in signal more effectively and one can use low-power compact lasers efficiently.

This paper describes a laser-induced fluorescence (LIF) detection system for imaging proteins separated in a microfluidic device. The diameter of a laser beam is first increased through a beam expander, and subsequently focused into a line using a cylindrical lens. The resultant laser line is used to image an entire capillary or channel in which protein separation took place. The fluorescence emission is collected with a cooled, scientific grade charge-coupled device (CCD) camera. The detection limit was determined using a series of concentrations of fluorescein solutions. The temporal and spatial effects of photobleaching from laser irradiation were analyzed and the parameters to reduce the effect of photobleaching are discussed. We used the imaging system to demonstrate rapid analysis of proteins using isoelectric focusing.

A probe was developed to detect biomolecular binding events on a substrate in the microwave regime in real time and without labels. The probe consists of a coplanar transmission line fabricated on a glass slide that can detect dielectric changes in close proximity of the substrate-liquid interface. The probe behavior was evaluated by adsorbing polyelectrolyte monolayers of alternating charge. Biomolecular detection was demonstrated by immobilizing protein A on the glass surface and detecting rabbit IgG molecules in a flow channel. The sensitivity of the probe was conservatively estimated to be ~100 pg/mm².

Sensors based on macroporous silicon (M-PSI) have demonstrated the ability to detect the presence of certain chemical and biological materials. The devices utilize silicon sensing membranes with deep trench structures (macropores) formed by electrochemical etching to depths up to 100μm. The sensor structure is unique in that it exploits the vertical dimension of the planar silicon substrate, utilizing the large internal surface area of the membrane as the active sensing region. Upon exposure to organic solvents (i.e. ethanol, acetone, benzene) the devices exhibit a characteristic impedance signature. Discrimination is achieved by recognizing a specific response characteristic, or by placing appropriate probe materials to provide an electrically detectable signal upon exposure to the target substance. M-PSi sensing devices have demonstrated an electrical response to DNA hybridization and shown discrimination between binding and non-binding events. The size of the pores in the sensing elements can host larger molecules such as proteins, which extends the use of the devices to other fields of biotechnology. The sensors have been designed and fabricated in array configurations. A flexible electronics interface platform has been developed to accommodate the use of the sensors for a variety of applications.

A novel process of casting the polydimethyl-siloxane (PDMS) microstructure with hybrid photoresists mold has been developed to fabricate the 3D microstructure. This new processing includes two parts: the 3D mother mold fabrication and PDMS casting processing. The 3D mother mold, which consists of the three-dimensional partial-spherical microstructure and micro channel, was successfully fabricated and characterized. The 3D micro structure and the micro channel of the mother mold, made of two different photosensitive materials, AZ100XT and SU8 photoresist respectively, are merged very smoothly at the joint area. A mother mold of a 2200-μm-diameter chamber with a 500-μm-width channel was presented in this paper. For the best precise dimensional control, we used this mother mold to fabricate the PDMS mold for PDMS casting processing. The surface average roughness of the final 3D structure is 30 nm. This novel processing provides a new technology for achieving smooth 3D chamber surface joined with microchannel. This new technology can be applied in various lab-on-a-chip and microfluidic devices such as micropump and microvalve for which the great sealing, no dead volume and high back pressure are critical requirements. In this paper, the design, fabrication process and surface profile characterizations of this processing are presented in details.

This paper investigates two-dimensional, time-dependent electroosmotic flows driven by an AC electric field via patchwise surface heterogeneities distributed along the microchannel walls. The time-dependent flow fields through the microchannel are simulated for various patchwise heterogeneous surface patterns using the backwards-Euler time stepping numerical method. Different heterogeneous surface patterns are found to create significantly different electrokinetic transport phenomena. It is shown that the presence of oppositely charged surface heterogeneities on the microchannel walls results in the formation of localized flow circulations within the bulk flow. These circulation regions grow and decay periodically in accordance with the applied periodic AC electric field intensity. The circulations provide an effective means of enhancing species mixing in the microchannel. A suitable design of the patchwise heterogeneous surface pattern permits the mixing channel length and the retention time required to attain a homogeneous solution to be reduced significantly.

The purpose of this study is to produce a platform device with the ability to detect a variety of pathogens based upon antigen-antibody interactions. The sensor comprises a nanoporous GeSe channel waveguide fabricated on a substrate, with an intermediate cladding buffer layer [GeSe₂], which is required when the substrate does not transmit at the desired λ. The light from a laser source is then coupled through a fiber and prism into the waveguide and collected with the help of a lens into a detector. The top cladding layer is a Ge₂₈Sb₁₂Se₆₀ thin film in which biomolecules can be 'tethered' via functionalization of the surface. Therefore the surface chemistry of the thin film and the specificity of antibody to its antigen are important considerations. This paper will focus primarily on the surface characterization of the top cladding layer using XPS, AFM, ellipsometry, contact angle measurements and diffuse reflectance analysis.