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This paper presents a microfluidic device that can passively reduce fluctuations in an upstream fluid flow and generate a largely steady flow in a microfluidic system. The device features a series of compliant membranes that form deformable walls of a microchannel. F or an incoming pulsatile flow, passive vibrations of the membranes allow the fluid to be accumulated for an overflow and discharged for an underflow, resulting in drastically reduced fluctuations in flow rate and pressure. A lumped-parameter model is developed to simulate the device characteristics, in which the coupling of membrane vibrations and fluid flow in individual channel sections associated with a single membrane is first represented by squeeze-film theory and inertia-free structural dynamics. The entire device is next represented as a system by connecting individual channel sections in series. The model is numerically solved, and the results agree with experiments. The theoretical and experimental results both show that a five-membrane device can stabilize a pulsating flow by a factor of about 20. These results also reveal that smaller average flow rate, smaller fluctuation, and higher fluctuation frequency lead to more effective flow stabilization. With these characteristics, the flow stabilizer can be used to obtain steady flow in microfluidic systems. [2008-0305] Index Terms---Compliant membrane, flow rectification, flow stabilization, fluid-structure interaction, lumped-parameter model.

We report fabrication and characterization of microfluidic devices made of thermoplastic and elastomeric polymers. These hard-soft hybrid material devices are motivated by the combined need for large scale manufacturability, enhanced barrier properties to gas permeation and evaporation of aqueous solutions compared to poly(dimethyl siloxane) (PDMS) devices, and compatibility with deformation-based actuation. Channel features are created on rigid polymers such as polyethylene terephthalate glycol (PETG), cyclic olefin copolymer (COC), and polystyrene (PS) by hot embossing. These 'hard tops' are bonded to elastomeric 'soft bottoms' (polyurethane (PU) or PDMS-parylene C-PDMS) to create devices that can be used for microfluidic cell culture where deformation-based fluid actuation schemes are used to perfuse and recirculate media. The higher barrier properties of this device compared to PDMS devices enable cell culture with less evaporation and creation of hypoxic conditions.

Here we propose optofluidic spherical microlenses that can change their focal distance by varying the refractive index of the liquid that composes them. These lenses are fabricated in the bulk of a polymeric mixture. Results of a characterization study of the profile of the lenses, the image forming capability, and the behavior of the focal distance as a function of the refractive index are presented. Ionic liquids are suggested as a source of liquids useful for fabricating this type of lens. OCIS codes: 130.3990, 350.3950, 160.5470, 160.4670.

Efficient capture of target analyte on biosensor platforms is a prerequisite for reliable and specific detection of pathogenic microorganisms in a microfluidic chip. Antibodies have been widely used as ligands; however, because of their occasional unsatisfactory performance, a search for alternative receptors is underway. Heat shock protein 60 (Hsp60), a eukaryotic mitochondrial chaperon protein is a receptor for Listeria adhesion protein (LAP) during Listeria monocytogenes infection. This paper reports application of biotinylated Hsp60 as a capture molecule for living (viable) L monocytogenes in a microfluidic environment. Hsp60, immobilized on the surface of streptavidin-coated silicon dioxide exhibited specific capture of pathogenic Listeria against a background of other Listeria species, Salmonella, Escherichia, Bacillus, Pseudomonas, Serratia, Hafnia, Enterobacter, Citrobacter, and Lactobacillus. The capture efficiency of L monocytogenes was 83 times greater than another Listeria receptor, the monoclonal antibody, mAb-C11E9. Additionally, the capture rate was further increased on a Hsp60-coated biochip by 60% when a dielectrophoresis force was applied for 5 min at the beginning of the final 1 h incubation step. Our data show that Hsp60 could be used for specific detection of L monocytogenes on a biochip sensor platform.

Chromatin immunoprecipitation (ChIP) is a powerful and widely applied technique for detecting association of individual proteins with specific genomic regions; the technique requires several complex steps and is tedious. In this paper, we develop a microbead-packed microfluidic chip which eliminates most of the laborious, time-consuming, and skill-dependent processes of the ChIP procedure. A computational fluid dynamics model was established to analyze fluidic behavior in a microbead-packed microchannel. With the use of the new chip, a ChIP procedure was performed to purify the GAPDH (glyceraldehyde 3-phosphate dehydrogenase) gene from human embryonic kidney cells (cell line 293). The ChIP capability of the microfluidic chip was evaluated and compared with that of a commercial assay kit; the precipitation performance of both methods was almost identical as shown by quantitative measurement of DNA. However, our chip offers the advantage of low resin volume, and the experimental time is greatly reduced. In addition, our method is less dependent on individual technical skill. Therefore, we expect that our microfluidic chip-based ChIP method will be widely used in DNA-, gene-, and protein-related research and will improve experimental efficiency.

A new packaging method for microfluidic devices is proposed of polymer overmolding to form a fluidic manifold integrated with the device in a single step. The anticipated advantages of the proposed method of packaging are ease of assembly and low part count, making it suitable for low cost and high volume manufacturing. This paper reports the results of a preliminary investigation into this concept. Glass and silicon inserts of 25 x 20 mm in size, used to represent microfluidic devices, were overmolded in an injection molding process with a range of polymers. The inserts were found to survive the molding process intact. The adhesion between overmold and insert was investigated by subjecting the interface between the overmold and insert surface to a hydrostatic pressure of up to 100 Ibf/[in.sup.2] (6.9 bar). The durability of the interfacial adhesion to hydrolysis was investigated by immersion in water at 50[degrees]C for 24 h before testing. Direct measurements of adhesion strength between polymer and glass were also attempted by tensile tests on lap-jointed samples. The best and most durable adhesion for glass and silicon inserts was found for polyamide (PA) 12, which is a low hygroscopicity PA. The ranking of polymers by their performances in the pressurization tests was consistent with the ranking by the calculated work-of-adhesion values for polymer/glass and polymer/silicon joints. [2006-0211] Index Terms--Bonding, fluidics, interconnections, packaging.

Microscale total analysis systems ([micro]TAS) allow high-throughput analyses by integrating multiple processes, parallelization, and automation. Here we combine unit operations of [micro]TAS to create a device that can perform multidimensional separations using a three-dimensional hybrid microfluidic/nanofluidic device composed of alternating layers of patterned poly(methyl methacrylate) and nanocapillary array membranes constructed from nuclear track-etched polycarbonate. Two consecutive electrophoretic separations are performed, the first being an achiral separation followed by a chiral separation of a selected analyte band. Separation conditions are optimized for a racemic mixture of fluorescein-isothiocynate-labeled amino acids, serine and aspartic acid, chosen because there are endogenous D-forms of these amino acids in animals. The chiral separation is implemented using micellar electrokinetic chromatography using [beta]-cyclodextrin as the chiral selector and sodium taurocholate as the micelle-forming agent. Analyte separation is monitored by dual-beam laser-induced fluorescence detection. After separation in the first electrophoretic channel, the preselected analyte is sampled by the second-stage separation using an automated collection sequence with a zero-crossing algorithm. The controlled fluidic environment inherent to the three-dimensional architecture enables a series of separations in varying fluidic environments and allows sample stacking via different background electrolyte pH conditions. The ability to interface sequential separations, selected analyte capture, and other fluidic manipulations in the third dimension significantly improves the functionality of multilayer microfluidic devices.

A novel passive microfluidic silicon mixer has been designed, optimized and fabricated. The architecture of the mixer consists of a simple 'T' junction, made up by a 20 [micro]m wide by 82 [micro]m deep channel, followed by three repeats of an alcove, each with a triangular obstruction, arranged in a zigzag fashion. Numerical simulations were employed to optimize the geometry, particularly the dimensions of the alcoves, the relative orientation and the spacing between them, and the degree of intrusion associated with them. The simulation results demonstrate that chaotic flow due to recirculation within the alcoves results in transverse velocity that promotes effective fluid mixing. The microfluidic mixer with the simulation-optimized geometry was fabricated with photolithographic techniques and characterized by optical imaging, fluorescence, and Raman microscope spectroscopy. At a sample flow rate of 20 [micro]L/s, the mixer exhibits a short mixing deadtime of ~22 [micro]s and a high mixing efficiency under both low and high viscosity conditions. The alcove-based microfluidic silicon mixer offers unique advantages for its short deadtime and slow sample consumption rate. In addition, it provides a valuable component for laboratory-on-a-chip applications for its ease of development into multiple networks for massively parallel analytical processes.

A microfluidic device designed for electrochemical studies on a microliter scale has been utilized for the examination of impurity levels in ionic liquids (ILs). Halide impurities are common following IL synthesis, and this study demonstrates the ability to quantify low concentrations of halide in a range of ILs to levels of ~5 ppm, even in ILs not currently measurable using other methods such as ion chromatography. To validate the mixer device, the electrochemistry of ferrocene was also examined and compared with spectroscopic and bulk electrochemistry measurements. An automated 'sample preparation, delivery, and calibration' method was developed, and the chip successfully used for linear sweep, cyclic voltammetry (under both quiescent and steady-state flowing conditions), square wave voltammetry, and differential pulse voltammetry. An effective method of electrochemically cleaning the electrodes is also presented.

We describe the fabrication of a controllable microfluidic valve coupled with an electrochemical pump, which has been designed to deliver reagents to an integrated microfluidic biosensing system. Fluid, retained within an insertion reservoir using a stop valve, was pumped using electrochemical actuation, providing a low power, low voltage integrated Laboratory-on-a-Chip for reproducible, small volume fluidic manipulation. The properties of the valve were characterized using both X-ray photoelectron spectroscopy and contact angle measurements, enabling the calculation of the magnitude of the forces involved (which were subsequently verified through experimental measurement). Electrochemical generation of oxygen and hydrogen acted as an on-demand pressure system to force fluid over the stop valve barrier. The process of filling-up the biosensing chamber was characterized in terms of the time to fill, the energy used, and the peak power consumed. The potential of the device was illustrated using a glucose biosensor.

We present a microfluidic device for multiplexed immunoassays based on DNA-directed immobilization. Because of the versatile building blocks used for this technique, it is possible to build up user configurable protein microarrays within the microfluidic system by means of DNA-directed self-assembly, which can be used for immunoassay applications. We demonstrate the performance of our system by parallel detection of cytokines in a multiplex immunoassay, employing silver deposition labeling and optical read-out.

The development of any particular analytical or preparative applications using electrochemical techniques in microfluidic devices requires integration of microelectrodes. This involves detailed predictions for optimizing the design of devices and selecting the best hydrodynamic conditions. For this purpose, we undertook a series of works aimed at a precise investigation of mass transport near electrodes with focus on analytical measurements. Part I of this series (Anal. Chem. 2007, 79, 8502-8510) evaluated the common case of a single microband electrode embedded within a microchannel under laminar flow. The present work (Part 2) investigated the case of a pair of microband electrodes operating either in generator-generator or generator-collector modes. The influence of the confining effect and flow velocity on the amperometric responses was examined on the basis of numerical simulations under steady-state regime. Several situations were identified, each of them corresponding to specific interactions taking place between the electrodes. Related conditions were extracted to establish a zone diagram describing all the situations. These predictions were systematically validated by experimental measurements. The results show that amperometric detections within microchannels can be performed at dual electrodes with higher analytical performances than at single ones.

The ability to rapidly and efficiently isolate specific viruses, bacteria, or mammalian cells from complex mixtures lies at the heart of biomedical applications ranging from in vitro diagnostics to cell transplantation therapies. Unfortunately, many current selection methods for cell separation, such as magnetic activated cell sorting (MACS), only allow the binary separation of target cells that have been labeled via a single parameter (e.g., magnetization). This limitation makes it challenging to simultaneously enrich multiple, distinct target cell types from a multicomponent sample. We describe here a novel approach to specifically label multiple cell types with unique synthetic dielectrophoretic tags that modulate the complex permittivities of the labeled cells, allowing them to be sorted with high purity using the multitarget dielectrophoresis activated cell sorter (MT-DACS) chip. Here we describe the underlying physics and design of the MT-DACS microfluidic device and demonstrate ~1000-fold enrichment of multiple bacterial target cell types in a single-pass separation.

Glass is a desired material for many microfluidics applications. It is chemically resistant and has desirable characteristics for capillary electrophoresis. The process to make a glass chip, however, is lengthy and inconvenient, with the most difficult step often being the bonding of two planar glass substrates. Here we describe a new glass bonding technique, which requires only washing of the glass surfaces with a calcium solution and 1-2 h of bonding at 115[degrees]C. We found calcium uniquely allows for this simple and efficient low-temperature bonding to occur, and none of the other cations we tried (e.g., [Na.sup.+], [Mg.sup.2+], [Mn.sup.3+]) resulted in satisfactory bonding. We determined this bond is able to withstand high applied field strengths of at least up to 4 kV x [cm.sup.-1]. When intense pressure was applied to a fluid inlet, a circular portion of the coverslip beneath the well exploded outward but very little of the glass--glass interface debonded. In combination with the directed hydrofluoric acid etching of a glass substrate using a poly(dimethylsiloxane) (PDMS) etch guide, we were able to make glass chips with better than 90% yield within 6 h. This technique is compatible with inexpensive unpolished glass and is limited in resolution by the PDMS etch guide used and the intrinsic properties of isotropic etching.

We report a new two-axis active optical fiber manipulator for on-chip optical manipulation and detection in microfluidic environment. The system comprising of air chambers, fiber channels, controllable moving walls, and membrane structures were fabricated by using microelectromechanical systems technology. By adjusting air pressures to control the deflection of the pneumatic chambers placed orthogonal to and underneath the fiber channels, accurate alignment of a pair of approximately coaxial optical fibers, which was indicated by maximizing fiber-to-fiber optical-coupling measured in real time, has been achieved. A maximum displacement of a buried fiber as large as 13 [micro]m at an applied pressure of 40 lb/[in.sup.2] for one air chamber has been demonstrated. It was sufficient to accurately align two approximately coaxial optical fibers to maximize the optical coupling efficiency. The maximum coupling efficiency for two single-mode optical fibers facing each other at a distance of 200 [micro]m was measured to be 4.1%. The following features have been successfully demonstrated with this system: 1) stable optical trapping and stretching of a single red blood cell; 2) stable optical trapping of multiple microparticles; 3) optically driven controlled motion of single and multiple microparticles; and 4) integration of a counterpropagating dual-beam trap with single-beam optical tweezers. In addition to optical trapping and manipulation, the proposed device is promising for applications requiring coaxial input/output fibers for in-line optical analysis. Furthermore, it can be easily integrated with other microfluidic devices such as microcapillary electrophoresis channels or microflow cytometers for DNA, protein, and cell analysis. [2007-0182] Index Terms--Microcoupler, microelectromechanical systems (MEMS), microfluidics chip, moving wall, optical detection, optical trappers.

The formation of microscale single- and double-emulsion droplets with various sizes is crucial for a variety of industrial applications. In this paper, we report a new microfluidic device which can actively fine-tune the size of single- and double-emulsion droplets in liquids by utilizing controllable moving-wall structures. Moreover, various sizes of external and internal droplets for double emulsions are also successfully formed by using this device. Three pneumatic side chambers are placed at a T-junction and flow-focusing channels to construct the controllable moving-wall structures. When compressed air is applied to the pneumatic side chambers, the controllable moving-wall structures are activated, thus physically changing the width of the microchannels. The size of the internal droplets at the intersection of the T-junction channel is then fine-tuned due to the increase in the shear force. Then, the internal droplets are focused into a narrow stream hydrodynamically and finally chopped into double-emulsion droplets using another pair of moving-wall structures downstream. For single emulsions, oil-in-water droplets can be actively fine-tuned from 50.07 to 21.80 [micro]m under applied air pressures from 10 to 25 psi with a variation of less than 3.53%. For a water-in-oil single emulsion, droplets range from 50.32 to 14.76 [micro]m with a variation of less than 4.62% under the same applied air pressures. For double emulsions, the sizes of the external and internal droplets can be fine-tuned with external/ internal droplet diameter ratios ranging from 1.69 to 2.75. The development of this microfluidic device is promising for a variety of applications in the pharmaceutical, cosmetics, and food industries. [2007-0230] Index Terms--Double emulsion, flow focusing, microdroplet, microelectromechanical systems (MEMS), microfluidics, moving wall, T-junction.

A novel microfluidic device with an array of analytical chambers was developed in order to perform single-cell-based gene-function analysis. A series of analytical processes was carried out using the device, including electrophorefic manipulation of single cells and electrochemical measurement of gene function. A poly(dimethylsiloxane) microstructure with a microfluidic channel (150 [micro]m in width, 10 [micro]m in height) and an analytical chamber (100 x 20 x 10[micro][m.sup.3]) were fabricated and aligned on a glass substrate with an array of Au microelectrodes. Two microelectrodes positioned in the analytical chamber were employed as a working electrode for the electrophoretic manipulation of cells and electrochemical measurements. A yeast strain (Saccharomyces cerevisiae Y190) carrying the [beta]-galactosidase reporter gene was used to demonstrate that the device could detect the enzyme. Target cells flowing through the main channel were introduced into the chamber by electrophoresis using the ground electrode laid on the main channel. When the cell was treated with 17[beta]-estradiol, gene expression was triggered to produce [beta]-galactosidase, catalyzing the hydrolysis of p-aminophenyl-[beta]-D-galactopyranoside to form p-aminophenol (PAP). The enzymatically generated PAP was detected by cyclic voltammetry and amperometry at the single-cell level in the chamber of the device. Generator-collector mode amperometry was also applied to amplify the current response originating from gene expression in the trapped single cells. After electrochemical measurement, the trapped cells were easily released from the chamber using electrophoretic force.

In the present study, replication of flow-through microfilters in the newly developed microfluidic lab-on-a-chip for blood typing by microinjection molding process was experimentally investigated. As a precise replication of the microfilters was required in order to effectively filter out agglutinated red blood cells, the effects of important processing conditions on the replication of the flow-through microfilters were investigated. By using a mold insert fabricated by a nickel electroplating process and a newly designed mold base, microinjection molding experiments were carried out. A three-dimensional solid model reconstruction method was proposed with the help of specific features characterizing the geometry of microfilters, and accordingly, the feature values of the replicated microfilters were measured by a noncontact optical measurement system. So reconstructed solid modeling result was then used to investigate the effects of various processing conditions, such as a flow rate, a mold temperature, and a packing pressure. Amongst the processing conditions investigated in the present study, the flow rate was found to be the most important one. [DOI: 10.1115/1.2896142] Keywords: microinjection molding, lab-on-a-chip, microfluidic system, microfilter, replication quality

Methods for fabricating poly(methyl methacrylate) microchips using a novel two-stage embossing technique and solvent welding to form microchannels in microfluidic devices are presented. The hot embossing method involves a two-stage process to create the final microchip design. In its simplest form, a mold made of aluminum is fabricated using CNC machining to create the desired microchannel design. In this work, two polymer substrates with different glass transition temperatures ([T.sub.g]), polyetherimide (PEI) and poly(methyl methacrylate) (PMMA), were used to make the reusable secondary master and the final chip. First, the aluminum mold was used to emboss the PEI, a polymeric substrate with [T.sub.g] 216 [degrees]C. The embossed PEI was then used as a secondary mold for embossing PMMA, a polymeric substrate with a lower [T.sub.g] (~105 [degrees]C). The resulting PMMA substrate possessed the same features as those of the aluminum mold. Successful feature transfer from the aluminum mold to the PMMA substrate was verified by profilometry. Bonding of the embossed layer and a blank PMMA layer to generate the microchip was achieved by solvent welding. The embossed piece was first filled with water that formed a solid sacrificial layer when frozen. The ice layer prevented channel deformation when the welding solvent (dichloroethane) was applied between the two chips during bonding. Electrophoretic separations of fluorescent dyes, rhodamine B (Rh B) and fluorescein (FL), were performed on PMMA microchips to demonstrate the feasibility of the fabrication process for microreplication of useful devices for separations. The PMMA microchip was tested under an electric field strength of 705 V [cm.sup.-1]. Separations of the test mixture of Rh B and FL generated 55 500 and 66 300 theoretical plates/meter, respectively.

This paper reports on liquid (deionized water) microlenses that are intrinsically formed and integrated within microfluidics through pneumatic manipulation of fluids inside microchannels. Such microlenses are formed via liquid-air interfaces of liquid droplets, which are pinned at T-shaped junctions of channels. In addition to being tunable in focal lengths (a few hundreds of micrometers to [infinity]) along the microchannels parallel to the substrate used, these microlenses can uniquely be repositioned, removed, and reformed at predetermined locations of the T-shaped junctions within microchannels on demand under pneumatic controls. The design and formation of a microfluidic channel network for the in situ formation are first described. Then, the pneumatic control of the fluids, including the formation, movement, and size control of a lens droplet, is discussed, and the in situ formation processes for single and multiple liquid microlenses are described. The in situ formation processes for a single microlens and a two-lens combination only take tens of seconds, eliminating various conventional microfabrication processes and multiple layers of materials. Finally, the detail of characterization of these microlenses is given. [2007-0176] Index Terms--Liquid microlens, microfluidics, optofluidics, pneumatic control, tunable.

We describe a microfluidic scheme based on remotely controlled self-assembled containers that allows spatio-temporal control over nanoliter-scale chemical reactions. We discuss finite-element simulations of the inductive coupling of radio-frequency radiation to the containers; this coupling enables remotely triggered release of chemical reactants. We demonstrate on-demand chemical release from stationary and mobile containers patterned with different porosities. We also explore reactions between chemicals released from two containers that form liquid products and deposit solid precipitates. We argue that these remotely controlled metallic containers provide an attractive platform for carrying out reconfigurable microfluidics 'without channels.' [2007-0221] Index Terms--Chemistry, microfluidics, radio frequency (RF), remote control, self-assembly.

This paper presents a new miniature reversetranscription polymerase chain-reaction (RT-PCR) system integrating a sample pretreatment device for fast nucleic-acid amplification and diagnosis of viruses and bacteria. In the system, a two-way serpentine-shape (s-shape) pneumatic micropump and a magnetic bioseparator were developed for separation and enrichment of viruses and bacteria. This new bioseparator can also be used as microheating chambers to perform RT-PCR. Taking advantage of the specific interaction between the antibodies on the surface of magnetic beads and the surface antigens on viruses or bacteria, the target virus and bacteria were recognized and further separated and purified from the biosamples by a magnetic field generated by the bioseparator. The target purified virus/bacteria was then lysed to release ribonucleic acid (RNA) or deoxyribonucleic acid (DNA) for the subsequent RT-PCR processes. Experimental results showed that the target virus/bacteria was successfully separated and enriched by the high specificity and selectivity of antibody-conjugated magnetic beads, and the subsequent amplification of RNA/DNA was automatically completed by utilizing the on-chip microheaters and the micro temperature sensor. The high mixing efficiency of the two-way s-shape pump and the rapid heating/cooling rate of the microheating chambers can significantly shorten the pretreatment and diagnosis processes. As a whole, the developed system may provide a powerful platform for sample pretreatment and fast disease diagnosis. [2007-0089] Index Terms--Enrichment, magnetic bead, microelectromechanical systems (MEMS), microfluidics, pretreatment, reverse-transcription polymerase chain reaction (RT-PCR).

Digital (i.e., droplet-based) microfluidics, by the electrowetting-on-dielectric (EWOD) mechanism, has shown great potential for a wide range of applications, such as lab-on-a-chip. While most reported EWOD chips use a series of electrode pads essentially in 1-D line pattern designed for specific tasks, the desired universal chips allowing user-reconfigurable paths would require the electrode pads in 2-D pattern. However, to electrically access the electrode pads independently, conductive lines need to be fabricated underneath the pads in multiple layers, raising a cost issue particularly for disposable chip applications. In this paper, we report the building of digital microfluidic plates based on a printed circuit board (PCB), in which multilayer electrical access lines were created inexpensively using the mature PCB technology. However, due to its surface topography and roughness and resulting high resistance against droplet movement, the as-fabricated PCB surfaces require high (~500 V) voltages unless coated with or immersed in oil. Our goal is the EWOD operations of droplets not only on oil-covered surfaces but also on dry ones. To meet the varying levels of performances, three types of gradually complex post-PCB microfabrication process are developed and evaluated. By introducing land-grid-array sockets in the packaging, a scalable digital microfluidic system with a reconfigurable and low-cost chip is also demonstrated. [2007-0049] Index Terms--Electrowetting, electrowetting on dielectric (EWOD), lab-on-a-chip, microfluidics, micro total analysis system, printed circuit board (PCB).

We present a new approach for fabricating multilayer microfluidic devices in poly(methyl methacrylate). Paraffin wax was used as a phase-changing sacrificial layer to protect microstructures during solvent bonding. Microchannels in the top and bottom pieces were aligned with through-holes in the middle layer, resulting in microchannels that cross one another. No discernible delamination of the layers or leakage between channels was observed at pressures as high as 300 psi. The current versus voltage linearity in the crossover channel indicates that no Joule heating occurs at voltages of at least 2.0 kV. Moreover, a potential in the crossover channel did not affect the current in the separation channel. Rapid and efficient separation of fluorescently labeled amino acids was performed in these devices. Pressurized buffer flow or voltage applied in the crossover channel caused no leakage into or electrical interference with the separation channel. Our results demonstrate that sacrificial layers with solvent bonding can be implemented readily in the fabrication of robust and fluidically complex multilayer polymer microchips. These capabilities should facilitate the development of a new generation of sophisticated microfluidic systems.

This paper reports on the design, fabrication, and characterization of a new class of wideband liquid RF microelectromechanical-system reflective and absorptive switches. A number of different liquids are considered, including mercury, Galinstan, and ultrapure and ionic water. We first briefly review the performance of liquid--metal switches made by mercury and Galinstan. Such switches demonstrate excellent off-state insertion loss of less than 1.3 dB up to 100 GHz (the loss includes a 1500-[micro]m-long line) and on-state isolation of better than 20 dB from 20 to 100 GHz. The main part of this paper, however, focuses on significantly transforming these designs to actually absorb and not reflect the incident power in their on-state, while at the same time maintaining their excellent off-state performance. Absorptive behavior is particularly important for high-power applications. Simpler materials such as water are proven to be very effective for wideband absorptive switches. In particular, three classes of water-based absorptive switches are discussed depending on the level of the signal coupling to water. At 10-40 GHz, the optimal design exhibits off-state insertion loss of less than 1.3 dB, on- and off-state return loss of less than 10 dB, and on-state isolation of 27.5-dB isolation at 40 GHz. Index Terms--Absorptive switch, microfluidics, RF microelectromechanical system (RF MEMS), water, wideband.

The advantages of microanalyzers, usually fabricated in silicon, glass, or polymers, are well-known. The design and construction of fluidic platforms are well-developed areas due to the perfectly established microfabrication technologies used. However, there is still the need to achieve devices that include not only the fluid management system but also the measurement electronics, so that real portable miniaturized analyzers can be obtained. Low-temperature co-fired ceramics technology permits the incorporation of actuators, such as micropumps and microvalves, controlled either magnetically, piezoelectrically, or thermally. Furthermore, electronic circuits can be also easily built exploiting the properties of these ceramics and the fact that they can be fabricated using a multilayer approach. In this work, taking advantage of the possibility of combining fluidics and electronics in a single substrate and using the same fabrication methodology, a chemical microanalyzer that integrates microfluidics, the detection system, and also the data acquisition and digital signal processing electronics is presented. To demonstrate the versatility of the technology, two alternative setups have been developed. In the first one, a modular configuration is proposed. In this case, the same electronic module can be used to determine different chemical parameters by simply exchanging the chemical module. In the second one, the monolithic integration of all the elements was accomplished, allowing the construction of compact and dedicated devices. Chloride ion microanalyzers have been constructed to demonstrate the operability of both device configurations. In all cases, the results obtained showed adequate analytical features.

An array of four sensing microdome optodes (potassium, sodium, calcium, and chloride) was incorporated into a centrifugal microfluidics platform to obtain a multiion analysis system. The behavior of each sensing microdome was in good agreement with a theoretical model describing the response. The selectivity of each optode over common interfering ions was established and was used to identify calibrant solutions that can be employed for the simultaneous calibration of all four optodes without significant cross-interference. The microfluidic platform was designed to facilitate both three-point calibration of the optodes and triplicate analysis of a sample within a single run, which increases the accuracy of the determination. The optimized microfluidic system was used to determine simultaneously the concentration of potassium, sodium, calcium, and chloride in aquarium water (with the composition of Lake Tanganyika water) with less than 6% error. The simple process of fabrication of these microdomes and their incorporation into a centrifugal microfluidic platform should facilitate the development of portable ion-sensing analysis systems.

In this paper, we present a simple, rapid, and low-cost procedure for fabricating glass microfluidic chips. This procedure uses commercially available microscopic slides as substrates and a thin layer of AZ 4620 positive photoresist (PR) as an etch mask for fabricating glass microfluidic components, rather than using expensive quartz glasses or Pyrex glasses as substrates and depositing an expensive metal or polysilicon/amorphous silicon layer as etch masks in conventional method. A long hard-baking process is proposed to realize the durable PR mask capable of withstanding a long etching process. In order to remove precipitated particles generated during the etching process, a new recipe of buffered oxide etching with addition of 20% HCl is also reported. A smooth surface microchannel with a depth of more than 110 [micro]m is achieved after 2 h of etching. Meanwhile, a simple, fast, but reliable bonding process based on UV-curable glue has been developed which takes only 10 min to accomplish the efficient sealing of glass chips. The result shows that a high bonding yield (~100%) can be easily achieved without the requirement of clean room facilities and programmed high-temperature furnaces. The presented simple fabrication process is suitable for fast prototyping and manufacturing disposable microfluidic devices. [2007-0032] Index Terms--Microfabrication, microfluidic devices, UV bonding, wet etching.

This paper presents a novel microfluidic photomasking strategy. Laminar microfluidic streaming is exploited to achieve photomasking with dynamic spatial control. Two light-absorbing streams hydrodynamically focus a transparent stream to define a line of light transmission. A device incorporating two such microfluidic layers, which are aligned orthogonally, enables light transmission only where the transparent streams overlap. Control of fluid flow in the microfluidic layers enables dynamic spatial control of the exposed region. The dynamic microfluidic photomasking strategy is tested through transmitted light microscopy and applied to microfabrication via photoresist patterning. When applied to microfabrication in a frontal photopolymerization mode, this method affords both planar and depthwise control of feature geometry. [2006-0088] Index Terms--Focusing, hydrodynamics, microfluidics, optofluidics, photomasking.

This paper presents a sequential dielectroliboretic field-flow separation method for particle populations using a chip with a 3-D electrode structure. A unique characteristic of our chili is that the walls of the microfluidic channels also constitute the device's electrodes. This property confers the opportunity to use the electrodes' shape to generate not only the electric field gradient required for dielectrophoretic force but also a fluid velocity gradient. This interesting combination gives rise to a new solution for the dielectrophoretic separation of two particle populations. The proposed sequential field-flow separation method consists of four steps. First, the microchannel is hilled with the mixture of the two populations of particle. Second, the particle populations are trapped in different locations of the microfluidic channels. The population, which exhibits positive dielectrophoresis (DEP), is trapped in the area where the distance between the electrodes is the minimum, while the other population that exhibits negative DEP is trapped in locations of maximum distance between electrodes. In the next step, increasing the flow in the microchannels will result in an increased hydrodynamic force that sweeps the cell population trapped by positive DEP out of the chip. In the last step, the electric field is removed, and the second population is swept out and collected at the outlet. For theoretical and experimental exemplification of the separation method, a population of viable and nonviable yeast cells was considered. [2006-0157] Index Terms--Cell separation, dielectrophoresis (DEP), microfluidic device, 3-D silicon electrodes.

The evolution process and turbulence properties of a transversely oscillating flow induced by a fluidic oscillator are studied in a gravity-driven water tunnel. A planar jet is guided to impinge a specially designed crescent surface of a target blockage that is enclosed in a cavity of a fluidic oscillator. The geometric configuration of the cavity transforms the inherent stability characteristics of the jet from convective instability to absolute instability, so that the jet precedes the persistent back and forth swinging in the cavity. The swinging jet is subsequently directed through two passages and issued alternatively out of the fluidic oscillator. Two short plates are installed near the exits of the alternatively issuing pulsatile jets to deflect the jets toward the central axis. The deflected jets impinge with each other and form a pair of counter-rotating vortices in the near wake of the oscillator with a stagnation point at the impingement point. The stagnation point of the counter-rotating vortex pair moves back and forth transversely because of the phase difference existing between the two issued jets. The merged flow evolving from the counter-rotating vortices formed by the impingement of the two pulsatile jets therefore presents complex behavior of transverse oscillation. The topological models corresponding to the flow evolution are constructed to illustrate the oscillation process of the oscillating flow. Significant momentum dispersion and large turbulence intensity are induced by the transverse oscillation of the merged flow. The statistical turbulence properties show that the Lagrangian integral time and length scales of the turbulence eddies (the fine-scale structure) produced in the oscillating flow are drastically reduced. [DOI: 10.1115/1.2746905]

In this paper, we present the development of disposable single-use microfluidic pumps entirely based on a straightforward wafer-level fabrication scheme, which allows for precise integrated active dosing in the microliter range. To accomplish stroke-lengths needed for microliter-range applications, we utilize a new method of bending of a unimorph-composite-actuator film. The unimorph composite actuator consists of a temperature-sensitive silicone elastomer composite, i.e., polydimethylsiloxane, with incorporated expandable microspheres. The fabricated micropumps successfully demonstrated precise liquid-volume control, both at low and high flow rates, and show a standard deviation of 6.7% for consecutive pump experiments. Moreover, the method of fluorescent thermometry was used to measure the thermal load on liquid volumes dispensed with the micropumps. The liquid temperature reaches a maximum of 50[degrees]C during the operation. The presented fully integrated single-use micropumps are electrically controllable, do not require external means for liquid actuation, are made of low-cost materials only, and might potentially be used in drug-delivery applications. [2006-0277] Index Terms--Fabrication, fluidics, micropumps, wafer-scale integration.

A contactless microfluidic dosing system based on actuation by an electrochemical reaction has been designed and tested. The device is composed of two chambers separated by a polydimethylsiloxane (PDMS) plate. One chamber is loaded with the liquid sample to be dosed while the electrochemical reaction occurs in the second chamber. Due to the strict separation between the reaction responsible of the actuation and the sample, the latter is preserved from contamination and/or electrochemical modification. Silicon master microstructures were fabricated using cleanroom technologies and subsequently replicated into the PDMS. A full microsystem has been fabricated and tested. Microliter sample solutions can be easily and reproducibly dispensed in a time frame of a few tens of seconds. [2006-0244] Index Terms--Contactless dispensing, dosing, electrochemical actuation, microfluidics, polydimethylsiloxane (PDMS) replication, silicon microfabrication.

To measure a microflow rate and to accelerate the reaction between proteins by an unbalanced impingement of feedback flow, we have proposed and verified the design of a self-flapping microfluidic oscillator. Three specific features--the large aspect ratio of the micronozzle, the structure of the sudden-expansion inlet, and the asymmetric feedback channels--are developed to induce stable oscillation. The large aspect ratio of the micronozzle diminishes the influence of viscous force, and the inlet structure triggers flow instability. The conjunction of both factors promotes the occurrence of the Coanda effect, and initiates oscillation. The asymmetric feedback channels produce an unbalanced impingement of the inlet flow, thus reinforcing the initial oscillation to become stably periodic. Beyond the function of a microflowmeter, the oscillatory characteristics are applicable to accelerate the biochemical reaction between two fluorescent proteins, B-phycoerythrin and an allophycocyanin alpha subunit. With fluorescence induced with a laser, we detected the proteins at a specific wavelength to define the region of interaction caused by the oscillatory motions, which clearly enhances the rate of reaction of these fluids. To focus on the reaction phenomenon of twin fluids, we demonstrated biotin-streptavidin binding that was detected via a fluorescence-resonance-energy-transfer (FRET) pair of fluorescent proteins. The FRET signal demonstrated conclusively that the biochemical reaction was promoted through the oscillatory function. [2006-0233] Index Terms--Fluorescence resonance energy transfer (FRET), microflowmeter, microfluidic oscillator, [mu]-laser-induced-fluorescence (LIF).

An approach for interfacing nanometer-scale capillaries (pores) that includes an affinity element for molecular recognition, capture, and release of a preselected analyte to microfluidic devices is described.

This paper describes the design, fabrication, and testing of microfluidic devices enabled by electrodes embedded vertically in the side walls of SU-8 microchannels. With vertical electrodes on the side walls, one can generate higher lateral electrical fields uniform along the vertical direction in the channel (perpendicular to the substrate). By designing the electrode shapes and configurations, uniform and nonuniform electrical fields in the lateral (planar) directions can be applied to manipulate flow or particles in microchannels for switching or sorting applications. The uniform field is demonstrated in a magnetohydrodynamic (MHD) microfluidic device for directing cells to different channel outlets while the nonuniform field is demonstrated in the generation of dielectrophoresis (DEP) forces for microbead focusing. Metal electrodes are fabricated by electroplating to form vertical electrodes aligned with the channel walls. The multilayer SU-8 lithography technique enables the four walls of the channel to he all SU-8. The thin precoated SU-8 layer on the substrate improves structure integrity of the SU-8 microchannels. The mechanical flexibility of PDMS compensates for the surface nonuniformity from the previous patterning steps to conformably call the channel. The ability to integrate versatile electrodes design broadens the realm of electrical control and sensing for microfluidic applications. [2006-0106] Index Terms--Conformably bonding, dielectrophoresis (DEP), electroplating, flexible PDMS, lateral manipulation, magnetohydrodynmic (MHD), microfluidics, multilayer SU-8, surface topology, vertical electrodes.

This paper presents a self-adaptive microfluidic variable resistor that accomplishes passive control of liquid flows by exploiting the large compliance of elastomeric polymers such as polydimethylsiloxane. The device features a compliant microstructure embedded in a microchannel that consists of a flexible thin flap and a stiff stopper located in close proximity. The shape and size of the gap between the flap and stopper vary with the applied pressure, resulting in small resistance with respect to forward flow and large resistance with respect to reverse flow. The variable flow resistor shows an interesting diode behavior because its flow resistance is drastically different for different directions of applied pressures and is self-adaptive in the sense that its flow resistance varies with reverse pressure in such a way that the resulting flow rate remains a constant. That is, the flow resistor can be used as a check valve and, more importantly, a passive flow regulator. Prototype devices have demonstrated regulation of nearly constant water flow rates from 0.21 to 1.2 ml/min, with variations less than 3%, under driving pressures that vary significantly from 100 to over 200 kPa. Three-dimensional fluid-structure interaction simulations have been performed to investigate the interactions between the fluid flow and flap deflection. The simulation results agree with the experimental data and provide insight into the device characteristics. Based on its single-layer planar configuration, passive operation over a large pressure range, and use of a flexible and inexpensive polymer, this self-adaptive variable flow resistor is well suited to flow control in lab-on-a-chip systems. [2006-0129] Index Terms--Flow regulator, fluid structure interaction (FSI), fluidic resistor, microfluidics, valve.

This paper describes a novel composite actuator for controlled liquid actuation in microsystems which is based on a thermally responsive elastomer. The composite actuator consists of expandable microspheres incorporated in a polydimethylsiloxane (PDMS) matrix and entails the merits of both PDMS and expandable microspheres. The main characteristic of the composite actuator is to expand upon heat. The expansion is irreversible and the relative volume increase is measured up to 270% of its original volume after heating to 80 [degrees]C. The composite was used to fabricate single-use microfluidic pumps and valves. We show the displacement of liquids in the range of nanoliters even against counter pressures up to 100 kPa. Moreover, liquid flow in microchannels was entirely blocked by means of the integrated valves. The valves can withstand pressures up to 140 kPa. The devices are fabricated using low-cost materials only, and the composite actuator allows using wafer-level processing. The fluidic components based on the novel composite are highly integrable and do not require external actuators. Index Terms--Fluidics, microactuators, micropumps.

Evaporation is a critical problem when handling submicroliter volumes of fluids. This paper characterizes this problem as it applies to microfluidic cell culture in poly(dimethylsiloxane) (PDMS) devices and provides a practical solution. Evaporation-mediated osmolality shifts through PDMS membranes with varying thicknesses (10, 1, 0.2, or 0.1 mm) were measured over 96 h. Even in humidified cell culture incubators, evaporation through PDMS and associated shifts in the osmolality of culture media was significant and prevented mouse embryo and human endothelial cell growth and development. A simple diffusion model, where the measured diffusion coefficient for PDMS matches reported values of ~[10.sup.-9] [m.sup.2]/s, accounts for these evaporation and osmolality shifts. To overcome this problem, a PDMS-parylene-PDMS hybrid membrane was developed that greatly suppresses evaporation and osmolality shifts, yet possesses thinness and the flexibility necessary to interface with deformation-based microfluidic actuation systems, maintains the clarity for optical microscopy, and enables the successful development of single-cell mouse embryos into blastocysts under static conditions and culture of human endothelial cells under dynamic recirculation of submicroliter volumes of media. These insights and methods demonstrated specifically for embryo and endothelial cell studies will be generally useful for understanding and overcoming evaporation-associated effects in microfluidic cell cultures.

A microfluidic chip that allows for the continuous monitoring of cellular secretions from multiple independent living samples was developed. Performance of the device was characterized through the analysis of insulin secretion from islets of Langerhans. The chip contained four individual channel networks, each capable of performing electrophoresis-based immunoassays of the perfusate from islets. In the networks, islets were housed in a chamber that was continuously perfused with pressure-driven biological media at 0.6 [micro]L [min.sup.-1]. Electroosmosis was used to pull perfusate containing secreted insulin into 4-cm-long reaction channels where it mixed with fluorescein isothiocyanate-labeled insulin and anti-insulin antibody for 60 s. The reaction streams were sampled at 6.25-s intervals and analyzed in parallel using an on-chip capillary electrophoresis separation with laser-induced fluorescence detection by a scanning confocal microscope. The limit of detection for insulin was 10 nM. The device was used to complete over 1450 immunoassays of biological samples in less than 40 min, allowing the parallel monitoring of insulin release from four islets every 6.25 s.

A 16-channel microfluidic chip with an integrated contact conductivity sensor array is presented. The microfluidic network consisted of 16 separation channels that were hot-embossed into polycarbonate (PC) using a high-precision micromilled metal master. All channels were 40 [micro]m deep and 60 [micro]m wide with an effective separation length of 40 mm. A gold (Au) sensor array was lithographically patterned onto a PC cover plate and assembled to the fluidic chip via thermal bonding in such a way that a pair of Au microelectrodes (60 [micro]m wide with a 5 [micro]m spacing) was incorporated into each of the 16 channels and served as independent contact conductivity detectors. The spacing between the corresponding fluidic reservoirs for each separation channel was set to 9 mm, which allowed for loading samples and buffers to all 40 reservoirs situated on the microchip in only five pipetting steps using an 8-channel pipettor. A printed circuit board (PCB) with platinum (Pt) wires was used to distribute the electrophoresis high-voltage to all reservoirs situated on the fluidic chip. Another PCB was used for collecting the conductivity signals from the patterned Au microelectrodes. The device performance was evaluated using microchip capillary zone electrophoresis ([micro]-CZE) of amino acid, peptide, and protein mixtures as well as oligonucleotides that were separated via microchip capillary electrochromatography ([micro]-CEC). The separations were performed with an electric field (E) of 90 V/cm and were completed in less than 4 min in all cases. The conductivity detection was carded out using a bipolar pulse voltage waveform with a pulse amplitude of [+ or -] 0.6 V and a frequency of 6.0 kHz. The conductivity sensor array concentration limit of detection (SNR = 3) was determined to be 7.1 [micro]M for alanine. The separation efficiency was found to be 6.4 x [10.sup.4], 2.0 x [10.sup.3], 4.8 x [10.sup.3], and 3.4 x [10.sup.2] plates for the [micro]-CEC of the oligonucleotides and [micro]-CZE of the amino acids, peptides, and proteins, respectively, with an average channel-to-channel migration time reproducibility of 2.8%. The average resolution obtained for [micro]-CEC of the oligonucleotides and [micro]-CZE of the amino acids, peptides, and proteins was 4.6, 1.0, 0.9, and 1.0, respectively. To the best of our knowledge, this report is the first to describe a multichannel microchip electrophoresis device with integrated contact conductivity sensor array.

A facile nonlithographic method for expedient fabrication of microfluidic devices of poly-(dimethylsiloxane) is described. Using the method, microfluidic devices are fabricated for detection of bacterial spores on the basis of enhancement of the emission of terbium (III) ions.

A novel polydimethyl siloxane (PDMS) microfluidic spotter system has been developed for the patterning of surface microarrays that require individually addressing each spot area and high probe density. Microfluidic channels are used to address each spot region, and large spot arrays can be addressed in parallel. Fluorescence intensity measurement of dye-spotted samples compared to control and pipetted drops demonstrated a minimum of three-fold increase in dye surface density compared to pin-spotted dyes. Surface plasmon resonance (SPR) measurement of protein-spotted samples as compared to pin-spotted samples demonstrated an 86-fold increase in protein surface concentration. The spotting system has been applied successfully to protein microarrays for SPR applications, in both a 12-spot linear and 48-spot two-dimensional (2-D) array format. This novel spotter system can be applied to the production of high-throughput arrays in the fields of genomics, proteomics, immunoassays, and fluorescence or luminescence assays. [1565] Index Terms--Microarray, microfluidic, polydimethyl siloxane (PDMS), spotter.

We describe the design, fabrication, and operation of two types of flow cytometers based on microfluidic devices made of a single cast of poly(dimethylsiloxane). The stream of particles or cells injected into the devices is hydrodynamically focused in both transverse and lateral directions, has a uniform velocity, and has adjustable diameter and shape. The cytometry system built around the first microfluidic device has fluorescence detection accuracy comparable with that of a commercial flow cytometer and can analyze as many as 17 000 particles/s. This high-throughput microfluidic device could be used in inexpensive stand-alone cytometers or as a part of integrated microanalysis systems. In the second device, a stream of particles is focused to a flow layer of a submicrometer thickness that allows imaging the particles with a high numerical aperture microscope objective. To take long-exposure, low-light fluorescence images of live cells, the device is placed on a moving stage, which accurately balances the translational motion of particles in the flow. The achieved resolution is comparable to that of still micrographs. This high-resolution device could be used for analysis of morphology and fluorescence distribution in cells in continuous flow.

This paper describes the combined use of controlled nanoassembly and microfabrication (photolithography) to construct multi-walled, carbon, nanotube-based fluidic devices. The nanoassembly technique utilizes dielectrophoresis to position individual nanotubes across the gap between two electrodes patterned on a wafer. The dielectrophoretic migration process was studied theoretically and experimentally. Once a tube had been trapped between a pair of electrodes, photoresist was spun over the wafer and developed to form microfluidic interfaces. Liquid condensation in and evaporation from the nanotubes were observed with optical microscopy. The nanotube-based fluidic devices can be used for studies of fluid transport under extreme confinement and as sensitive sensors.

We present a novel approach to perform high-sensitivity NMR imaging and spectroscopic analysis on microfluidic devices. The application of NMR, the most information-rich spectroscopic technique, to microfluidic devices remains a challenge because the inherently low sensitivity of NMR is aggravated by small fluid volumes leading to low NMR signal and geometric constraints resulting in poor efficiency for inductive detection. We address the latter by physically separating signal detection from encoding of information with remote detection. Thereby, we use a commercial imaging probe with sufficiently large diameter to encompass the entire device, enabling encoding of NMR information at any location on the chip. Because large-diameter coils are too insensitive for detection, we store the encoded information as longitudinal magnetization and flow it into the outlet capillary. There, we detect the signal with optimal sensitivity, using a solenoidal microcoil, and reconstruct the information encoded in the fluid. We present a generally applicable design for a detection-only microcoil probe that can be inserted into the bore of a commercial imaging probe. Using hyperpolarized [sup.129]Xe gas, we show that this probe enables sensitive reconstruction of NMR spectroscopic information encoded by the large imaging probe while keeping the flexibility of a large coil.

This paper reports a protocol that improves the resolving power of isoelectric focusing (IEF) in a polymeric microfluidic chip. This method couples several stages of IEF in series by first focusing proteins in a straight channel using broad-range ampholytes and then refocusing segments of the first channel into secondary channels that branch from the first one at T-junctions. Experiments demonstrate that several fluorescent proteins that had focused within a segment of the straight channel in the first stage were refocused at significantly higher resolution due to the shallower pH gradient and higher electrical field gradient. Two variants of green fluorescent protein from the second-stage IEF fractionation were further separated in a third stage. Three stages of IEF were completed in less than 25 min at electric field strengths ranging from 50 to 214 V/cm.

A stable, rugged, and easy-to-use microfluidic platform has been developed and evaluated. The system is based on multilayer silicone elastomer chips with integrated flow channels and active components, such as pressure-actuated valves. The silicone chips and stainless steel interconnect tubes are embedded in a block of an epoxy resin to give the chip and the interconnects outstanding mechanical stability. Full optical accessibility of the chip for demanding optical detection and manipulation applications, such as fluorescence and bright-field microscopy and optical tweezing, is achieved through the use of an optically transparent epoxy resin and microscope cover glasses as the packaging materials. Furthermore, this packaging technique uses a purely mechanical seal between the elastomer and cover glass, enabling applications that use chemically functionalized glass surfaces as the bottom of the flow channels. In addition, a socket was developed in which the microfluidic chips can be plugged in to provide all external connections for reagent delivery and pressure control of the integrated valves. The utility of these devices is demonstrated by showing that single DNA molecules can be attached to protein-coated walls of the flow channels and manipulated with optical tweezers.

A new preconcentration device was developed for analysis of proteins by capillary electrophoresis (CE). The micro-fluidic device uses an electric field to capture proteins that pass through the system. The capture zone is maintained in the flow stream by the interaction between hydrodynamic and electrical forces. The device consists of a flow channel made of PEEK tubing with two electrical junctions, each of which is covered with a conductive membrane. A syringe pump provides the flow stream and also allows the injection of up to 13.5 [micro]L of a dilute sample. The system can be easily connected to a CE device post-capture for off-line preconcentration of proteins. For the proteins used in this study, preconcentration factors up to 40-fold can be achieved. CE detection limits for bovine carbonic anhydrase, [alpha]-lactalbumin and [beta]-lactoglobulins A and B were in the nanomolar range using UV detection at 200 nm. Preconcentration is dependent on both time and initial protein concentration. We show the possibility of using an off-line fluidic preconcentrator device employing counterflow capillary electrophoresis with minimum sample manipulation, achieving detection limits similar to on-line approaches.

The design and operation of a microfluidic device for sample preparation in MALDI mass spectrometry of peptides and proteins is described. It is particularly useful for proteomics applications and for mass determination of proteins in salt- and detergent-containing solutions. The system consists of a flow channel with two conductive areas or electrical junctions where proteins and peptides are retained by means of an electric field. The microfluidic device is made of PEEK tubing, and the junctions are covered with a conductive polymeric membrane. A syringe pump connected to the device produces a flow stream, and injection of sample is carried out manually via hydrodynamic pressure. Proteolytic peptides and intact proteins in salt- and detergent-containing acidic media were captured at the cathode junction followed by exchange of the original solution to a solvent suitable for subsequent mass spectrometry. Using this principle, a significant desalting effect was obtained for tryptic peptides in mass-mapping experiments. Protein sequence coverages were high (up to 40%) at subpicomole levels with results better than those obtained using reversed-phase solid-phase extraction. In contrast to the latter technique, the microfluidic device has the capacity to efficiently remove detergents such as CHAPS before peptide mapping and protein analysis.