Use of directly molded poly(methyl methacrylate) channels for microfluidic applications

A direct molding method for creating a homogeneous, polymer microfluidic channel is presented. By utilizing capillary rise and subsequent absorption of poly(methyl methacrylate) (PMMA) solution into a solvent-permeable poly(dimethyl siloxane) (PDMS) mold, various circular or elliptic polymer microchannels were fabricated without channel bonding and additional surface modification processes. In addition, the channel diameter was tunable from several micrometres to several hundreds of micrometres by controlling concentration and initial amount of polymer solution for a given PDMS mold geometry. The molded PMMA channels were used for two applications: blocking absorption of Rhodamine B dye and constructing artificial endothelial cell-cultured capillaries. It was observed that the molded PMMA channels effectively prevented absorption and diffusion of Rhodamine molecules over 5 h time span, demonstrating approximately 40 times higher blocking efficiency as compared to porous PDMS channels. Also, calf pulmonary artery endothelial cells (CPAEs) adhered, spread, and proliferated uniformly within the molded microchannels to form near confluency within 3 days and remained viable at day 6 without notable cell death, suggesting high biocompatibility and possibility for emulating in vivo-like three-dimensional architecture of blood vessels.     

Fabrication of circular microfluidic channels by combining mechanical micromilling and soft lithography

The fabrication of microfluidic channels with complex three-dimensional (3D) geometries presents a major challenge to the field of microfluidics, because conventional lithography methods are mainly limited to rectangular cross-sections. In this paper, we demonstrate the use of mechanical micromachining to fabricate microfluidic channels with complex cross-sectional geometries. Micro-scale milling tools are first used to fabricate semi-circular patterns on planar metallic surfaces to create a master mold. The micromilled pattern is then transferred to polydimethylsiloxane (PDMS) through a two-step reverse molding process. Using these semi-circular PDMS channels, circular cross-sectioned microchannels are created by aligning and adhering two channels face-to-face. Straight and serpentine-shaped microchannels were fabricated, and the channel geometry and precision of the metallic master and PDMS molds were assessed through scanning electron microscopy and non-contact profilometry. Channel functionality was tested by perfusion of liquid through the channels. This work demonstrates that micromachining enabled soft lithography is capable of fabricating non-rectangular cross-section channels for microfluidic applications. We believe that this approach will be important for many fields from biomimetics and vascular engineering to microfabrication and microreactor technologies.     

A Microchip‐Based System for Immobilizing PC 12 Cells and Amperometrically Detecting Catecholamines Released After Stimulation with Calcium

In this paper, we describe a microchip‐based system for amperometrically monitoring the amount of catecholamines released from rat pheochromocytoma (PC 12) cells. Key to this system is a novel, yet simple method for the immobilization of PC 12 cells in poly(dimethylsiloxane) (PDMS)‐based microchannels. The procedure involves selectively coating microchannels with collagen followed by introduction of PC 12 cells over the PDMS structure, with the cells being immobilized only on the coated portion of the channels. The cell‐coated microchannels can then be reversibly sealed to a glass plate containing electrodes for amperometric detection, resulting in an immobilized cell reactor with integrated microelectrodes. Nafion‐coated microelectrodes made by micromolding of carbon inks were used to measure calcium‐induced catecholamine release from the cells. Varying concentrations of PC 12 cells immobilized in the microchannels led to a catecholamine release ranging from 20 to 160 μM when the cells were stimulated with a calcium solution. This microchip approach leads to a three‐dimensional culture that can be used with this or other cells lines to study the effect of external stimuli on neurotransmitter release.     

Role of surface anchoring and geometric confinement on focal conic textures in smectic-A liquid crystals

A high surface area-to-volume ratio in microchannels increases the importance of surface interactions within them. In layered liquids, such as smectic liquid crystals, surface interactions play an important role in the formation of defect textures. We use 8CB liquid crystal, which is in the smectic-A phase at room temperature, as a model layered liquid. PDMS surfaces can be tuned to be hydrophilic or hydrophobic, and due to the nature of liquid crystalline molecules, we show that this results in planar or homeotropic anchoring conditions, respectively. In a confined system, contrary to the bulk, generated defects cannot grow freely. In the present work, we show that the confinement offered by PDMS microchannels along with the capability of creating mixed anchoring conditions within them results in the formation of particular ordered defect textures through increased surface interactions in smectic-A liquid crystals. Our observations imply that microscale confinement is useful for controlling the size, size distribution, and packing structure of microscale defect structures within these materials. In addition, we show that by placing a droplet of smectic-A liquid crystal on a PDMS surface containing microscale parallel cracks, ordered focal conic defects form between two adjacent cracks. The distance between two adjacent cracks dictates the size of the defects. These observations could lead to useful ideas for exploring new technologies for flexible optical devices or displays that utilize smectic-A liquid crystals.     

Fabrication of PDMS/glass microchips by twofold replication of PDMS and its application in genetic analysis

In this paper, we describe a simple method for fabrication of high quality poly(dimethylsiloxane) (PDMS)/glass microchip by twofold replica molding of PDMS. This technique first served to transfer the negative microchannels from the glass template to the PDMS substrate as a master, and then this PDMS master with positive microchannels was used to replicate the PDMS replica with negative microchannels. Finally, the PDMS replica was bound to a glass sheet by UV radiation. The fabricated microchips were successfully applied for the detection of C677T mutation from the human methylenetetrahydrofolate reductase gene.     

Stabilization of two-phase octanol/water flows inside poly(dimethylsiloxane) microchannels using polymer coatings

In this paper we present our first results on the realization of stable water/octanol, two-phase flows inside poly(dimethylsiloxane) (PDMS) microchannels. Native PDMS microchannels were coated with high molecular weight polymers to change the surface properties of the microchannels and thus stabilize the laminar flow profile. The polymers poly(2-hydroxyethyl methacrylate), poly(vinyl pyrrolidone), poly(ethylene oxide), poly(ethylene glycol), and poly(vinyl alcohol) were assessed for their quality as stabilization coatings after deposition from flowing and stationary solutions. Additionally, the influence of coating the microchannels homogeneously with a single kind of polymer or heterogeneously with two different polymers was investigated. From the experimental observations, it can be concluded that homogeneous polymer coatings with poly(2-hydroxyethyl methacrylate) and poly(vinyl pyrrolidone) led to the effective stabilization of laminar water/octanol flows. Furthermore, heterogeneous coatings led to two-phase flows which had a better-defined and more stable interface over long distances (i.e., 40-mm-long microchannels). Finally, the partitioning of fuchsin dye in the coated microchannels was demonstrated, establishing the feasibility of the use of the polymer-coated PDMS microchannels for determination of logP values in laminar octanol/water flows.     

Functionality and stability of heparin immobilized onto poly(dimethylsiloxane)

Poly(dimethylsiloxane) (PDMS) has become an attractive material when working in the field of microfluidics, mainly because of the rapid prototyping process it involves. The increased surface volume ratio in microchannels makes the interaction between sample and material surface highly important, evident when handling complex biological samples such as plasma or blood. This study demonstrates a new grade of non-covalent heparin surface that adds efficient anticoagulant property to the PDMS material. The surface modification is a simple and fast one-step process performed at neutral pH, optimal when working with closed microsystems. The heparin formed a uniform and functional coating on hydrophobic PDMS with comparatively high level of antithrombin-binding capacity. In addition, long-term studies revealed that the immobilized heparin was more or less stable in the microchannels over a time of three weeks. Recalcified plasma in contact with native PDMS showed complete coagulation after 1h, while no fibrin formation was detected in plasma incubated on heparin-coated PDMS within the same time. In conclusion, we see the heparin coating developed and evaluated in this study as a tool that greatly facilitates the use of PDMS in microfluidics dealing with plasma or blood samples.     

Internal modification of poly(dimethylsiloxane) microchannels with a borosilicate glass coating

We report on an original technique for the in situ coating of poly(dimethylsiloxane) (PDMS) microchannels with borosilicate glass, starting from an active nonaqueous and alkali-free precursor solution. By chemical reaction of this active solution inside the microchannel and subsequent thermal annealing, a protective and chemically inert glass borosilicate coating is bonded to the PDMS. Attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy and nuclear magnetic resonance spectroscopy of the active solution show that it is composed of a silicon oxide network with boron connectivity. Thermal gravimetric analysis demonstrates the absence of organic content when curing is done above 150 degrees C. The borosilicate nature of the glass coating covalently bonded to the PDMS is demonstrated using ATR-FTIR spectroscopy and X-ray photoelectron spectroscopy. Atomic force microscopy and scanning electron microscopy show a smooth and crack-free coating. The latter is used as an efficient protective barrier against diffusion in PDMS of fluorescent rhodamine B dye that is dissolved either in water or in toluene. Moreover, the coating prevents swelling and consequent structural damage of the PDMS when the latter is exposed to harsh chemicals such as toluene.     

Rapid fabrication of microchannels using microscale plasma activated templating (microPLAT) generated water molds

Poly(dimethylsiloxane) (PDMS) is a common material used in fabricating microfluidic devices. The predominant PDMS fabrication method, soft lithography, relies on photolithography for fabrication of micropatterned molds. In this technical note, we report an alternative molding technique using microscale PLasma Activated Templating (microPLAT). The use of photoresist in soft lithography is replaced by patterned water droplets created using microPLAT. When liquid PDMS encapsulates patterned water and then solidifies, the cavities occupied by water become structures such as microchannels. Using this method, device fabrication is less time consuming, more cost efficient and flexible, and ideal for rapid prototyping. An additional important feature of the water-molding process is that it yields structural profiles that are difficult to achieve using photolithography.     

Electroosmotic flow in poly(dimethylsiloxane) microchannels

This paper reports on the study of electroosmotic flow (EOF) in poly(dimethylsiloxane) (PDMS) microchannels on the basis of indirect amperometric detection method. Gradual increase of EOF rate in freshly prepared PDMS microchannels was observed with the running buffer of phosphate buffer solution (PBS). With the same concentration (10 mM) of PBS containing different cations and the same pH value (7.0) and, the time of the stable EOF in PDMS microchannels under the applied separation voltage of 1000 V was 49.8 s (Li+ -PBS), 57.1 s (Na+ -PBS), 91 s (K+ -PBS), respectively. Meanwhile, the different adsorption of cations (Li+, Na+ and K+) on hydrophobic PDMS wall was observed through their separation in PDMS microchannels. Such experimental results demonstrated that the EOF in PDMS microchannels came from the cations and anions adsorbed on PDMS wall. This study would not only help us understand the surface state of PDMS, but also provide a useful guidance for establishing the effective surface modification methods in PDMS microchip CE.     

Characterization of molecular transport in poly(dimethylsiloxane) microchannels for electrophoresis fabricated with synchrotron radiation-lithography and UV-photolithography

In the present paper, a study was undertaken of molecular transport in ploy(dimethylsiloxane) microchannels that were fabricated by ultraviolet (UV)-photolithography and synchrotron radiation (SR)-lithography characterized and compared for microchip capillary electrophoresis by evaluating in-channel molecular dispersion. A fluorescent tag, sulforhodamine B was used as the probing molecule. It was found that microchannels made by SR-lithography fabrication were superior to those made by UV-photolithography fabrication in terms of molecular transport performance. A deep insight into surface conditions characterized by scanning electron microscopy suggested it was related to the difference in surface roughness. Chromatographic retention in electropherograms further supported such a conclusion, which depended on the phase ratio of the channel surface. The results revealed for PDMS microchannels in this work were in good agreement with the phenomenon found for glass microchannels in the literature.     

Fabrication of complex multilevel microchannels in PDMS by using three-dimensional photoresist masters

This paper demonstrates a new method of implementing complex microchannels in PDMS, which is simply constructed using three-dimensional photoresist structures as a master mold for the PDMS replica process. The process utilizes UV-insensitive LOR resist as a sacrificial layer to levitate the structural photoresist. In addition, the thickness of photoresist structures can be controlled by multi-step UV exposure. By using these techniques, various three-dimensional photoresist structures were successfully implemented, including the recessed cantilevers, suspended bridges, and the complex plates with micro-pits or micro-villi. We demonstrate that the three-dimensional photoresist structures are applicable to implementing complex multiple microchannels in PDMS by using the PDMS replica method.     

Immunosensing of Staphylococcus enterotoxin B (SEB) in milk with PDMS microfluidic systems using reinforced supported bilayer membranes (r-SBMs)

A versatile and novel method has been developed for microfluidic immunosensing of the food-borne pathogen Staphylococcus enterotoxin B (SEB) in poly(dimethylsiloxane) (PDMS) chips. Supported bilayer membranes (SBMs) were generated by vesicle fusion in oxidized PDMS microchannels for minimizing non-specific adsorption of biomolecules. The stability of SBMs was strengthened with a streptavidin layer to make them air-stable and allow for subsequent display of the biotin-functionalized antibodies. The reinforced supported bilayer membranes (r-SBMs) are fluid, exhibiting a lateral diffusion coefficient of approximately 1.9 microm(2) s(-1), and no detectable change of mobility was found after dehydration/rehydration. This is a substantial improvement over phosphatidylcholine (PC) membranes on PDMS, which suffered a roughly 10% reduction in the mobile fraction and 30% decrease in mobility after dehydration. Non-specific protein adsorption in the membrane-treated channels was reduced 100-1000 fold as compared to PDMS surfaces without a membrane coating. A flow-based microfluidic immunosensor for SEB was developed using antibodies linked to the r-SBMs in PDMS channels, and a detection limit of 0.5 ng mL(-1) was obtained from the linear portion of the calibration curve. The microchip was applied to detection of SEB in milk, and similar response and sensitivity were obtained, demonstrating the sensor's remarkable performance for real world samples. The r-SBMs overcome the stability hurdle in SBM-modified surfaces, opening up possibilities for transport and storage of membrane-functionalized microchips in the dehydrated form without compromising the performance, and facilitating the commercialization of disposable SBM-based microdevices.     

Fabrication of poly(dimethylsiloxane) microfluidic system based on masters directly printed with an office laser printer

Applications of poly(dimethylsiloxane) (PDMS)-based microfluidic systems are more popular nowadays. Previous fabrication methods of the masters for PDMS microchannels require complicated steps and/or special device. In this paper, we demonstrated that the toner printed on the transparency film with the office laser printer (1200 dpi) can be used as the positive relief of the masters. The transparency film was printed in two steps in order to obtain the same printing quality for the crossed lines. With the laser-printed master, the depth of the fabricated PDMS microchannels was ca. 10 microm and the smallest width was ca. 60 microm. Surface characteristics of the PDMS/PDMS microchannels were performed with SEM. Their electrokinetic properties were investigated by the aids of the measurement of electroosmotic flow (EOF) and the Ohm's curve. Using the PDMS/PDMS microchip CE systems, electroactive biological molecules and non-electroactive inorganic ions were well separated, respectively. This simple approach could make it easy to carry out the studies of PDMS microfluidic systems in more general labs without special devices.     

Development of a new contactless dielectrophoresis system for active particle manipulation using movable liquid electrodes

This study presents a new DEP manipulation technique using a movable liquid electrode, which allows manipulation of particles by actively controlling the locations of electrodes and applying on–off electric input signals. This DEP system consists of mercury as a movable liquid electrode, indium tin oxide (ITO)‐coated glass, SU‐8‐based microchannels for electrode passages, and a PDMS medium chamber. A simple squeezing method was introduced to build a thin PDMS layer at the bottom of the medium chamber to create a contactless DEP system. To determine the operating conditions, the DEP force and the friction force were analytically compared for a single cell. In addition, an appropriate frequency range for effective DEP manipulation was chosen based on an estimation of the Clausius–Mossotti factor and the effective complex permittivity of the yeast cell using the concentric shell model. With this system, we demonstrated the active manipulation of yeast cells, and measured the collection efficiency and the dielectrophoretic velocity of cells for different AC electric field strengths and applied frequencies. The experimental results showed that the maximum collection efficiency reached was approximately 90%, and the dielectrophoretic velocity increased with increasing frequency and attained the maximum value of 10.85 ± 0.95 μm/s at 100 kHz, above which it decreased.     

Microchannels filled with diverse micro- and nanostructures fabricated by glancing angle deposition

The integration of porous structures into microchannels is known to enable unique and useful separations both in electrophoresis and chromatography. Etched pillars and other nanostructures have received considerable interest in recent years as a platform for creating microchannels with pores tailored to specific applications. We present a versatile method for integration of three-dimensionally sculptured nano- and micro-structures into PDMS microchannels. Glancing angle deposition was used to fabricate nanostructures that were subsequently embedded in PDMS microchannels using a sacrificial resist process. With this technique, an assortment of structures made from a wide selection of materials can be integrated in PDMS microchannels; some examples of this versatility, including chiral and chevron nanostructures, are demonstrated. We also present a working device made using this process, separating 6/10/20 kbp and 10/48 kbp DNA mixtures in a DNA fractionator containing GLAD-deposited SiO(2) vertical posts as the separating medium. The separation mechanism was verified to resemble that found in prior fractionation devices, using total internal reflection fluorescence microscopy. GLAD fabrication enables insertion of three-dimensional structures into microchannels that cannot be fabricated with any existing techniques, and this versatility in structural design could facilitate new developments in on-chip separations.     

Fabrication of carbon microelectrodes with a micromolding technique and their use in microchip-based flow analyses

In this paper, we report a new technique to pattern carbon microelectrodes for use in microfluidics. This technique, termed micromolding of carbon inks, uses poly(dimethylsiloxane)(PDMS) microchannels to define the size of the microelectrode. First, PDMS microchannels of the approximate dimensions desired for the microelectrode are made by soft lithography. The PDMS is then reversibly sealed to a substrate and the microchannels are filled with carbon ink. After a heating step the PDMS mold is removed, leaving a carbon microelectrode with a size slightly smaller than the original PDMS microchannel. The resulting microelectrode (27 microm wide and 6 microm in height) can be reversibly sealed to a PDMS-based flow channel. Fluorescence microscopy showed that no leakage occurred around the chip/electrode seal, even up to flow rates of 10 microL min(-1). The electrode was characterized by microchip-based flow injection analysis. Injections of catechol in Hank's Balanced Salt Solution (pH 7.4), showed a linear response from 2 mM to 10 microM (r(2)= 0.995), with a sensitivity of 56.5 pA microM(-1) and an estimated limit of detection of 2 microM (0.27 picomole, S/N=3). Reproducibility of the electrode response was shown by repeated injections (n= 10) of a 500 microM catechol solution, resulting in a RSD of 4.6%. Finally, selectivity was demonstrated by coating the microelectrode with Nafion, a perfluoronated cation exchange polymer. Dopamine exhibited a response at the modified microelectrode while ascorbic acid was rejected by the Nafion-coating. These electrodes provide inexpensive detectors for microfluidic applications while also being viable alternatives to use of other carbon microelectrode materials, such as carbon fibers. Furthermore, the manner in which the microelectrodes are produced will be of interest to researchers who do not have access to state of the art microfabrication facilities.     

Tailoring the surface properties of poly(dimethylsiloxane) microfluidic devices

Poly(dimethylsiloxane) (PDMS) is an attractive material for microelectrophoretic applications because of its ease of fabrication, low cost, and optical transparency. However, its use remains limited compared to that of glass. A major reason is the difficulty of tailoring the surface properties of PDMS. We demonstrate UV grafting of co-mixed monomers to customize the surface properties of PDMS microfluidic channels in a simple one-step process. By co-mixing a neutral monomer with a charged monomer in different ratios, properties between those of the neutral monomer and those of the charged monomer could be selected. Mixtures of four different neutral monomers and two different charged monomers were grafted onto PDMS surfaces. Functional microchannels were fabricated from PDMS halves grafted with each of the different mixtures. By varying the concentration of the charged monomer, microchannels with electrophoretic mobilities between +4 x 10(-4) cm2/(V s) and -2 x 10(-4) cm2/(V s) were attainable. In addition, both the contact angle of the coated surfaces and the electrophoretic mobility of the coated microchannels were stable over time and upon exposure to air. By carefully selecting mixtures ofmonomers with the appropriate properties, it may be possible to tailor the surface of PDMS for a large number of different applications.     

Rapid separation of strychnine and brucine on a dynamically modified poly(dimethylsiloxane) microchip followed by electrochemical detection

A method has been developed for rapidly separating and detecting strychnine and brucine using a poly(dimethysiloxane) (PDMS) microchip and electrochemical (EC) detection. PDMS microchannels dynamically modified by Brij35 are shown to be more efficient than native ones. The two analytes are well separated within 90 s in 70 mmol/L acetate buffer (pH 5.5) containing 0.01% (v/v) Brij35. Detection limits were found to be 1.0 μmol/L for strychnine and 0.2 μmol/L for brucine at S/N=3. The method was used to determine trace strychnine and brucine in rat serum, and the results obtained correlate well with those obtained via high-performance liquid chromatography (HPLC).      

Embryonic development in the mouse is enhanced via microchannel culture

Microfluidic devices (microchannels) have been fabricated and tested for embryo culture. Three different microfabrication materials (silicon, polydimethylsiloxane (PDMS), and borosilicate) were used to fabricate the microchannels. The objective of this study was to determine if static microchannels permitted culture of mouse embryos to the blastocyst stage. Groups of 10 two-cell ICR x B6SJL/F1 mouse embryos were cultured for 96 hours in 4 different physical culture systems: 1) silicon/borosilicate microchannels, 2) PDMS/borosilicate microchannels, and 3) standard microdrops. Embryos cultured in the silicon/borosilicate and PDMS/borosilicate microchannels exhibited a faster rate of cleavage (P < 0.05), and produced more blastocysts (P < 0.01) than control microdrops. Furthermore, microchannels had a lower percentage of degenerated embryos than control embryos (P < 0.01). The results suggest that the microchannel culture systems may provide a culture environment that more closely mimics the in vivo environment.