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At least 1,351 records · Page 75Linked to original sources

PubMed2005

Long-term monitoring of bacteria undergoing programmed population control in a microchemostat.

Using an active approach to preventing biofilm formation, we implemented a microfluidic bioreactor that enables long-term culture and monitoring of extremely small populations of bacteria with single-cell resolution. We used this device to observe the dynamics of Escherichia coli carrying a synthetic "population control" circuit that regulates cell density through a feedback mechanism based on quorum sensing. The microfluidic bioreactor enabled long-term monitoring of unnatural behavior programmed by the synthetic circuit, which included sustained oscillations in cell density and associated morphological changes, over hundreds of hours.

4-Butyrolactone↗

PubMed2005

Transition of MEMS technology to nanofabrication.

The transition of MEMS technology to nano fabrication is a solution to the growing demand for smaller and high-density feature sizes in the nanometer scale. Nanoimprint lithography (NIL) techniques for fabricating micro- and nano-features are discussed including hot embossing lithography (HEL), UV Molding (UVM) and micro contact printing (microCP). Recent results in micro and nano-pattern transfer are presented where features ranged from < 100 nm to several centimeters. We also present a comparative study between standard glass microfluidic chips and their HEL counterparts by metrology. Hot-embossed microfluidic chips are shown to be faithful replicates of their parent stamps. NIL is presented as a promising avenue for low-cost, high throughput micro and nano-device fabrication.

Equipment Design↗

PubMed2006

Surface modification methods for enhanced device efficacy and function.

Currently available microfluidic devices can accomplish a variety of tasks useful in molecular biology. When moving analytical processes to a microenvironment, the properties of the device surface play a larger role in the functioning of the device. Surface modification may become necessary or advantageous for the purpose of control of the functional mechanics of the device, keeping cell components from adsorbing, attaching antibodies to the surface for detection of biological components, and attaching a functional bonding complex. Modification of the surface of microfluidic devices for the control of flow and device function, or for functionalization of the surface to tailor the device to a specific use, can be accomplished in numerous bench-top, postfabrication procedures. The use of polyelectrolyte multilayers, ultraviolet grafting of polymers, and polydimethylsiloxane/surfactant coating to control flow and mitigate adsorption is discussed. In addition, the functionalization of devices through amine termination of surfaces, and immobilization of biotin within a phosphotidylcholine bilayer is detailed.

Biotin↗

PubMed2005

Protein biochip systems for the clinical laboratory.

Classical methods of protein analysis such as electrophoresis, ELISA and liquid chromatography are generally time-consuming, labor-intensive and lack high-throughput capacity. In addition, all existing methods used to measure proteins necessitate multiple division of the original sample and individual tests carried out for each substance, with an associated cost for each test. The chip system allows several tests to be performed simultaneously without dividing the original patient sample. This system facilitates the development of multiplexed assays that simultaneously measure many different analytes in a small sample volume. These emerging technologies fall into two categories: 1) spotted array-based tools, and 2) microfluidic-based tools. Miniaturized and multiplexed immunoassays allow a great deal of information to be obtained from a single sample. These analytical systems are referred to as "lab-on-a-chip" devices. This article presents current trends and advances in miniaturized multiplexed immunoassay technologies, reviewing different systems from research to point-of-care assays. We focus on a subset of chip-based assays that may be used in a clinical laboratory and are directly applicable for biomedical diagnosis. Recent advances in biochip assays combine the power of miniaturization, microfluidics, micro- to nanoparticles, and quantification. A number of applications are just beginning to be explored. The power of biochip assays offers great promise for point-of-care clinical testing and monitoring of many important analytes.

Clinical Laboratory Techniques↗

PubMed2006

Extracellular recordings of field potentials from single cardiomyocytes.

Open microfluidic channels were used to separate the extracellular space around a cardiomyocyte into three compartments: the cell ends and a central partition (insulating gap). The microchannels were filled with buffer solution and overlaid with paraffin oil, thus forming the cavities for the cell ends. The central part of the cardiomyocyte rested on the partition between two adjacent microchannels and was entirely surrounded by the paraffin oil. This arrangement increased the extracellular electrical resistance to > 20 MOmega and facilitated the recording of the time course of the change in extracellular voltage and current during subthreshold and suprathreshold stimuli. The waveform of the extracellular current and voltage in response to an extracellular depolarizing stimulus comprised an initial monophasic signal followed by a biphasic signal with a delay of 2-15 ms. The latter was associated with a transient contraction and therefore caused by an action potential. The biphasic signal became monophasic after the depolarization of one cell end by raised extracellular [K+]. This form of differential recording revealed the repolarization phase of the action potential. At rest, the sarcomere length within the gap was 12% +/- 4.8% longer than outside the gap, but intracellular Ca2+ transients occurred to the same extent as that observed in the outer pools. This data demonstrate the feasibility of the use of a microfluidic bath design to limit the extracellular resistance between two ends of an isolated cardiomyocyte.

Action Potentials↗

PubMed2006

Novel determination system for urea in alcoholic beverages by using an FIA system with an acid urease column.

A novel determination method for urea using an acid urease column-FIA system was developed, and the system was applied to the determination of urea in rice wine. This novel FIA system was characterized by CO2 detection due to the property of acid urease and by a microfluidic gas-diffusion device with the use of an ultra-thin hollow fiber membrane. A biosensing system fabricated in this study was assembled with a double-plunger pump, a sample-injection valve, an immobilized acid urease column as a recognition element for the assay of urea, a gas-diffusion unit, and a flow-type spectrophotometer. The gas-diffusion unit consisted of a double-tubing structure in which the outer tubing was made of PTFE (i.d. 1.0 mm; o.d. 1.5 mm) and the inner tubing was of porous PTFE (i.d. 0.19 mm; o.d. 0.25 mm). Standard urea solutions (20 microl) were measured through monitoring variations in the absorbance of a coloring agent solution resulting from a pH shift due to carbon dioxide molecules being enzymatically generated. A wide and linear relationship was obtained between the concentration of urea (16 microM - 1.0 mM) and the change in absorbance. This FIA system has great advantages that the system did not suffer from ammonia and ethanol in samples. This system, armed with a microfluidic gas-diffusion device, was applicable to the determination of various substrates of many kinds of decarboxylase, amino-acid oxidase, and amino-acid oxygenase, producing CO2 and NH3 molecules.

Acids↗

PubMed2006

Transparent polymeric cell culture chip with integrated temperature control and uniform media perfusion.

Modern microfabrication and microfluidic technologies offer new opportunities in the design and fabrication of miniaturized cell culture systems for online monitoring of living cells. We used laser micromachining and thermal bonding to fabricate an optically transparent, low-cost polymeric chip for long-term online cell culture observation under controlled conditions. The chip incorporated a microfluidic flow equalization system, assuring uniform perfusion of the cell culture media throughout the cell culture chamber. The integrated indium-tin-oxide heater and miniature temperature probe linked to an electronic feedback system created steady and spatially uniform thermal conditions with minimal interference to the optical transparency of the chip. The fluidic and thermal performance of the chip was verified by finite element modeling and by operation tests under fluctuating ambient temperature conditions. HeLa cells were cultured for up to 2 weeks within the cell culture chip and monitored using a time-lapse video recording microscopy setup. Cell attachment and spreading was observed during the first 10-20 h (lag phase). After approximately 20 h, cell growth gained exponential character with an estimated doubling time of about 32 h, which is identical to the observed doubling time of cells grown in standard cell culture flasks in a CO2 incubator.

Cell Culture Techniques↗

PubMed2004

Towards sorting of biolibraries using single-molecule fluorescence detection techniques.

The selection of specific binding molecules like peptides and proteins from biolibraries using, for instance, phage display methods can be quite time-consuming. It is therefore desirable to develop a strategy that is much faster in selection and sorting of potential binders out of a biolibrary. In this contribution we separately discuss the current achievements in generation of biolibraries, single-molecule detection techniques and microfluidic devices. A high-throughput microfluidic platform is then proposed that combines the propulsion of liquid containing fluorescent components of the biolibrary through microchannels, single-molecule fluorescence photon burst detection and real-time sorting of positive hits.

Fluorescence↗

PubMed2001

Proteomics on a chip: promising developments.

The field of proteomics is expanding rapidly due to the completion of the human genome and the realization that genomic information is often insufficient to comprehend cellular mechanisms. This considerable expansion of proteomics towards high-throughput platforms is stressing its current technical capabilities. In recent years, technologies in microfluidic and array technologies have appeared for proteomics. These novel approaches might help solve current technical challenges in proteomics. This review presents a general survey of the recent development in microfluidic and array technologies from a proteomics perspective.

Amino Acid Sequence↗

PubMed2001

Patterned self-assembled beads in silicon channels.

A novel technique enabling selective bead trapping in microfluidic devices without the use of physical barriers is presented in this paper. It is a fast, convenient and simple method, involving microcontact printing and self-assembly, that can be applied to silicon, quartz or plastic substrates. In the first step, channels are etched in the substrate. The surface chemistry of the internal walls of the channels is then modified by microcontact printing. The chip is submerged in a bead slurry where beads self-assemble based on surface chemistry and immobilize on the internal walls of the channels. Silicon channels (100 microm wide and 50 microm deep) have been covered with monolayers of streptavidin-, amino- and hydroxy-functionalized microspheres and resulted in good surface coverage of beads on the channel walls. A high-resolution pattern of lines of self-assembled streptavidin beads, as narrow as 5 microm, has also been generated on the bottom of a 500 microm wide and 50 microm deep channel. Flow tests were performed in sealed channels with the different immobilized beads to confirm that the immobilized beads could withstand the forces generated by water flowing in the channels. The presented results indicate that single beads can be precisely positioned within microfluidic devices based on self-assembly which is useful as screening and analysis tools within the field of biochemistry and organic chemistry.

Animals↗

PubMed2002

On-chip enzymatic assays.

This article reviews different possibilities for conducting enzymatic assays on microchip platforms, along with potential advantages, limitations, and selected examples of such biochips. Enzyme-based chips combine the analytical power and reagent economy of microfluidic devices with the selectivity and amplification features of biocatalytic reactions. "Lab-on-chip" devices thus allow enzymatic assays to be performed more rapidly, easily, and economically. Such assays usually rely on on-chip mixing and reactions (of the substrates and enzymes) in connection to separations (of the substrates or products). The realization of on-chip enzymatic assays thus requires understanding of how enzymatic reactions behave on a small scale and can be interfaced with separation microchips, and how the microfluidics can be tailored to suit the requirements of particular enzymatic assays. The goal is to obtain sufficient reaction times, without compromising the quality of the analytical separation. The versatility of such on-chip enzymatic assays offers great promise for decentralized testing of clinically or environmentally important substrates.

Electrophoresis, Capillary↗

PubMed2002

A nanoliter rotary device for polymerase chain reaction.

Polymerase chain reaction (PCR) has revolutionized a variety of assays in biotechnology. The ability to implement PCR in disposable and reliable microfluidic chips will facilitate its use in applications such as rapid medical diagnostics, food control testing, and biological weapons detection. We fabricated a microfluidic chip with integrated heaters and plumbing in which various forms of PCR have been successfully demonstrated. The device uses only 12 nL of sample, one of the smallest sample volumes demonstrated to date. Minimizing the sample volume allows low power consumption, reduced reagent costs, and ultimately more rapid thermal cycling.

Polymerase Chain Reaction↗

PubMed2002

Electroosmotic pumping in microchips with nonhomogeneous distribution of electrolytes.

A general equation to calculate the node pressure at a junction in a microfluidic network is presented. The node pressure is generated from both the hydrodynamic flow due to the external applied hydraulic pressures and the electrokinetic flow resulted from the applied electric field. Pure electroosmotic flow has a plug-flow profile and pressure flow has a parabolic flow profile. In a first order approximation, these two flows can be treated separately, and the total flow is the sum of the two. An externally applied pressure simply creates a constant offset in the node pressure as long as the flow resistances remain the same. In a nonhomogeneous microfluidic network, where the electrical resistivity or the electroosmotic mobility is not constant everywhere, the differences in electroosmotic flow in various sections of the network will create an electroosmotically induced pressure at the internal nodes. Our theoretical approach can easily be extended to networks with more than one internal node. One prediction of this theory is that any variation in electroosmotic mobility or solution resistivity in different network branches will generate a pressure, and can thus be used as a pump. As an example, we demonstrate electroosmotic pumping in a high-low buffer system.

Electrolytes↗

PubMed2002

DNA analysis on microfabricated electrophoretic devices with bubble cells.

Microfluidic devices with bubble cells have been fabricated on poly(methyl methacrylate) (PMMA) plates and have been employed for the analysis of DNA using polyethylene oxide (PEO) solutions. First, the separation channel was fabricated using a wire-imprinting method. Then, wires with greater sizes or a razor blade glued in a polycarbonate plate was used to fabricate bubble cells, with sizes of 190-650 microm. The improvements in resolution and sensitivity have been achieved for large DNA (> 603 base pair, bp) using such devices, which depend on the geometry of the bubble cell. The main contributor for optimal resolution is mainly due to DNA migration at lower electric field strengths inside the bubble cell. On the other hand, slight losses of resolution for small DNA fragments have been found mainly due to diffusion, supported by the loss of resolution when separating two small solutes. With a bubble cell of 75 microm (width) x 500 microm (depth), the sensitivity improvement up to 17-fold has been achieved for the 271 bp fragment in the separation of PhiX-174/HaeIII DNA restriction fragments. We have also found that a microfluidic device with a bubble cell of 360 microm x 360 microm is appropriate for DNA analysis. Such a device has been used for separating DNA ranging from 8 to 2176 bp and polymerase chain reaction (PCR) products amplified after 30 cycles, with rapidity and improvements in the sensitivity as well as resolution.

Base Sequence↗

PubMed2002

Trapping of DNA by dielectrophoresis.

Under suitable conditions, a DNA molecule in solution will develop a strong electric dipole moment. This induced dipole allows the molecule to be manipulated with field gradients, in a phenomenon known as dielectrophoresis (DEP). Pure dielectrophoretic motion of DNA requires alternate current (AC) electric fields to suppress the electrophoretic effect of the molecules net charge. In this paper, we present two methods for measuring the efficiency of DEP for trapping DNA molecules as well as a set of quantitative measurements of the effects of strand length, buffer composition, and frequency of the applied electric field. A simple configuration of electrodes in combination with a microfluidic flow chamber is shown to increase the concentration of DNA in solution by at least 60-fold. These results should prove useful in designing practical microfluidic devices employing this phenomenon either for separation or concentration of DNA.

Bacteriophage lambda↗

PubMed2002

Fabrication and use of a transient contractional flow device to quantify the sensitivity of mammalian and insect cells to hydrodynamic forces.

A microfluidic device was fabricated via photolithographic techniques which can create transient elongational and shear forces ranging over three orders of magnitude while still maintaining laminar flow conditions. The contractional fluid flow inside the microfluidic device was simulated with FLUENT (a computational fluid dynamics computer program) and the local deformation forces were characterized with the scalar quantity, local energy dissipation rate. The sensitivities of four cell lines (CHO, HB-24, Sf-9, and MCF7) were tested in the device. The results indicate that all four cell lines are able to withstand relatively intense energy dissipation rates (up to 10(4)-10(5) kW/m(3)), which is orders of magnitude higher than the maximum local energy dissipation rates generated by impellers in bioreactors, but comparable to that associated with small bursting bubbles. While the concept that suspended animal cells are relatively robust with respect to purely hydrodynamic forces in bioprocess equipment is well known, these results quantitatively demonstrate these observations.

Animals↗

PubMed2003

Anisotropic stretch-induced hypertrophy in neonatal ventricular myocytes micropatterned on deformable elastomers.

Because cell shape and alignment, cell-matrix adhesion, and cell-cell contact can all affect growth, and because mechanical strains in vivo are multiaxial and anisotropic, we developed an in vitro system for engineering aligned, rod-shaped, neonatal cardiac myocyte cultures. Photolithographic and microfluidic techniques were used to micropattern extracellular matrices in parallel lines on deformable silicone elastomers. Confluent, elongated, aligned myocytes were produced by varying the micropattern line width and collagen density. An elliptical cell stretcher applied 2:1 anisotropic strain statically to the elastic substrate, with the axis of greatest stretch (10%) either parallel or transverse to the myofibrils. After 24 h, the principal strain parallel to myocytes did not significantly alter myofibril accumulation or expression of atrial natriuretic factor (ANF), connexin-43 (Cx-43), or N-cadherin (by indirect immunofluorescent antibody labeling and immunoblotting) compared with unstretched controls. In contrast, 10% transverse principal strain resulted in continuous staining of actin filaments (rhodamine phalloidin); increased immunofluorescent labeling of ANF, Cx-43, and N-cadherin; and upregulation of protein signal intensity by western blotting. By using microfabrication and microfluidics to control cell shape and alignment on an elastic substrate, we found greater effects for transverse than for longitudinal stretch in regulating sarcomere organization, hypertrophy, and cell-to-cell junctions.

Actins↗

PubMed2004

Deformability-based flow cytometry.

BACKGROUND: Elasticity of cells is determined by their cytoskeleton. Changes in cellular function are reflected in the amount of cytoskeletal proteins and their associated networks. Drastic examples are diseases such as cancer, in which the altered cytoskeleton is even diagnostic. This connection between cellular function and cytoskeletal mechanical properties suggests using the deformability of cells as a novel inherent cell marker. METHODS: The optical stretcher is a new laser tool capable of measuring cellular deformability. A unique feature of this deformation technique is its potential for high throughput, with the incorporation of a microfluidic delivery of cells. RESULTS: Rudimentary implementation of the microfluidic optical stretcher has been used to measure optical deformability of several normal and cancerous cell types. A drastic difference has been seen between the response of red blood cells and polymorphonuclear cells for a given optically induced stress. MCF-10, MCF-7, and modMCF-7 cells were also measured, showing that while cancer cells stretched significantly more (five times) than normal cells, optical deformability could even be used to distinguish metastatic cancer cells from nonmetastatic cancer cells. This trimodal distribution was apparent after measuring a mere 83 cells, which shows optical deformability to be a highly regulated cell marker. CONCLUSIONS: Preliminary work suggests a deformability-based cell sorter similar to current fluorescence-based flow cytometry without the need for specific labeling. This could be used for the diagnosis of all diseases, and the investigation of all cellular processes, that affect the cytoskeleton.

Breast Neoplasms↗