Microfluidic devices for the analysis of apoptosis

Apoptosis is the outcome of a metabolic cascade that results in cell death in a controlled manner. Due to its important role in maintaining balance in organisms, in mechanisms of diseases, and tissue homeostasis, apoptosis is of great interest in the emerging fields of systems biology. Research into cell death regulation and efforts to model apoptosis processes have become powerful drivers for new technologies to acquire ever more comprehensive information from cells and cell populations. The microfluidic technology promises to integrate and miniaturize many bioanalytical processes, which offers an alternative platform for the analysis of apoptosis. This review aims to highlight the recent developments of microfluidic devices in measuring the hallmarks as well as the dynamic process of cellular apoptosis. The potential capability and an outlook of microfluidic devices for the study of apoptosis are addressed.     

Microfluidic devices: useful tools for bioprocess intensification

The dawn of the new millennium saw a trend towards the dedicated use of microfluidic devices for process intensification in biotechnology. As the last decade went by, it became evident that this pattern was not a short-lived fad, since the deliverables related to this field of research have been consistently piling-up. The application of process intensification in biotechnology is therefore seemingly catching up with the trend already observed in the chemical engineering area, where the use of microfluidic devices has already been upgraded to production scale. The goal of the present work is therefore to provide an updated overview of the developments centered on the use of microfluidic devices for process intensification in biotechnology. Within such scope, particular focus will be given to different designs, configurations and modes of operation of microreactors, but reference to similar features regarding microfluidic devices in downstream processing will not be overlooked. Engineering considerations and fluid dynamics issues, namely related to the characterization of flow in microchannels, promotion of micromixing and predictive tools, will also be addressed, as well as reflection on the analytics required to take full advantage of the possibilities provided by microfluidic devices in process intensification. Strategies developed to ease the implementation of experimental set-ups anchored in the use of microfluidic devices will be briefly tackled. Finally, realistic considerations on the current advantages and limitation on the use of microfluidic devices for process intensification, as well as prospective near future developments in the field, will be presented.     

Microfluidic devices for studies of shear-dependent platelet adhesion

Adhesion of platelets to blood vessel walls is a shear stress dependent process that promotes arrest of bleeding and is mediated by the interaction of receptors expressed on platelets with various extracellular matrix (ECM) proteins that may become exposed upon vascular injury. Studies of dynamic platelet adhesion to ECM-coated substrates in conventional flow chambers require substantial fluid volumes and are difficult to perform with blood samples from a single laboratory mouse. Here we report dynamic platelet adhesion assays in two new microfluidic devices made of PDMS. Small cross-sections of the flow chambers in the devices reduce the blood volume requirements to <100 microl per assay, making the assays compatible with samples of whole blood obtained from a single mouse. One device has an array of 8 flow chambers with shear stress varying by a factor of 1.93 between adjacent chambers, covering a 100-fold range from low venous to arterial. The other device allows simultaneous high-resolution fluorescence imaging of dynamic adhesion of platelets from two different blood samples. Adhesion of platelets in the devices to three common ECM substrate coatings was verified to conform with published results. The devices were subsequently used to study the roles of extracellular and intracellular domains of integrin alphaIIbbeta3, a platelet receptor that is a central mediator of platelet aggregation and thrombus formation. The study involved wild-type mice and two genetically modified mouse strains and showed that the absence of the integrin impaired adhesion at all shear stresses, whereas a mutation in its intracellular domain reduced the adhesion only at moderate and high stresses. Because of small sample volumes required, the devices could be employed in research with genetically-modified model organisms and for adhesion tests in clinical settings with blood from neonates.     

Microfluidic devices fabricated in poly(dimethylsiloxane) for biological studies

This review describes microfluidic systems in poly(dimethylsiloxane) (PDMS) for biological studies. Properties of PDMS that make it a suitable platform for miniaturized biological studies, techniques for fabricating PDMS microstructures, and methods for controlling fluid flow in microchannels are discussed. Biological procedures that have been miniaturized into PDMS-based microdevices include immunoassays, separation of proteins and DNA, sorting and manipulation of cells, studies of cells in microchannels exposed to laminar flows of fluids, and large-scale, combinatorial screening. The review emphasizes the advantages of miniaturization for biological analysis, such as efficiency of the device and special insights into cell biology.     

Direct measurement of extracellular electrical signals from mammalian olfactory sensory neurons in planar triode devices

An artificial nose was developed to mimic aspects of sensory transduction of the peripheral mammalian olfactory system. We directly cultured and differentiated rat olfactory sensory neurons (OSNs) on indium-tin oxide electrodes of planar triode substrates without a coupling agent. Direct voltage (~50 μV) and current (~250 nA) signals were measured simultaneously when OSNs on the planar triode substrates were exposed to odorant mixtures. The response signals were sensitive to the concentration of the odorant mixture, with a typical lifetime, shape, and adaptation profile as seen in responses upon repeated stimulation in vivo. We found that the rising time to the peak current was ~161 ms, while the signal back to baseline was in 1.8 s, which are in agreement with the natural intracellular electrophysiological responses. These results provide the first evidence that mature OSNs grown in a planar triode device are able to detect direct electrophysiological responses to odorants.     

Microfluidic designs and techniques using lab-on-a-chip devices for pathogen detection for point-of-care diagnostics

Effective pathogen detection is an essential prerequisite for the prevention and treatment of infectious diseases. Despite recent advances in biosensors, infectious diseases remain a major cause of illnesses and mortality throughout the world. For instance in developing countries, infectious diseases account for over half of the mortality rate. Pathogen detection platforms provide a fundamental tool in different fields including clinical diagnostics, pathology, drug discovery, clinical research, disease outbreaks, and food safety. Microfluidic lab-on-a-chip (LOC) devices offer many advantages for pathogen detection such as miniaturization, small sample volume, portability, rapid detection time and point-of-care diagnosis. This review paper outlines recent microfluidic based devices and LOC design strategies for pathogen detection with the main focus on the integration of different techniques that led to the development of sample-to-result devices. Several examples of recently developed devices are presented along with respective advantages and limitations of each design. Progresses made in biomarkers, sample preparation, amplification and fluid handling techniques using microfluidic platforms are also covered and strategies for multiplexing and high-throughput analysis, as well as point-of-care diagnosis, are discussed.     

Monolayers of twisted binaphthyls for aromatic crystallization at low nucleation densities and high growth rates

Monolayers of 1,1'-bi-2-naphthol (BN) derivatives, of which the two naphthalene rings are twisted along the carbon(1)-carbon(1') single bond, were studied for their conformational effect on the growth of pentacene crystals on their monolayer surface. BN monolayers with H and Br at 6,6'-positions (H-BN and Br-BN) were prepared by immersion-coating in toluene solution of the corresponding BNSiCl2. Pentacene was thermally evaporated onto the H-BN and Br-BN monolayers, silica, octadecylsilyl (ODTS) SAM, and a micropattern of H-BN and ODTS SAM. Pentacene crystals were also grown on the SAMs of 1-naphthylsilyl(NPh), phenylsilyl(Ph), and diphenylsilyl (DPh) groups, which are aromatic and have contact angle values similar to those of the the BN monolayers. AFM images of the crystals at the early stage of growth indicated that the BN monolayers suppressed the nucleation while facilitating the growth of nuclei to larger crystals. The low nucleation density and high growth rate are accounted for by the amorphous nature of the twisted BN monolayer surface where the intermolecular interaction between neighboring adsorbates is likely to be suppressed. The results offer new insights into designing surfaces for controlling the crystallization kinetics of organic materials.     

Microfluidic devices for electrochemical measurement of photosynthetic activity of cyanobacteria microcystis cells

A microfluidic device with analytical chambers for electrochemical measurements has been employed to detect photosynthetic activity at single cell level. The flowing cells (Microcystis viridis) in a main channel are individually guided to the chamber with microelectrodes by an electrophoretic manipulation. The reduction current of oxygen was continuously monitored to determine the photosynthetic activity upon light irradiation. The average rates for oxygen generation were estimated and found to be 10(-18) mol/s level.     

Microfluidic devices with disposable enzyme electrode for electrochemical monitoring of glucose concentrations

This article describes the fabrication of tube-like microchannels made of UV curable polymer on a glass substrate and the device assembling with a disposable enzyme-working electrode for high-sensitivity electrochemical detection. While both reference and counter electrodes are patterned on the surface of the glass substrate, the working electrode is flipped on the top of the channel with an open access, providing a face-to-face probing configuration. When the enzyme electrode is contaminated or degraded, it can be easily replaced by a new one, keeping the main body of the device and the detection schema unchanged. Using glucose oxidase-coated gold electrodes, we were able to determine a linear amperometry response to the glucose concentrations in the range of 2-16 mM. By replacing the as-prepared working electrode by the one after thermal treatments, we showed a much more degraded enzyme electrode activity, enabling efficient determination of the electrode quality as well as the whole process optimization.     

Microfluidic devices for fluidic circulation and mixing improve hybridization signal intensity on DNA arrays

Reactions of biomolecules with surface mounted materials on microscope slides are often limited by slow diffusion kinetics, especially in low volumes where diffusion is the only means of mixing. This is a particular problem for reactions where only small amounts of analyte are available and the required reaction volume limits the analyte concentration. A low volume microfluidic device consisting of two interconnected 9 mm x 37.5 mm reaction chambers was developed to allow mixing and closed loop fluidic circulation over most of the surface of a microscope slide. Fluid samples are moved from one reaction chamber to the other by the rotation of a magnetic stirring bar that is driven by a standard magnetic stirrer. We demonstrate that circulation and mixing of different reagents can be efficiently accomplished by this closed loop device with solutions varying in viscosity from 1 to 16.2 centipoise. We also show by example of a microarray hybridization that the reaction efficiency can be enhanced 2-5 fold through fluid mixing under conditions where diffusion is rate limiting. For comparison, similar results were achieved with a disposable commercial device that covers only half of the reaction area of the closed loop device.     

The viscosity of triethylamine vapor at low densities and in the saturated vapor phase

For the first time, results of high-precision measurements of the viscosity coefficient of triethylamine vapor at low densities are reported. The relative measurements with an all-quartz oscillating-disk viscometer were carried out along seven isochores at densities from 0.002 to 0.009 mol m⁻³ in the temperature range between 298 and 498 K. The uncertainty is estimated to be ±$\pm$0.2% at ambient temperature, increasing up to ±$\pm$0.3% at higher temperatures. First isothermal values were recalculated from the original experimental data and then evaluated with a first-order expansion for the viscosity, in terms of density. In addition, viscosity values of the saturated vapor were determined at low temperatures. The results are utilized to model the viscosity coefficient of triethylamine vapor at moderately low densities. A so-called individual correlation on the basis of the extended theorem of corresponding states was employed to describe the zero-density viscosity coefficient, whereas the Rainwater–Friend theory was used to represent the initial density dependence expressed as second viscosity virial coefficient.     

Microfluidic sensing devices employing in situ-formed liquid crystal thin film for detection of biochemical interactions

Although biochemical sensing using liquid crystals (LC) has been demonstrated, relatively little attention has been paid towards the fabrication of in situ-formed LC sensing devices. Herein, we demonstrate a highly reproducible method to create uniform LC thin film on treated substrates, as needed, for LC sensing. We use shear forces generated by the laminar flow of aqueous liquid within a microfluidic channel to create LC thin films stabilized within microfabricated structures. The orientational response of the LC thin films to targeted analytes in aqueous phases was transduced and amplified by the optical birefringence of the LC thin films. The biochemical sensing capability of our sensing devices was demonstrated through experiments employing two chemical systems: dodecyl trimethylammonium bromide (DTAB) dissolved in an aqueous solution, and the hydrolysis of phospholipids by the enzyme phospholipase A(2) (PLA(2)).     

A low band gap conjugated polymer for supercapacitor devices

We present the electrochemical polymerization and characterization of a stable and dopable polyindophenine derivative starting from monomer 5. The fabrication of supercapacitor devices was carried out by direct electrochemical polymerization on an ITO-coated slide. Furthermore, the galvanostatic and potentiostatic experiments conducted on these supercapacitors have shown that the polyindophenine-derivative material has very good cyclability, over a potential range of 1.4 V, and a maximum capacitance of 140 F g(-1).     

Molecular dynamics simulations of self-diffusion coefficient and thermal conductivity of methane at low and moderate densities

This article demonstrates a highly accurate molecular dynamics (MD) simulation of thermal conductivity of methane using an ab initio intermolecular potential. The quantum effects of the vibrational contribution to thermal conductivity are more efficiently accounted for in the present MD model by an analytical correction term as compared to by the Monte Carlo method. The average deviations between the calculated thermal conductivity and the experimental data are 0.92% for dilute methane and 1.29% for methane at moderate densities, as compared to approximately 20% or more in existing MD calculations. The results demonstrate the importance of considering vibrational contribution to the thermal conductivity which is mainly through the self-diffusion process.     

Configurational probabilities for symmetric dimers on a lattice: an analytical approximation with exact limits at low and high densities

A new approach is developed for lattice density functional theory of interacting symmetric dimers at high temperatures. Equations of equilibrium for two-dimensional square and three-dimensional cubic lattices are derived for the complete set of configurations in the first three shells around the central dimer, and rules of truncation for higher shells are based on exact results from the mathematical theory of domino tilings. This provides exact limits for both low and high densities. The new model predicts contributions of particular configurations which are in agreement with Monte Carlo simulations over the whole range of densities, including agreement with pocket Monte Carlo simulations at high densities.     

Memory function for the velocity autocorrelation function at moderate densities

The memory function for the velocity autocorrelation function is evaluated at low and moderate densities for the square well potential. Short time deviations from the Boltzmann-Lorentz result are calculated for both incomplete scattering events and bound pairs. It is shown that the bound states are responsible for certain structural anomalies found in recent molecular dynamics simulation at low temperature.     

Ultraviolet-based bonding for perfluoropolyether low aspect-ratio microchannels and hybrid devices

Producing solvent-resistant microfluidic devices is a challenge for analytical chemistry and biochemistry. We demonstrate a simple and low-cost fabrication approach for the realization of solvent-resistant microchannels based on perfluoropolyether elastomers, exhibiting very low aspect ratios (0.01). The strength of the microchannels sealing is evaluated through the maximum internal pressure (1.52 MPa) prior to device failure, due to delamination at the bonded interface. This approach allows the elastic properties of silicone elastomers, suitable for high quality external connections, to be combined with the non-swelling character of perfluoropolyethers.     

Copolymerization of styrene and diethyl fumarate with zinc bromide. III. Electroinitiation at low current densities

The electroinitiated preparation of equimolar copolymers of styrene and diethyl fumarate is described. The reaction medium consisted of these monomers dissolved in a methanol–zinc bromide solution. Rates of polymerization increased with increasing applied current and the copolymer yield increased linearly with the total number of Faradays transferred. The copolymer composition is 1:1 and is essentially invariant with the degree of conversion (<15%), electroinitiation rate, and monomer feed ratios. A reaction mechanism involving donor–acceptor complexes and electroinitiated excitation of these complexes at the electrode is postulated.     

Fluorinated carbon nanofibres for high energy and high power densities primary lithium batteries

The electrochemical performances of fluorinated carbon nanofibres have been tested for a use as cathode material in primary lithium battery using LiBF₄ PC:DME 1M as electrolyte. For a very narrow fluorination range (420–450 °C), the fluorine content in the carbon nanofibres increases up to CF_0.78 and so do both the energy and the power densities. A maximum of 8057 W kg⁻¹ power density has been reached. Moreover, a current density of 6C can be used for such fluorinated carbon nanofibres. Such high electrochemical values can be correlated to the amount of unfluorinated carbon located in the core of the carbon nanofibres. Owing to solid state ¹³C NMR which can accurately evaluated this fraction, a minimum of 10% of unfluorinated carbon nanofibre is necessary in order to insure a good conducting behaviour.     

Microfluidic electroporation for selective release of intracellular molecules at the single-cell level

Analysis of intracellular materials at the single-cell level presents opportunities for probing the heterogeneity of a cell population. Lysis by electroporation has been gaining popularity as a rapid method for disruption of the cell membrane and release of intracellular contents. In this report, we selectively released specific intracellular molecules for interrogation at the single-cell level by tuning the parameters of electroporation. We examined the release of a small molecule, calcein (MW approximately 600), and a 72-kDa protein kinase, Syk, tagged by enhanced green fluorescent protein (EGFP) from chicken B cells during electroporation at the single-cell level. We studied the effects of the field intensity and the field duration on the release of the two molecules. We found that calcein in general was released at lower field intensities and shorter durations than did SykEGFP. By tuning the electrical parameters, we were able to deplete calcein from the cells before SykEGFP started to release. This approach potentially provides a high-throughput alternative for probing different intracellular molecules at the single-cell level compared to chemical cytometry by eliminating complete disruption of the cell membrane.