Flow In Microfluidic Channels Research Articles

Optical chromatographic sample separation of hydrodynamically focused mixtures.

Optical chromatography relies on the balance between the opposing optical and fluid drag forces acting on a particle. A typical configuration involves a loosely focused laser directly counter to the flow of particle-laden fluid passing through a microfluidic device. This equilibrium depends on the intrinsic properties of the particle, including size, shape, and refractive index. As such, uniquely fine separations are possible using this technique. Here, we demonstrate how matching the diameter of a microfluidic flow channel to that of the focusing laser in concert with a unique microfluidic platform can be used as a method to fractionate closely related particles in a mixed sample. This microfluidic network allows for a monodisperse sample of both polystyrene and poly(methyl methacrylate) spheres to be injected, hydrodynamically focused, and completely separated. To test the limit of separation, a mixed polystyrene sample containing two particles varying in diameter by less than 0.5 μm was run in the system. The analysis of the resulting separation sets the framework for continued work to perform ultra-fine separations.

Stop flow lithography in perfluoropolyether (PFPE) microfluidic channels.

Stop Flow Lithography (SFL) is a microfluidic-based particle synthesis method for creating anisotropic multifunctional particles with applications that range from MEMS to biomedical engineering. Polydimethylsiloxane (PDMS) has been typically used to construct SFL devices as the material enables rapid prototyping of channels with complex geometries, optical transparency, and oxygen permeability. However, PDMS is not compatible with most organic solvents which limit the current range of materials that can be synthesized with SFL. Here, we demonstrate that a fluorinated elastomer, called perfluoropolyether (PFPE), can be an alternative oxygen permeable elastomer for SFL microfluidic flow channels. We fabricate PFPE microfluidic devices with soft lithography and synthesize anisotropic multifunctional particles in the devices via the SFL process--this is the first demonstration of SFL with oxygen lubrication layers in a non-PDMS channel. We benchmark the SFL performance of the PFPE devices by comparing them to PDMS devices. We synthesized particles in both PFPE and PDMS devices under the same SFL conditions and found the difference of particle dimensions was less than a micron. PFPE devices can greatly expand the range of precursor materials that can be processed in SFL because the fluorinated devices are chemically resistant to most organic solvents, an inaccessible class of reagents in PDMS-based devices due to swelling.

Effect of pH waves on capacitive charging in microfluidic flow channels

Novel energy-efficient desalination techniques, such as capacitive deionization (CDI), are a key element for the future of the fresh water supply, which is increasingly under stress due to the ever-growing world population and ongoing climate changes. CDI is a desalination technique where salt ions are removed from a flow channel by the application of an electrical potential difference across this channel and are stored in electrical double layers. The aim of this work is to visualize and explain the charging process of CDI using a new microfluidic approach. Namely, we implement the geometry of CDI on a chip and visualize the ion distributions in the channel using fluorescence microscopy. In contrast to normal CDI, our system was operated in the absence of flow, using non-porous electrodes. By using two pH-sensitive fluorescence dyes, we found the formation of pH waves across the channel, even though the system is operated at low potential differences in order to suppress Faradaic reactions, such as water splitting. From simulations of the transport process, we found that a small current density in the order of 0.1 A m(-2) can trigger the formation of such pH waves. CDI generally benefits from large electrode areas relative to the channel cross section. However, this large area ratio will also increase the magnitude of these waves, which might lead to a reduction in desalination efficiency

Magnetoactive sponges for dynamic control of microfluidic flow patterns in microphysiological systems.

We developed a microfluidic flow-control system capable of dynamically generating various flow patterns on demand. The flow-control system is based on novel magnetoactive sponges embedded in microfluidic flow channels. Applying a non-uniform magnetic field compresses the magnetoactive sponge, significantly reducing porosity and hydraulic conductivity. Tuning the applied magnetic field can dynamically vary the flow rate in the microfluidic channel. Pulsatile and physiological flow patterns with frequency between 1 and 3 Hz, flow rates between 0.5 and 10 μL min(-1) and duration over 3 weeks have been achieved. Smooth muscle cells in engineered blood vessels perfused for 7 days aligned perpendicular to the flow direction under pulsatile but not steady flow, similar to the in vivo orientation. Owing to its various advantages over traditional flow-control methods, the new system potentially has important applications in microfluidic-based microphysiological systems to simulate the physiological nature of blood flow.

Life under flow: A novel microfluidic device for the assessment of anti-biofilm technologies

In the current study, we have developed and fabricated a novel lab-on-a-chip device for the investigation of biofilm responses, such as attachment kinetics and initial biofilm formation, to different hydrodynamic conditions. The microfluidic flow channels are designed using computational fluid dynamic simulations so as to have a pre-defined, homogeneous wall shear stress in the channels, ranging from 0.03 to 4.30 Pa, which are relevant to in-service conditions on a ship hull, as well as other man-made marine platforms. Temporal variations of biofilm formation in the microfluidic device were assessed using time-lapse microscopy, nucleic acid staining, and confocal laser scanning microscopy (CLSM). Differences in attachment kinetics were observed with increasing shear stress, i.e., with increasing shear stress there appeared to be a delay in bacterial attachment, i.e., at 55, 120, 150, and 155 min for 0.03, 0.60, 2.15, and 4.30 Pa, respectively. CLSM confirmed marked variations in colony architecture, i.e.,: (i) lower shear stresses resulted in biofilms with distinctive morphologies mainly characterised by mushroom-like structures, interstitial channels, and internal voids, and (ii) for the higher shear stresses compact clusters with large interspaces between them were formed. The key advantage of the developed microfluidic device is the combination of three architectural features in one device, i.e., an open-system design, channel replication, and multiple fully developed shear stresses.

Probing concentration-dependent behavior of DNA-binding proteins on a single-molecule level illustrated by Rad51

Low throughput is an inherent problem associated with most single-molecule biophysical techniques. We have developed a versatile tool for high-throughput analysis of DNA and DNA-binding molecules by combining microfluidic and dense DNA arrays. We use an easy-to-process microfluidic flow channel system in which dense DNA arrays are prepared for simultaneous imaging of large amounts of DNA molecules with single-molecule resolution. The Y-shaped microfluidic design, where the two inlet channels can be controlled separately and precisely, enables the creation of a concentration gradient across the microfluidic channel as well as rapid and repeated addition and removal of substances from the measurement region. A DNA array stained with the fluorescent DNA-binding dye YOYO-1 in a gradient manner illustrates the method and serves as a proof of concept. We have applied the method to studies of the repair protein Rad51 and could directly probe the concentration-dependent DNA-binding behavior of human Rad51 (HsRad51). In the low-concentration regime used (100nM HsRad51 and below), we detected binding to double-stranded DNA (dsDNA) without positive cooperativity.

Computer simulations of colloidal particles under flow in microfluidic channels

We study the propagation of single, neutrally buoyant rigid spheres under pressure-driven flow by means of extensive computer simulations that correctly account for hydrodynamic interactions. We first consider a system geometry consisting of two parallel plane walls and achieve very good agreement with experimental results [M. E. Staben and R. H. Davis, Int. J. Multiphase Flow, 2005, 31, 529]. In the second part of our analysis, we simulate the flow of tracer particles through a hexagonal array of cylindrical obstacles, whose axis lies parallel to the gradient–vorticity plane of the flow. We find that the presence of the obstacles causes a significant slowdown of the tracer particles and that their velocities respond in a highly non-linear way to an increasing pressure drop.

Measuring Lipid Bilayer Bending Energy in a Dual-Beam Optical Trap

While cell membrane bending is central to many physiological processes, techniques for measuring the bending energy of lipid bilayers have reported widely divergent results. Here, we show that a dual-beam optical trap (DBOT) can be used to apply finely controlled tensions to lipid bilayer membranes in a giant unilamellar vesicle (GUV) format. Optical force from the trap stretches the GUV; video microscopy is used to measure the change in membrane area during stretching. As laser power is increased, the surface area of the GUV also increases. Laser power is translated to membrane tension using a ray optics approach. The resulting tension-area relationship is fit to a model of membrane mechanics to yield a bending modulus.The entire DBOT system is integrated with a microfluidic flow channel in such a manner as to facilitate the high-throughput analysis of large populations of GUVs. Performing such an analysis, we have shown that the presence of cholesterol has no effect on the bending modulus of bilayers made from the unsaturated lipid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). Both pure POPC bilayers and bilayers consisting of 80% POPC, 20% cholesterol have a bending modulus around 8 kT.This technique is a promising route to detailed, accurate data relating lipid bilayer composition to membrane mechanical properties.

Leukocyte Function Antigen-1, Kindlin-3, and Calcium Flux Orchestrate Neutrophil Recruitment during Inflammation

Neutrophil arrest and migration on inflamed endothelium involves a conformational shift in CD11a/CD18 (leukocyte function antigen-1; LFA-1) to a high-affinity and clustered state that determines the strength and lifetime of bond formation with ICAM-1. Cytoskeletal adapter proteins Kindlin-3 and Talin-1 anchor clustered LFA-1 to the cytoskeleton and facilitate the transition from neutrophil rolling to arrest. We recently reported that tensile force acts on LFA-1 bonds inducing their colocalization with Orai1, the predominant membrane store operated Ca(2+) channel that cooperates with the endoplasmic reticulum to elicit cytosolic flux. Because Kindlin-3 was recently reported to initiate LFA-1 clustering in lymphocytes, we hypothesized that it cooperates with Orai1 and LFA-1 in signaling local Ca(2+) flux necessary for shear-resistant neutrophil arrest. Using microfluidic flow channels combined with total internal reflection fluorescence microscopy, we applied defined shear stress to low- or high-affinity LFA-1 and imaged the spatiotemporal regulation of bond formation with Kindlin-3 recruitment and Ca(2+) influx. Orai1 and Kindlin-3 genes were silenced in neutrophil-like HL-60 cells to assess their respective roles in this process. Kindlin-3 was enriched within focal clusters of high-affinity LFA-1, which promoted physical linkage with Orai1. This macromolecular complex functioned to amplify inside-out Ca(2+) signaling in response to IL-8 stimulation by catalyzing an increased density of Talin-1 and consolidating LFA-1 clusters within sites of contact with ICAM-1. In this manner, neutrophils use focal adhesions as mechanosensors that convert shear stress-mediated tensile force into local bursts of Ca(2+) influx that catalyze cytoskeletal engagement and an adhesion-strengthened migratory phenotype.

Pulsatile Poiseuille flows in microfluidic channels with back-and-forth mode

The numerical solver for the velocity field equation describing laminar pulsatile flows driven by a timedependent pressure drop in the straight microfluidic channel of square cross-section is developed. In the computational algorithm, an orthogonal collocation on finite element scheme for spatial discretizations is combined with an adaptive Runge-Kutta method for time integration. The algorithm with the 1,521 computational nodes and the accuracy up to O(10 –5 ) is applied to the flow in the back-and-forth standing mode with the channel hydraulic diameter (Dh) in the range 10 – 500 µm and the oscillating frequency (f) of 1 to 100 Hz. As a result, a periodic steady state is defined as the flow condition where there would be no net movement after long time elapses. Following by the retardation phenomena in a cycle, reversal of the axial velocity is observed at the channel center. Major attention is focused on the influences of the size of channel cross-section and the oscillating frequency. Increasing Dh and f results in the decrease in the amplitude of mean velocity but the increase in the start-up time. Larger time delay occurs by low-frequency pulsation.

Fusion of single proteoliposomes with planar, cushioned bilayers in microfluidic flow cells

Many biological processes rely on membrane fusion, and therefore assays to study its mechanisms are necessary. Here we report an assay with sensitivity to single-vesicle, and even to single-molecule events using fluorescently labeled vesicle-associated v-SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) liposomes and target-membrane-associated t-SNARE-reconstituted planar, supported bilayers (t-SBLs). Docking and fusion events can be detected using conventional far-field epifluorescence or total internal reflection fluorescence microscopy. In this assay, fusion is dependent on SNAP-25, one of the t-SNARE subunits that is required for fusion in vivo. The success of the assay is due to the use of: (i) bilayers covered with a thin layer of poly(ethylene glycol) (PEG) to control bilayer-bilayer and bilayer-substrate interactions, and (ii) microfluidic flow channels that present many advantages, such as the removal of nonspecifically bound liposomes by flow. The protocol takes 6-8 d to complete. Analysis can take up to 2 weeks.

FT-IR Spectroscopic Imaging of Reactions in Multiphase Flow in Microfluidic Channels

Rapid, in situ, and label-free chemical analysis in microfluidic devices is highly desirable. FT-IR spectroscopic imaging has previously been shown to be a powerful tool to visualize the distribution of different chemicals in flows in a microfluidic device at near video rate imaging speed without tracers or dyes. This paper demonstrates the possibility of using this imaging technology to capture the chemical information of all reactants and products at different points in time and space in a two-phase system. Differences in the rates of chemical reactions in laminar flow and segmented flow systems are also compared. Neutralization of benzoic acid in decanol with disodium phosphate in water has been used as the model reaction. Quantitative information, such as concentration profiles of reactant and products, can be extracted from the imaging data. The same feed flow rate was used in both the laminar flow and segmented flow systems. The laminar flow pattern was achieved using a plain wide T-junction, whereas the segmented flow was achieved by introducing a narrowed section and a nozzle at the T-junction. The results show that the reaction rate is limited by diffusion and is much slower with the laminar flow pattern, whereas the reaction is completed more quickly in the segmented flow due to better mixing.

Mobility Gradient Induces Cross-Streamline Migration of Semiflexible Polymers.

Many aspects of modern material science and biology rely on the strategic manipulation and understanding of polymer dynamics in confining micro- and nanoflow. We directly observe and analyze nonequilibrium structural and dynamic properties of individual semiflexible actin filaments in pressure-driven microfluidic channel flow using fluorescence microscopy. Different conformational shapes, such as filaments fluctuating in an elongated manner, parabolically bent, as well as tumbling, are identified. With increasing flow velocity, a strong center-of-mass migration toward the channel walls is observed. This significant migration effect can be explained by a shear rate dependent spatial diffusivity due to a gradient in chain mobility of the semiflexible polymers.

Investigating the Effects of Dynamic External Stimuli on Single Cell Fitness and Gene Expression in Escherichia Coli

In this work, we study the bacterial cellular response to time-dependent external stimuli in single living cells. We developed a microfluidic platform for single cell analysis that allows for dynamic control of well-defined environmental growth conditions and culture media. Using this platform, we studied the effect of small molecule inducers on gene expression in the lac operon using fluorescent reporter proteins and cell growth rates as a proxy of cellular fitness. We applied temporally varying inducer concentrations by translating single cells between two adjacent fluid streams containing either growth medium or growth medium and inducer. We observed that single cell gene expression depends on growth rate and frequency of exposure to inducer concentrations. Single cell induction experiments are compared to control experiments with and without continuous fluid flow in microfluidic channels. For these experiments, single cell analysis is facilitated by a microfluidic-based hydrodynamic trap recently developed in our lab. The hydrodynamic trap enables confinement and manipulation of single cells in free solution using the sole action of fluid flow. Automated feedback control is integrated into the device using an “on-chip” valve, which allows for precise confinement of cells in free solution. Using this device, cells are confined at a fluid stagnation point generated in a cross-slot microfluidic geometry, thereby enabling non-perturbative trapping of cells for long time scales. Using optical microscopy, we observe single Escherichia coli cells growing and dividing both at room temperature and at 370C, and cell division rates in the microfluidic trap compare favorably to the growth rates of E. coli measured in bulk studies. We anticipate that the microfluidic-based trap is an ideal platform to study cellular regulation and gene network dynamics of single living cells in free solution.

Microfluidic synthesis of multifunctional Janus particles for biomedical applications.

Multifunctional Janus particles have a variety of applications in a wide range of fields. However, to achieve many of these applications, high-throughput, low-cost techniques are needed to synthesize these particles with precise control of the various structural/physical/chemical properties. Microfluidics provides a unique platform to fabricate Janus particles using carefully controlled liquid flow in microfluidic channels to form Janus droplets and various types of solidification methods to solidify them into Janus particles. In this Focus article, we summarize the most recent representative works on Janus particle fabrication in microfluidics. The applications of Janus particles in biomedical areas are emphasized. We believe that microfluidics-enabled multifunctional Janus particles could resolve multiple prevalent issues in biomedicine (e.g., disease monitoring at an early stage, high-throughput bioassays, therapeutic delivery) if persistent effort and collaboration are devoted to this direction.

Coalescence dynamics of surfactant-stabilized emulsions studied with microfluidics

We report the results of a study on emulsion stability in a microfluidic channel flow using an integrated microfluidic device. The microfluidic circuit enables production of a monodisperse oil-in-water emulsion and monitoring of emulsion stability upon shear-induced collisions. Sodium-n-dodecyl sulfate was used as emulsifier, and hexadecane as dispersed phase. We measure the mean drop size at the end of the collision channel as a function of the surfactant and sodium chloride bulk concentration, and as a function of the total flow rate. We find that emulsions are stable against coalescence for SDS bulk concentrations down to 10−6 M within the residence time of the droplets in the channel in the absence of added NaCl. The stability of the emulsion at these low SDS bulk concentrations is interpreted in terms of a reduced mobility of the droplet interfaces, which slows down drainage of the film of the continuous phase between the droplets. Emulsions stabilized by SDS with added NaCl in the continuous phase display a transition from a stable to unstable regime when increasing the NaCl bulk concentration from 0.1 M to 0.3 M, which is in agreement with predictions using simple DLVO force calculations for colloidal stability. We also estimate the characteristic coalescence time between droplets using a simple coalescence theory and compare the results with values obtained by us previously from trajectory analysis of colliding droplet pairs.

Controlling flow in microfluidic channels with a manually actuated pin valve

There is a need for a simple method to control fluid flow within microfluidic channels. To meet this need, a simple push pin with a polydimethylsiloxane (PDMS) tip has been integrated into microfluidic networks to be placed within the microchannel to obstruct flow. This new valve design can attain on/off control of fluid flow without an external power source using readily-available, low-cost materials. The valve consists of a 14 gauge (1.6 mm) one inch piece of metal tubing with a PDMS pad at the tip to achieve a fluidic seal when pressed against a microfluidic channel's substrate. The metal tubing or pin is then either manually pushed down to block or pulled up to allow fluid flow. The valve was validated using a pressure transducer and fluorescent dye to determine the breakthrough pressure the valve can withstand over multiple cycles. In the first cycle, the median value for pressure withstood by the valve was 8.8 psi with a range of 17.5-2.7 psi. The pressure the valves were able to withstand during each successive trial was lower suggesting they may be most valuable as a method to control the initial introduction of fluids into a microfluidic device. These valves can achieve flow regulation within microfluidic devices, have a small dead volume, and are simple to fabricate and use, making this technique widely suitable for a range of applications.

Validation of Screening Assays for Ligand Gated ion Channels on Qpatch HTX

QPatch HTX has the highest throughput in the QPatch series. With signature features such as unattended operations, eight pipettes and the integrated microfluidic glass flow channels of the QPlates, the QPatch HTXis well suited for testing ligand gated ion channels. The recent advent of a QPlate with 10 patch holes per measurement site, instead of the classical single patch holehas minimized problems with low expression or problems with stability of the recodings over time. The success rate of recording on QPatch HTX is therefore approaching 100% for a number of different applications. We have previously shown that compound profiling on a wide variety of ligand-gated ion channels is certainly possible on the QPatch. With the increased stability and success rates of QPatch HTX, we demonstrate that the QPatch also can be used as an efficient screening tool for ligand gated ion channels. In this study, we have tested 3 different ion channels in a screening assay on QPatch HTX: GABA α1β1γ2 (γ-amino butyric acid receptor), ASIC a1 (acid sensing ion channel) and nAchR α1 (nicotinic acetylcholine receptor). These assays each have their own unique set of challenges: Low functional expression, extremely high elicited current amplitudes, and fast desensitization respectively. In this poster we present data to compare throughput and assay quality between the QPatch HTX in multi-hole mode and the classic single-hole mode in a screening scenario.

Pumping-induced perturbation of flow in microfluidic channels and its implications for on-chip cell culture

We study the rate of response to changes in the rate of flow and the perturbations in flow in polydimethylsiloxane (PDMS) microfluidic chips that are subjected to several common flow-control systems. We find that the flow rate of liquid delivered from a syringe pump equipped with a glass syringe responds faster to the changes in the conditions of flow than the same liquid delivered from a plastic syringe; and the rate of flow delivered from compressed air responds faster than that from a glass syringe. We discover that the rate of flow that is driven by a syringe pump and regulated by an integrated pneumatic valve responds even faster, but this flow-control method is characterized by large perturbations. We also examine the possible effects of these large perturbations on NIH 3T3 cells in microfluidic channels and find that they could cause the detachment of NIH 3T3 cells in the microchannels.

Transient deflection response in microcantilever array integrated with polydimethylsiloxane (PDMS) microfluidics

We report the integration of a nanomechanical sensor consisting of 16 silicon microcantilevers with polydimethylsiloxane (PDMS) microfluidics. For microcantilevers positioned near the bottom of a microfluidic flow channel, a transient differential analyte concentration for the top versus bottom surface of each microcantilever is created when an analyte-bearing fluid is introduced into the flow channel (which is initially filled with a non-analyte containing solution). We use this effect to characterize a bare (nonfunctionalized) microcantilever array in which the microcantilevers are simultaneously read out with our recently developed high sensitivity in-plane photonic transduction method. We first examine the case of non-specific binding of bovine serum albumin (BSA) to silicon. The average maximum transient microcantilever deflection in the array is -1.6 nm, which corresponds to a differential surface stress of only -0.23 mN m(-1). This is in excellent agreement with the maximum differential surface stress calculated based on a modified rate equation in conjunction with finite element simulation. Following BSA adsorption, buffer solutions with different pH are introduced to further study microcantilever array transient response. Deflections of 20-100 nm are observed (2-14 mN m(-1) differential surface stress). At a flow rate of 5 μL min(-1), the average measured temporal width (FWHM) of the transient response is 5.3 s for BSA non-specific binding and 0.74 s for pH changes.