Continuous separation of particles using a microfluidic device equipped with flow rate control valves

We propose herein an improved microfluidic system for continuous and precise particle separation. We have previously proposed a method for particle separation called "pinched flow fractionation." Using the previously reported method, particles can be continuously separated according to differences in their diameters, simply by introducing liquid flows with and without particles into a specific microchannel structure. In this study, we incorporated PDMS membrane microvalves for flow rate control into the microfluidic device to improve the separation accuracy. By adjusting the flow rates distributed to each outlet, target particles could be precisely collected from the desired outlet. We succeeded in separating micron and submicron-size polymer particles. This method can be used widely for continuous and precise separation of various kinds of particles, and can function as an important part of microfluidic systems.     

Continuous and size-dependent sorting of emulsion droplets using hydrodynamics in pinched microchannels

In this report, a microfluidic system is presented for continuous and size-dependent separation of droplets utilizing microscale hydrodynamics. The separation scheme is based on laminar-flow focusing and spreading in a pinched microchannel, referred to as "pinched flow fractionation (PFF)", which was previously developed for the size-dependent separation of solid particles, such as polymer microparticles or cells. By simply introducing emulsion and the continuous phase into a microchannel, continuous separation could be achieved without using complicated operations or devices. We first examined whether this scheme could be applied for droplets, by using a pinched microchannel with one outlet, and observed the behaviors of monodisperse droplets generated at the upstream T-junction. Analysis via high-speed imaging revealed that the length of the pinched segment is critical for precise separation of droplets. Then, separation of a polydisperse oil-in-water emulsion that was prepared previously was demonstrated using a microfluidic device equipped with multiple outlets. These results showed the ability of the presented system to sort or select specific-sized droplets easily and accurately, which would be difficult to achieve using normal-scale schemes, such as centrifugation or filtration.     

Continuous particle separation in a microchannel having asymmetrically arranged multiple branches

A new method for continuous size separation and collection of particles in microfabricated devices, asymmetric pinched flow fractionation (AsPFF), has been proposed and demonstrated. This method improves the separation scheme of pinched flow fractionation (PFF), which utilizes a laminar flow profile inside a microchannel. In this study, multiple branch channels with different channel dimensions were arranged at the end of the pinched segment, so that the flow rate distributions to each branch channel were varied, and a large part of the liquid was forced to go through one branch channel (drain channel). In the proposed channel system, the flow profile inside the microchannel was asymmetrically amplified, enabling the separation of one-order smaller particles compared with PFF. After introducing the method, we examined the effect of the asymmetric amplification by controlling the outlet of the drain channel. Also, a mixture of 1.0 approximately 5.0 microm particles was separated, and erythrocytes were successfully separated from blood. The results indicate that the AsPFF method could be applied to the separation of much smaller-size particles, since more precise separation can be achieved simply by changing the geometries of branch channels.     

Continuous dielectrophoretic separation of particles in a spiral microchannel

Particle separation is a fundamental operation in the areas of biology and physical chemistry. A variety of force fields have been used to separate particles in microfluidic devices, among which electric field may be the most popular one due to its general applicability and adaptability. So far, however, electrophoresis‐based separations have been limited primarily to batchwise processes. Dielectrophoresis (DEP)‐based separations require in‐channel micro‐electrodes or micro‐insulators to produce electric field gradients. This article introduces a novel particle separation technique in DC electrokinetic flow through a planar double‐spiral microchannel. The continuous separation arises from the cross‐stream dielectrophoretic motion of particles induced by the non‐uniform electric field inherent to curved channels. Specifically, particles are focused by DEP to one sidewall of the first spiral, and then dielectrophoretically deflected toward the other sidewall of the second spiral at a particle‐dependent rate, leading to focused particle streams along different flow paths. This DEP‐based particle separation technique is demonstrated in an asymmetric double‐spiral microchannel by continuously separating a mixture of 5/10 μm particles and 3/5 μm particles.     

Inertial separation in a contraction-expansion array microchannel

We report a contraction-expansion array (CEA) microchannel that allows inertial size separation by a force balance between inertial lift and Dean drag forces in fluid regimes in which inertial fluid effects become significant. An abrupt change of the cross-sectional area of the channel curves fluid streams and produces a similar effect compared to Dean flows in a curved microchannel of constant cross-section, thereby inducing Dean drag forces acting on particles. In addition, the particles are influenced by inertial lift forces throughout the contraction regions. These two forces act in opposite directions each other throughout the CEA microchannel, and their force balancing determines whether the particles cross the channel, following Dean flows. Here we describe the physics and design of the CEA microfluidic device, and demonstrate complete separation of microparticles (polystyrene beads of 4 and 10 μm in diameter) and efficient exchange of the carrier medium while retaining 10 μm beads.     

Brownian dynamics simulation and experimental study of colloidal particle deposition in a microchannel flow

This paper reports an analysis of the irreversible deposition of colloidal particles from the pressure-driven flow in a microchannel within the framework of DLVO theory. A theoretical model is presented on the basis of the stochastic Langevin equation, incorporating the random Brownian motion of colloidal particles. Brownian dynamics simulation is used to compute the particle deposition in terms of the surface coverage. To validate the theoretical model, experiments are carried out using the parallel-plate flow cell technique, enabling direct videomicroscopic observation of the deposition kinetics of polystyrene latex particles in NaCl electrolytes. The theoretical predictions are compared with the experimental results, and good agreement is found.     

A velocity program using the Kanade–Lucas–Tomasi feature-tracking algorithm with demonstration for pressure and electroosmosis conditions

Investigating microfluidic flow profiles is of interest in the microfluidics field for the determination of various characteristics of a lab-on-a-chip system. Microparticle tracking velocimetry uses computational methods upon recording video footage of microfluidic flow to ultimately visualize motion within a microfluidic system across all frames of a video. Current methods are computationally expensive or require extensive instrumentation. A computational method suited to microparticle tracking applications is the robust Kanade–Lucas–Tomasi (KLT) feature-tracking algorithm. This work explores a microparticle tracking velocimetry program using the KLT feature-tracking algorithm. The developed program is demonstrated using pressure-driven and EOF and compared with the respective mathematical fluid flow models. An electrostatics analysis of EOF conditions is performed in the development of the mathematical using a Poisson's Equation solver. This analysis is used to quantify the zeta potential of the electroosmotic system. Overall, the KLT feature-tracking algorithm presented in this work proved to be highly reliable and computationally efficient for investigations of pressure-driven and EOF in a microfluidic system.     

Orthogonal optical force separation simulation of particle and molecular species mixtures under direct current electroosmotic driven flow for applications in biological sample preparation

Presented here are the results from numerical simulations applying optical forces orthogonally to electroosmotically induced flow containing both molecular species and particles. Simulations were conducted using COMSOL v4.2a Multiphysics® software including the particle tracking module. The study addresses the application of optical forces to selectively remove particulates from a mixed sample stream that also includes molecular species in a pinched flow microfluidic device. This study explores the optimization of microfluidic cell geometry, magnitude of the applied direct current electric field, EOF rate, diffusion, and magnitude of the applied optical forces. The optimized equilibrium of these various contributing factors aids in the development of experimental conditions and geometry for future experimentation as well as directing experimental expectations, such as diffusional losses, separation resolution, and percent yield. The result of this work generated an optimized geometry with flow conditions leading to negligible diffusional losses of the molecular species while also being able to produce particle removal at near 100% levels. An analytical device, such as the one described herein with the capability to separate particulate and molecular species in a continuous, high‐throughput fashion would be valuable by minimizing sample preparation and integrating gross sample collection seamlessly into traditional analytical detection methods.     

Chiral separations in capillary high-performance liquid chromatography and nonaqueous capillary electrochromatography using helically chiral poly(diphenyl-2-pyridylmethyl methacrylate) as chiral stationary phase.

Enantioseparations in nonaqueous capillary electrochromatography (CEC) are reported in this study for the first time, using wide-pore aminopropyl silica gel coated with helically chiral poly(diphenyl-2-pyridylmethyl methacrylate) (PDPM) as chiral stationary phase (CSP). The anodic electroosmotic flow (EOF) in a methanolic solution of ammonium acetate was used for the migration of neutral analytes through the packed bed in the capillaries. Four different techniques, high-performance liquid chromatography (HPLC) in common-size columns, capillary HPLC, pressure-assisted CEC and CEC were compared from the viewpoint of separation parameters. The latter three were performed with the same experimental setup, varying the relative contribution of the pressure-driven and the electrically driven flow to the overall mobility of the analyte. Capillary HPLC offers clear advantages compared to enantioseparations in common-size columns. However, for a given particle size of the packing material, CEC was not obviously advantageous compared to pressure-driven separations.     

A continuous DC-insulator dielectrophoretic sorter of microparticles

A lab-on-a-chip device is described for continuous sorting of fluorescent polystyrene microparticles utilizing direct current insulating dielectrophoresis (DC-iDEP) at lower voltages than previously reported. Particles were sorted by combining electrokinetics and dielectrophoresis in a 250 μm wide PDMS microchannel containing a rectangular insulating obstacle and four outlet channels. The DC-iDEP particle flow behaviors were investigated with 3.18, 6.20 and 10 μm fluorescent polystyrene particles which experience negative DEP forces depending on particle size, DC electric field magnitude and medium conductivity. Due to negative DEP effects, particles are deflected into different outlet streams as they pass the region of high electric field density around the obstacle. Particles suspended in dextrose added phosphate buffer saline (PBS) at conductivities ranging from 0.50 to 8.50 mS/cm at pH 7.0 were compared at 6.85 and 17.1 V/cm. Simulations of electrokinetic and dielectrophoretic forces were conducted with COMSOL Multiphysics^® to predict particle pathlines. Experimental and simulation results show the effect of medium and voltage operating conditions on particle sorting. Further, smaller particles experience smaller iDEP forces and are more susceptible to competing nonlinear electrostatic effects, whereas larger particles experience greater iDEP forces and prefer channels 1 and 2. This work demonstrates that 6.20 and 10 μm particles can be independently sorted into specific outlet streams by tuning medium conductivity even at low operating voltages. This work is an essential step forward in employing DC-iDEP for multiparticle sorting in a continuous flow, multiple outlet lab-on-a-chip device.     

Sheathless elasto-inertial particle focusing and continuous separation in a straight rectangular microchannel

Particle focusing in planar geometries is essentially required in order to develop cost-effective lab-on-a-chips, such as cell counting and point-of-care (POC) devices. In this study, a novel method for sheathless particle focusing, called "Elasto-Inertial Particle Focusing", was demonstrated in a straight microchannel. The particles were notably aligned along the centerline of the straight channel under a pressure-driven flow without any additional external force or apparatus after the addition of an elasticity enhancer: PEO (poly(ethylene oxide)) (～O(100) ppm). As theoretically predicted (elasticity number: El≈O(100)), multiple equilibrium positions (centerline and corners) were observed for the viscoelastic flow without inertia, whereas three-dimensional particle focusing only occurred when neither the elasticity nor the inertia was negligible. Therefore, the three-dimensional particle focusing mechanism was attributed to the synergetic combination of the elasticity and the inertia (elasticity number: El≈O(1-10)). Furthermore, from the size dependence of the elastic force upon particles, we demonstrated that a mixture of 5.9 and 2.4 μm particles was separated at the exit of the channel in viscoelastic flows. We expect that this method can contribute to develop the miniaturized flow cytometry and microdevices for cell and particle manipulation.     

DC dielectrophoresis separation of marine algae and particles in a microfluidic chip

This paper reports a microfluidic method of continuous separation of marine algae and particles by DC dielectrophoresis. The locally non-uniform electric field is generated by an insulating PDMS triangle hurdle fabricated within a PDMS microchannel. Both the particles and algae are subject to negative DEP forces at the hurdle where the gradient of local electric-field strength is the strongest. The DEP force acting on the particle or the algae depends on particles’ or algae’s volume, shape and dielectric properties. Thus the moving particles and algae will be repelled to different streamlines when passing the hurdle. In this way, combined with the electroosmotic flow, continuous separation of algae of two different sizes, and continuous separation of polystyrene particles and algae with similar volume but different shape were achieved. This first demonstration of DC DEP separation of polystyrene particles and algae with similar sizes illustrates the great influence of dielectric properties on particle separation and potentials for sample pretreatment.     

Controlled protein adsorption on a silica microparticle

In the present study, controlled protein adsorption on a rigid silica microparticle is investigated numerically using classical Langmuir and two-state models under electrokinetic flow conditions. The instantaneous particle locations are simulated along a straight microchannel using an arbitrary Lagrangian−Eulerian framework in the finite element method for the electrophoretic motion of the charged particle. Within the scope of the parametric study, the strength of the external electric field (E), particle diameter (D_p), the zeta potential of the particle (ζ_p), and the location of the microparticle away from the channel wall (H) are systematically varied. The results are also compared to the data of pressure-driven flow having a parabolic flow profile at the inlet whose maximum magnitude is set to the particle's electrophoretic velocity magnitude. The validation studies reveal that the code developed for the particle motion in the present simulations agrees well with the experimental results. It is observed that protein adsorption can be controlled using electrokinetic phenomena. The plug-like flow profile in electrokinetics is beneficial for a microparticle at every spatial location in the microchannel, whereas it is not valid for the pressure-driven flow. The electric field strength and the zeta potential of the particle accelerate the protein adsorption. The wall shear stress and shear rate are good indicators to predict the adsorption process for electrokinetic flow.     

Increased size‐sorting performance in gravitational SPLITT by using a pinched sample inlet design

Split‐flow thin fractionation is a continuous, flow‐assisted separation technique for sorting macromolecules and particulate matter on a preparative scale. On reducing the thickness of the sample inlet conduit of a gravitational split‐flow thin fractionation channel, size‐sorting performance is found to increase since particles that are continuously fed into the channel can be more rapidly compressed toward the upper wall of the channel. Experiments are carried out by measuring the number percentage of particles eluted at each outlet as a function of different thickness values of the sample inlet conduit. The effects that the total thickness of the gravitational split‐flow thin fractionation channel and the sample feed concentration have on the size‐fractionation performance are examined with the goal of determining the best pinched sample inlet, gravitational split‐flow thin fractionation channel design.     

Precise measurement and control of the pressure-driven flows for microfluidic systems

The pressure-driven device is designed and the flow rates of the microfluidic systems can be supplied by the pressure-driven flows, which can significantly reduce the flow-rate fluctuations coming from the pump source. For pressure-driven flows, the flow rates of the fluids can be predicted by measuring the pressure drop along a polytetrafluoroethylene (PTFE) tubing. Especially, by varying the geometrical parameters of the PTFE tubing, the predicted flow rates of the fluids are compared with the experimental measurements, and the testing precision of the pressure-driven flows can be obtained. Meanwhile, the dynamic characteristics of the open-loop and closed-loop control pressure-driven device are comparatively studied. Particularly, a proportional and integral (PI) controller is integrated with the closed-loop control pressure-driven device, and the effects of the parameters of the PI controller on the dynamic characteristics of the pressure-driven devices are mainly discussed. Most importantly, by improving the dynamic characteristics of the pressure-driven devices, precise measurement and control of the pressure-driven flows can be achieved for microfluidic systems.     

Electroosmotic flow velocity measurements in a square microchannel

Experiments were performed using a microparticle image velocimetry (MPIV) for 2D velocity distributions of electroosmotically driven flows in a 40-mm-long microchannel with a square cross section of 200×200 μm. Electroosmotic flow (EOF) bulk fluid velocity measurements were made in a range of streamwise electric field strengths from 5 to 25 kV/m. A series of seed particle calibration tests can be made in a 200×120×24,000-μm untreated polydimethyl siloxane (PDMS channel incorporating MPIV to determine the electrophoretic mobilities in aqueous buffer solutions of 1× TAE, 1× TBE, 10 mM NaCl, and 10 mM borate. A linear/nonlinear (due to Joule heating) flow rate increase with applied field was obtained and compared with those of previous studies. A parametric study, with extensive measurements, was performed with different electric field strength and buffer solution concentration under a constant zeta potential at wall for each buffer. The characteristics of EOF in square microchannels were thus investigated. Finally, a composite correlation of the relevant parameters was developed in the form of within ±1% accuracy for 99% of the experimental data.     

Negative dielectrophoresis-based particle separation by size in a serpentine microchannel

Dielectrophoresis has been widely used to focus, trap, concentrate, and sort particles in microfluidic devices. This work demonstrates a continuous separation of particles by size in a serpentine microchannel using negative dielectrophoresis. Depending on the magnitude of the turn-induced dielectrophoretic force, particles travelling electrokinetically through a serpentine channel either migrate toward the centerline or bounce between the two sidewalls. These distinctive focusing and bouncing phenomena are utilized to implement a dielectrophoretic separation of 1 and 3 μm polystyrene particles under a DC-biased AC electric field of 880 V/cm on average. The particle separation process in the entire microchannel is simulated by a numerical model.     

Enhancement by optical force of separation in pinched flow fractionation

A method for improving the size-based particle separation technique known as pinched flow fractionation (PFF) has been demonstrated experimentally and analyzed by performing numerical calculations. Since the particles in the pinched region are pushed by an optical scattering force, the original particle position with respect to the wall is modulated. This position modulation in the pinched region is amplified in the broadened region along the streamline. This enhancement of separation is achieved by imposing an optical force on the original PFF design. Three different polystyrene latexes (PSLs) with diameters of 2, 5, and 10 μm were separated with PFF and optically enhanced PFF (OEPFF) devices. The separations achieved with the two devices were compared and enhancements in the separation distance by factors of up to approximately 15 were achieved. Theoretical calculations were also performed to interpret these results.     

Hydrodynamic filtration for on-chip particle concentration and classification utilizing microfluidics

We propose here a new method for continuous concentration and classification of particles in microfluidic devices, named hydrodynamic filtration. When a particle is flowing in a microchannel, the center position of the particle cannot be present in a certain distance from sidewalls, which is equal to the particle radius. The proposed method utilizes this fact, and is performed using a microchannel having multiple side branch channels. By withdrawing a small amount of liquid repeatedly from the main stream through the side channels, particles are concentrated and aligned onto the sidewalls. Then the concentrated and aligned particles can be collected according to size through other side channels (selection channels) in the downstream of the microchannel. Therefore, continuous introduction of a particle suspension into the microchannel enables both particle concentration and classification at the same time. In this method, the flow profile inside a precisely fabricated microchannel determines the size limit of the filtered substances. So the filtration can be performed even when the channel widths are much larger than the particle size, without the problem of channel clogging. In this study, concentrations of polymer microspheres with diameters of 1-3 microm were increased 20-50-fold, and they were collected independently according to size. In addition, selective enrichment of leukocytes from blood was successfully performed.     

Stop-flow lithography in a microfluidic device

Polymeric particles in custom designed geometries and with tunable chemical anisotropy are expected to enable a variety of new technologies in diverse areas such as photonics, diagnostics and functional materials. We present a simple, high throughput and high resolution microfluidic method to synthesize such polymeric particles. Building off earlier work that we have done on continuous flow lithography (CFL) (D. Dendukuri, D. C. Pregibon, J. Collins, T. A. Hatton, P. S. Doyle, Nat. Mater., 2006, 5, 365-369; ref. 1), we have devised and implemented a new setup that uses compressed air driven flows in preference to syringe pumps to synthesize particles using a technique that we call stop-flow lithography (SFL). A flowing stream of oligomer is stopped before polymerizing an array of particles into it, providing for much improved resolution over particles synthesized in flow. The formed particles are then flushed out at high flow rates before the cycle of stop-polymerize-flow is repeated. The high flow rates enable orders-of-magnitude improvements in particle throughput over CFL. However, the deformation of the PDMS elastomer due to the imposed pressure restricts how quickly the flow can be stopped before each polymerization event. We have developed a simple model that captures the dependence of the time required to stop the flow on geometric parameters such as the height, length and width of the microchannel, as well as on the externally imposed pressure. Further, we show that SFL proves to be superior to CFL even for the synthesis of chemically anisotropic particles with sharp interfaces between distinct sections.