Optimization of an electrokinetic mixer for microfluidic applications

This work is concerned with the investigation of the concentration fields in an electrokinetic micromixer and its optimization in order to achieve high mixing rates. The mixing concept is based on the combination of an alternating electrical excitation applied to a pressure-driven base flow in a meandering microchannel geometry. The electrical excitation induces a secondary electrokinetic velocity component, which results in a complex flow field within the meander bends. A mathematical model describing the physicochemical phenomena present within the micromixer is implemented in an in-house finite-element-method code. We first perform simulations comparable to experiments concerned with the investigation of the flow field in the bends. The comparison of the complex flow topology found in simulation and experiment reveals excellent agreement. Hence, the validated model and numerical schemes are employed for a numerical optimization of the micromixer performance. In detail, we optimize the secondary electrokinetic flow by finding the best electrical excitation parameters, i.e., frequency and amplitude, for a given waveform. Two optimized electrical excitations featuring a discrete and a continuous waveform are discussed with respect to characteristic time scales of our mixing problem. The results demonstrate that the micromixer is able to achieve high mixing degrees very rapidly.     

A simplified design of the staggered herringbone micromixer for practical applications

We demonstrated a simple method for the device design of a staggered herringbone micromixer (SHM) using numerical simulation. By correlating the simulated concentrations with channel length, we obtained a series of concentration versus channel length profiles, and used mixing completion length L_m as the only parameter to evaluate the performance of device structure on mixing. Fluorescence quenching experiments were subsequently conducted to verify the optimized SHM structure for a specific application. Good agreement was found between the optimization and the experimental data. Since L_m is straightforward, easily defined and calculated parameter for characterization of mixing performance, this method for designing micromixers is simple and effective for practical applications.     

Inducing self-rotation of cells with natural and artificial melanin in a linearly polarized alternating current electric field

The phenomenon of self-rotation observed in naturally and artificially pigmented cells under an applied linearly polarized alternating current (non-rotating) electrical field has been investigated. The repeatable and controllable rotation speeds of the cells were quantified and their dependence on dielectrophoretic parameters such as frequency, voltage, and waveform was studied. Moreover, the rotation behavior of the pigmented cells with different melanin content was compared to quantify the correlation between self-rotation and the presence of melanin. Most importantly, macrophages, which did not originally rotate in the applied non-rotating electric field, began to exhibit self-rotation that was very similar to that of the pigmented cells, after ingesting foreign particles (e.g., synthetic melanin or latex beads). We envision the discovery presented in this paper will enable the development of a rapid, non-intrusive, and automated process to obtain the electrical conductivities and permittivities of cellular membrane and cytoplasm in the near future.     

An efficient micromixer based on multidirectional vortices due to baffles and channel curvature

An efficient planar micromixer based on multidirectional vortices in a curved channel with radial baffles is proposed and examined in this work. The curvature of the microchannel and the radial baffles induce vortices in different directions. The multidirectional vortices and the converging-diverging flow caused by the baffles contribute together to the enhancement of mixing. The micromixer is fabricated with polydimethylsiloxane by a single planar microlithography process and the mixing behaviors are observed by a confocal spectral microscope imaging system to validate the simulation obtained by a commercial code. The simulation and experimental results are in reasonable agreement. The concentration distributions and flow patterns obtained reveal the following trends. (i) The mixing efficiency of the basic C-shaped micromixer with the first baffle attached to the internal cylinder and the second attached to the external cylinder is better than that of the C-shaped micromixer with inverted arrangement of baffles. (ii) When the radius of the curved channel and the width of the passage between the baffle and the cylindrical wall are small enough and the Reynolds number (Re) is large enough, an extra separation vortex develops in the downstream of the second baffle. This phenomenon is one of the reasons of trend (i). (iii) A micromixer consisting of a few basic C-shaped micromixers connected by straight channels may generate a high degree of mixing for the case with a large Re.     

Integrated microfluidic chip for rapid DNA digestion and time-resolved capillary electrophoresis analysis

An integrated microfluidic chip is proposed for rapid DNA digestion and time-resolved capillary electrophoresis (CE) analysis. The chip comprises two gel-filled chambers for DNA enrichment and purification, respectively, a T-form micromixer for DNA/restriction enzyme mixing, a serpentine channel for DNA digestion reaction, and a CE channel for on-line capillary electrophoresis analysis. The DNA and restriction enzyme are mixed electroomostically using a pinched-switching DC field. The experimental and numerical results show that a mixing performance of 97% is achieved within a distance of 1 mm from the T-junction when a driving voltage of 90 V/cm and a switching frequency of 4 Hz are applied. Successive mixing digestion and capillary electrophoresis operation clearly present the changes on digesting φx-174 DNA in different CE runs. The time-resolved electropherograms show that the proposed device enables a φx-174 DNA sample comprising 11 fragments to be concentrated and analyzed within 24 min. Overall, the results presented in this study show that the proposed microfluidic chip provides a rapid and effective tool for DNA digestion and CE analysis applications.     

Studying enzymatic bioreactions in a millisecond microfluidic flow mixer

In this study, the pre-steady state development of enzymatic bioreactions using a microfluidic mixer is presented. To follow such reactions fast mixing of reagents (enzyme and substrate) is crucial. By using a highly efficient passive micromixer based on multilaminar flow, mixing times in the low millisecond range are reached. Four lamination layers in a shallow channel reduce the diffusion lengths to a few micrometers only, enabling very fast mixing. This was proven by confocal fluorescence measurements in the channel’s cross sectional area. Adjusting the overall flow rate in the 200 μm wide and 900 μm long mixing and observation channel makes it possible to investigate enzyme reactions over several seconds. Further, the device enables changing the enzyme/substrate ratio from 1:1 up to 3:1, while still providing high mixing efficiency, as shown for the enzymatic hydrolysis using β-galactosidase. This way, the early kinetics of the enzyme reaction at multiple enzyme/substrate concentrations can be collected in a very short time (minutes). The fast and easy handling of the mixing device makes it a very powerful and convenient instrument for millisecond temporal analysis of bioreactions.     

Focusing-enhanced mixing in microfluidic channels

A focusing-based microfluidic mixer was studied. The micromixer utilizes the focusing process required for cytometry to reduce the diffusion distance of molecules to be mixed in order to facilitate the passive diffusion-controlled mixing process. It was found that both the high flow rate ratio of the sheath flow to the flows to be mixed and the low flow rate of the mixing fluids resulted in the short mixing length required within the microfluidic channel. It was shown that a complete mixing was achieved within a distance of 4 mm in the micromixer for the focused mixing fluids at a flow rate of 2 μl∕min and a flow rate ratio of the sheath flow to the flows to be mixed at 4:1. The mixer described here is simple and can be easily fabricated and controlled.     

Micromixing visualization and quantification in a microscale multi-inlet vortex nanoprecipitation reactor using confocal-based reactive micro laser-induced fluorescence

A technique for visualizing and quantifying reactive mixing for laminar and turbulent flow in a microscale chemical reactor using confocal-based microscopic laser induced fluorescence (confocal μ-LIF) was demonstrated in a microscale multi-inlet vortex nanoprecipitation reactor. Unlike passive scalar μ-LIF, the reactive μ-LIF technique is able to visualize and quantify micromixing effects. The confocal imaging results indicated that the flow in the reactor was laminar and steady for inlet Reynolds numbers of 10, 53, and 93. Mixing and reaction were incomplete at each of these Reynolds numbers. The results also suggested that although mixing by diffusion was enhanced near the midplane of the reactor at Re_j = 53 and 93 due to very thin bands of acidic and basic fluid forming as the fluid spiraled towards the center of the reactor, near the top, and bottom walls of the reactor, the lower velocities due to fluid friction with the walls hindered the formation of these thin bands, and, thus, resulted in large regions of unmixed and unreacted fluid. At Re_j = 240, the flow was turbulent and unsteady. The mixing and reaction processes were still found to be incomplete even at this highest Reynolds number. At the reactor midplane, the flow images at Re_j = 240 showed unmixed base fluid near the center of the reactor, suggesting that just as in the Re_j = 53 and 93 cases, lower velocities near the top and bottom walls of the reactor hinder the mixing and rection of the acidic and basic streams. Ensemble averages of line-scan profiles for the Re_j = 240 were then calculated to provide statistical quantification of the microscale mixing in the reactor. These results further demonstrate that even at this highest Reynolds number investigated, mixing and reaction are incomplete. Visualization and quantification of micromixing using this reactive μ-LIF technique can prove useful in the validation of computational fluid dynamics models of micromixing within microscale chemical reactors.     

A “twisted” microfluidic mixer suitable for a wide range of flow rate applications

This paper proposes a new “twisted” 3D microfluidic mixer fabricated by a laser writing/microfabrication technique. Effective and efficient mixing using the twisted micromixers can be obtained by combining two general chaotic mixing mechanisms: splitting/recombining and chaotic advection. The lamination of mixer units provides the splitting and recombination mechanism when the quadrant of circles is arranged in a two-layered serial arrangement of mixing units. The overall 3D path of the microchannel introduces the advection. An experimental investigation using chemical solutions revealed that these novel 3D passive microfluidic mixers were stable and could be operated at a wide range of flow rates. This micromixer finds application in the manipulation of tiny volumes of liquids that are crucial in diagnostics. The mixing performance was evaluated by dye visualization, and using a pH test that determined the chemical reaction of the solutions. A comparison of the tornado-mixer with this twisted micromixer was made to evaluate the efficiency of mixing. The efficiency of mixing was calculated within the channel by acquiring intensities using ImageJ software. Results suggested that efficient mixing can be obtained when more than 3 units were consecutively placed. The geometry of the device, which has a length of 30 mm, enables the device to be integrated with micro total analysis systems and other lab-on-chip devices.     

Centrifugal multiplexing fixed-volume dispenser on a plastic lab-on-a-disk for parallel biochemical single-end-point assays

In this study, a multiple sample dispenser for precisely metered fixed volumes was successfully designed, fabricated, and fully characterized on a plastic centrifugal lab-on-a-disk (LOD) for parallel biochemical single-end-point assays. The dispenser, namely, a centrifugal multiplexing fixed-volume dispenser (C-MUFID) was designed with microfluidic structures based on the theoretical modeling about a centrifugal circumferential filling flow. The designed LODs were fabricated with a polystyrene substrate through micromachining and they were thermally bonded with a flat substrate. Furthermore, six parallel metering and dispensing assays were conducted at the same fixed-volume (1.27 μl) with a relative variation of ±0.02 μl. Moreover, the samples were metered and dispensed at different sub-volumes. To visualize the metering and dispensing performances, the C-MUFID was integrated with a serpentine micromixer during parallel centrifugal mixing tests. Parallel biochemical single-end-point assays were successfully conducted on the developed LOD using a standard serum with albumin, glucose, and total protein reagents. The developed LOD could be widely applied to various biochemical single-end-point assays which require different volume ratios of the sample and reagent by controlling the design of the C-MUFID. The proposed LOD is feasible for point-of-care diagnostics because of its mass-producible structures, reliable metering/dispensing performance, and parallel biochemical single-end-point assays, which can identify numerous biochemical.     

The effect of the surface functionalization and the electrolyte concentration on the electrical conductance of silica nanochannels

It is known that the conductance of nanochannels as a function of electrolyte concentration deviates from a linearly proportional relationship and approaches a value independent of the concentration as the electrolyte concentration is lowered. Most of the proposed models account for this behavior by considering a constant surface charge density and an ideal electrolyte solution. However, at low electrolyte concentrations, the ideal electrolyte approximation is no longer valid because the ions that result from the atmospheric carbon dioxide dissolution in water dominate the ionic concentration. In this paper, arrays of silica nanochannels were electrically characterized via conductance measurements. The conductance at low salt concentrations is modeled by a variable surface charge model that accounts for all ionic species in solution. This model was used to determine the variable surface charge of the bare silica nanochannels as well as of chemically modified nanochannels. The model correctly predicted the variation of the nanochannel conductance observed after silane (aminopropyldimethylethoxysilane) functionalization and single-strand DNA immobilization. Finally, pH modification of bulk KCl solutions was employed as an alternative method of changing the surface charge of silica nanochannels. Surface charge calculated from conductance measurements performed at different bulk pH values confirmed that the surface charge of the silica nanochannel walls is sensitive to the H⁺ concentration.     

Dynamic superconcentration at critical-point double-layer gates of conducting nanoporous granules due to asymmetric tangential fluxes

A transient 10⁶-fold concentration of double-layer counterions by a high-intensity electric field is demonstrated at the exit pole of a millimeter-sized conducting nanoporous granule that permits ion permeation. The phenomenon is attributed to a unique counterion screening dynamics that transforms half of the surface field into a converging one toward the ejecting pole. The resulting surface conduction flux then funnels a large upstream electro-osmotic convective counterion flux into the injecting hemisphere toward the zero-dimensional gate of the ejecting hemisphere to produce the superconcentration. As the concentrated counterion is ejected into the electroneutral bulk electrolyte, it attracts co-ions and produce a corresponding concentration of the co-ions. This mechanism is also shown to trap and concentrate co-ion microcolloids of micron sizes too (macroions) and hence has potential application in bead-based molecular assays.     

Biosensing in a microelectrofluidic system using optical whispering-gallery mode spectroscopy

Label-free detection of biomolecules using an optical whispering-gallery mode sensor in a microelectrofluidic channel is simulated. Negatively charged bovine serum albumin is considered as the model protein analyte. The analyte transport in aqueous solution is controlled by an externally applied electrical field. The finite element method is employed for solving the equations of the charged species transport, the Poisson equation of electric potential, the equations of conservation of momentum and energy, and the Helmholtz equations of electromagnetic waves. The adsorption process of the protein molecules on the microsensor head surface is monitored by the resonance frequency shifts. Frequency shift caused by temperature variation due to Joule heating is analyzed and found to be negligible. The induced shifts behave in a manner similar to Langmuir-like adsorption kinetics; but the time constant increases due to the presence of the external electrical field. A correlation of the frequency shift, the analyte feed concentration in the solution, and the applied voltage gradient is obtained, in which an excellent linear relationship between the frequency shift and the analyte concentration is revealed. The applied voltage gradient enhances significantly the analyte concentration in the vicinity of the sensor surface; thus, the sensor sensitivity which has a power function of the voltage gradient with exponent 2.85 in the controlled voltage range. Simulated detection of extremely low protein concentration to the pico-molar level is carried out.     

Mathematical and numerical model to study two-dimensional free flow isoelectric focusing

Even though isoelectric focusing (IEF) is a very useful technique for sample concentration and separation, it is challenging to extract separated samples for further processing. Moreover, the continuous sample concentration and separation are not possible in the conventional IEF. To overcome these challenges, free flow IEF (FFIEF) is introduced in which a flow field is applied in the direction perpendicular to the applied electric field. In this study, a mathematical model is developed for FFIEF to understand the roles of flow and electric fields for efficient design of microfluidic chip for continuous separation of proteins from an initial well mixed solution. A finite volume based numerical scheme is implemented to simulate two dimensional FFIEF in a microfluidic chip. Simulation results indicate that a pH gradient forms as samples flow downstream and this pH profile agrees well with experimental results validating our model. In addition, our simulation results predict the experimental behavior of pI markers in a FFIEF microchip. This numerical model is used to predict the separation behavior of two proteins (serum albumin and cardiac troponin I) in a two-dimensional straight microchip. The effect of electric field is investigated for continuous separation of proteins. Moreover, a new channel design is presented to increase the separation resolution by introducing cross-stream flow velocity. Numerical results indicate that the separation resolution can be improved by three folds in this new design compare to the conventional straight channel design.     

Three-dimensional cellular focusing utilizing a combination of insulator-based and metallic dielectrophoresis

Particle focusing in microfluidic devices is a necessary step in medical applications, such as detection, sorting, counting, and flow cytometry. This study proposes a microdevice that combines insulator-based and metal-electrode dielectrophoresis for the three-dimensional focusing of biological cells. Four insulating structures, which form an X pattern, are employed to confine the electric field in a conducting solution, thereby creating localized field minima in the microchannel. These electrodes, 56-μm-wide at the top and bottom surfaces, are connected to one electric pole of the power source. The electrodes connected to the opposite pole, which are at the sides of the microchannel, have one of three patterns: planar, dual-planar, or three-dimensional. Therefore, low-electric-field regions at the center of the microchannel are generated to restrain the viable HeLa cells with negative dielectrophoretic response. The array of insulating structures aforementioned is used to enhance the performance of confinement. According to numerical simulations, three-dimensional electrodes exhibit the best focusing performance, followed by dual-planar and planar electrodes. Experimental results reveal that increasing the strength of the applied electric field or decreasing the inlet flow rate significantly enhances focusing performance. The smallest width of focusing is 17 μm for an applied voltage and an inlet flow rate of 35 V and 0.5 μl/min, respectively. The effect of the inlet flow rate on focusing is insignificant for an applied voltage of 35 V. The proposed design retains the advantages of insulator-based dielectrophoresis with a relatively low required voltage. Additionally, complicated flow controls are unnecessary for the three-dimensional focusing of cells.     

A nanoporous optofluidic microsystem for highly sensitive and repeatable surface enhanced Raman spectroscopy detection

We report the demonstration of an optofluidic surface enhanced Raman spectroscopy (SERS) device that leverages a nanoporous microfluidic matrix to improve the SERS detection performance by more than two orders of magnitude as compared to a typical open microfluidic channel. Although it is a growing trend to integrate optical biosensors into microfluidic channels, this basic combination has been detrimental to the sensing performance when applied to SERS. Recently, however, synergistic combinations between microfluidic functions and photonics (i.e., optofluidics) have been implemented that improve the detection performance of SERS. Conceptually, the simplest optofluidic SERS techniques reported to date utilize a single nanofluidic channel to trap nanoparticle-analyte conjugates as a method of preconcentration before detection. In this work, we leverage this paradigm while improving upon the simplicity by forming a 3D nanofluidic network with packed nanoporous silica microspheres in a microfluidic channel; this creates a concentration matrix that traps silver nanoclusters and adsorbed analytes into the SERS detection volume. With this approach, we are able to achieve a detection limit of 400 attomoles of Rhodamine 6G after only 2 min of sample loading with high chip-to-chip repeatability. Due to the high number of fluidic paths in the nanoporous channel, this approach is less prone to clogging than single nanofluidic inlets, and the loading time is decreased compared to previous reports. In addition, fabrication of this microsystem is quite simple, as nanoscale fabrication is not necessary. Finally, integrated multimode fiber optic cables eliminate the need for optical alignment, and thus the device is relevant for portable and automated applications in the field, including point-of-sample and point-of-care detection. To illustrate a relevant field-based application, we demonstrate the detection of 12 ppb of the organophosphate malathion in water using the nanofluidic SERS microsystem.     

Numerical modeling of motion trajectory and deformation behavior of a cell in a nonuniform electric field

The motion trajectory and deformation behavior of a neutral red blood cell (RBC) in a microchannel subjected to an externally applied nonuniform electric field are numerically investigated, where both the membrane mechanical force and the dielectrophoresis (DEP) force are considered. The simulation results demonstrate that the DEP force is significantly influenced by several factors, namely, the RBC size, electrode potential, electric frequency, RBC permittivity, and conductivity, which finally results in the different behaviors of the cell motion and deformation in the nonuniform electric field.     

Distinctive translational and self-rotational motion of lymphoma cells in an optically induced non-rotational alternating current electric field

In this paper, the translational motion and self-rotational behaviors of the Raji cells, a type of B-cell lymphoma cell, in an optically induced, non-rotational, electric field have been characterized by utilizing a digitally programmable and optically activated microfluidics chip with the assistance of an externally applied AC bias potential. The crossover frequency spectrum of the Raji cells was studied by observing the different linear translation responses of these cells to the positive and negative optically induced dielectrophoresis force generated by a projected light pattern. This digitally projected spot served as the virtual electrode to generate an axisymmetric and non-uniform electric field. Then, the membrane capacitance of the Raji cells could be directly measured. Furthermore, Raji cells under this condition also exhibited a self-rotation behavior. The repeatable and controlled self-rotation speeds of the Raji cells to the externally applied frequency and voltage were systematically investigated and characterized via computer-vision algorithms. The self-rotational speed of the Raji cells reached a maximum value at 60 kHz and demonstrated a quadratic relationship with respect to the applied voltage. Furthermore, optically projected patterns of four orthogonal electrodes were also employed as the virtual electrodes to manipulate the Raji cells. These results demonstrated that Raji cells located at the center of the four electrode pattern could not be self-rotated. Instead any Raji cells that deviated from this center area would also self-rotate. Most importantly, the Raji cells did not exhibit the self-rotational behavior after translating and rotating with respect to the center of any two adjacent electrodes. The spatial distributions of the electric field generated by the optically projected spot and the pattern of four electrodes were also modeled using a finite element numerical simulation. These simulations validated that the electric field distributions were non-uniform and non-rotational. Hence, the non-uniform electric field must play a key role in the self-rotation of the Raji cells. As a whole, this study elucidates an optoelectric-coupled microfluidics-based mechanism for cellular translation and self-rotation that can be used to extract the dielectric properties of the cells without using conventional metal-based microelectrodes. This technique may provide a simpler method for label-free identification of cancerous cells with many associated clinical applications.     

Electrokinetic focusing and filtration of cells in a serpentine microchannel

Focusing cells into a single stream is usually a necessary step prior to counting and separating them in microfluidic devices such as flow cytometers and cell sorters. This work presents a sheathless electrokinetic focusing of yeast cells in a planar serpentine microchannel using dc-biased ac electric fields. The concurrent pumping and focusing of yeast cells arise from the dc electrokinetic transport and the turn-induced ac∕dc dielectrophoretic motion, respectively. The effects of electric field (including ac to dc field ratio and ac field frequency) and concentration (including buffer concentration and cell concentration) on the cell focusing performance were studied experimentally and numerically. A continuous electrokinetic filtration of E. coli cells from yeast cells was also demonstrated via their differential electrokinetic focusing in a serpentine microchannel.     

Novel index for micromixing characterization and comparative analysis

The most basic micromixer is a T- or Y-mixer, where two confluent streams mix due to transverse diffusion. To enhance micromixing, various modifications of T-mixers are reported such as heterogeneously charged walls, grooves on the channel base, geometric variations by introducing physical constrictions, etc. The performance of these reported designs is evaluated against the T-mixer in terms of the deviation from perfectly mixed state and mixing length (device length required to achieve perfect mixing). Although many studies have noticed the reduced flow rates for improved mixer designs, the residence time is not taken into consideration for micromixing performance evaluation. In this work, we propose a novel index, based on residence time, for micromixing characterization and comparative analysis. For any given mixer, the proposed index identifies the nondiffusive mixing enhancement with respect to the T-mixer. Various micromixers are evaluated using the proposed index to demonstrate the usefulness of the index. It is also shown that physical constriction mixer types are equivalent to T-mixers. The proposed index is found to be insightful and could be used as a benchmark for comparing different mixing strategies.