Parasitic trap cancellation using multiple frequency dielectrophoresis, demonstrated by loading cells into cages

This paper presents a method of using multiple frequencies to counteract electric field distortions that interfere with the dielectrophoretic (DEP) manipulation of particles or cells. To demonstrate the technique, simulations were performed for a scenario in which cells were to be loaded into a cage whose walls created parasitic trapping sites that prevented cells from entering it. By employing negative DEP on one electrode in conjunction with positive DEP on another, these traps could be almost completely cancelled. The model predictions were validated experimentally: multiple frequency DEP was used to load many cells into a cage in a matter of seconds in fluid flows of up to 300 microm s(-1), which could not be done with single frequency DEP. Actively cancelling field distortions permits the presence of features that would otherwise be prohibited near regions of dielectrophoretic manipulation, significantly expanding the environments in which dielectrophoresis can be used.     

Dielectrophoresis of Caenorhabditis elegans

We demonstrate for the first time the dielectrophoretic trapping and manipulation of a whole animal, the nematode Caenorhabditis elegans. We studied the effect of the electric field on the nematode as a function of field intensity and frequency. We identified a range of electric field intensities and frequencies that trap worms without apparent adverse effect on their viability. Worms tethered by dielectrophoresis (DEP) exhibit behavioral responses to blue light, indicating that at least some of the nervous system functions are unimpaired by the electrical field. DEP is useful to dynamically tether nematodes, sort nematodes according to size, and separate dead worms from live ones.     

Frequency discretization in dielectrophoretic assisted cell sorting arrays to isolate neural cells

We present an automated dielectrophoretic assisted cell sorting (DACS) device for dielectric characterization and isolation of neural cells. Dielectrophoretic (DEP) principles are often used to develop cell sorting techniques. Here we report the first statistically significant neuronal sorting using DACS to enrich neurons from a heterogeneous population of mouse derived neural stem/progenitor cells (NSPCs) and neurons. We also study the dielectric dispersions within a heterogeneous cell population using a Monte-Carlo (MC) simulation. This simulation model explains the trapping behavior of populations as a function of frequency and predicts sorting efficiencies. The platform consists of a DEP electrode array with three multiplexed trapping regions that can be independently activated at different frequencies. A novel microfluidic manifold enables cell sorting by trapping and collecting cells at discrete frequency bands rather than single frequencies. The device is used to first determine the percentage of cells trapped at these frequency bands. With this characterization and the MC simulation we choose the optimal parameters for neuronal sorting. Cell sorting experiments presented achieve a 1.4-fold neuronal enrichment as predicted by our model.     

Selective concentration of human cancer cells using contactless dielectrophoresis

This work is the first to demonstrate the ability of contactless dielectrophoresis (cDEP) to isolate target cell species from a heterogeneous sample of live cells. Since all cell types have a unique molecular composition, it is expected that their dielectrophoretic (DEP) properties are also unique. cDEP is a technique developed to improve upon traditional and insulator-based DEP devices by replacing embedded metal electrodes with fluid electrode channels positioned alongside desired trapping locations. Through the placement of the fluid electrode channels and the removal of contact between the electrodes and the sample fluid, cDEP mitigates issues associated with sample/electrode contact. MCF10A, MCF7, and MDA-MB-231 human breast cells were used to represent early, intermediate, and late-staged breast cancer, respectively. Trapping frequency responses of each cell type were distinct, with the largest difference between the cells found at 20 and 30 V. MDA-MB-231 cells were successfully isolated from a population containing MCF10A and MCF7 cells at 30 V and 164 kHz. The ability to selectively concentrate cells is the key to development of biological applications using DEP. The isolation of these cells could provide a workbench for clinicians to detect transformed cells at their earliest stage, screen drug therapies prior to patient treatment, increasing the probability of success, and eliminate unsuccessful treatment options.     

Frequency dependence of gold nanoparticle superassembly by dielectrophoresis

Dielectrophoresis is an effective method for capturing nanoparticles and assembling them into nanostructures. The frequency of the dielectrophoretic alternating current (ac) electric field greatly influences the morphology of resultant nanoparticle assemblies. In this study, frequency regimes associated with specific gold nanoparticle assembly morphologies were identified. Gold nanoparticles suspended in water were captured by microelectrodes at different electric field frequencies onto thin silicon nitride membranes. The resultant assemblies were examined by transmission electron microscopy. For this system, the major frequency-dependent influence on morphology appears to arise not from the Clausius-Mossotti factor of the dielectrophoretic force itself, but instead from ac electroosmotic fluid flow and the influence of the electrical double layer at the electrode-solution interface. Frequency regimes of technological interest include those forming one-dimensional nanoparticle chains, microwires, combinations of microwires and nanoparticle chains suitable for nanogap electrode formation, and dense three-dimensional assemblies with very high surface area.     

Microfluidic single-cell cultivation chip with controllable immobilization and selective release of yeast cells

We present a microfluidic cell-culture chip that enables trapping, cultivation and release of selected individual cells. The chip is fabricated by a simple hybrid glass-SU-8-PDMS approach, which produces a completely transparent microfluidic system amenable to optical inspection. Single cells are trapped in a microfluidic channel using mild suction at defined cell immobilization orifices, where they are cultivated under controlled environmental conditions. Cells of interest can be individually and independently released for further downstream analysis by applying a negative dielectrophoretic force via the respective electrodes located at each immobilization site. The combination of hydrodynamic cell-trapping and dielectrophoretic methods for cell releasing enables highly versatile single-cell manipulation in an array-based format. Computational fluid dynamics simulations were performed to estimate the properties of the system during cell trapping and releasing. Polystyrene beads and yeast cells have been used to investigate and characterize the different functions and to demonstrate biological compatibility and viability of the platform for single-cell applications in research areas such as systems biology.     

Quantitative modeling of dielectrophoretic traps

We present quantitative modeling software for simulating multiple forces acting on a single particle in a microsystem. In this paper, we focus on dielectrophoretic (DEP) trapping of single cells against fluid flow. The software effectively models the trapping behavior for a range of particles including beads, mammalian cells, viruses, and bacteria. In addition, the software can be used to reveal useful information about the DEP traps - such as multipolar DEP force effects, trap size-selectivity, and effects from varying the flow chamber height. Our modeling software thus serves as a predictive tool, enabling the design of novel DEP traps with superior performance over existing trap geometries. In addition, the software can evaluate a range of trap dimensions to determine the effects on trapping behavior, thus optimizing the trap geometry before it is even fabricated. The software is freely available to the scientific community at: .     

Aligning fast alternating current electroosmotic flow fields and characteristic frequencies with dielectrophoretic traps to achieve rapid bacteria detection

Tailor-designed alternating current electroosmotic (AC-EO) stagnation flows are used to convect bioparticles globally from a bulk solution to localized dielectrophoretic (DEP) traps that are aligned at the flow stagnation points. The multiscale trap, with a typical trapping time of seconds for a dilute 70 microL volume of 10(3) particles per cc sample, is several orders of magnitude faster than conventional DEP traps and earlier AC-EO traps with parallel, castellated, or finger electrodes. A novel serpentine wire capable of sustaining a high voltage, up to 2500 V(RMS), without causing excessive heat dissipation or Faradaic reaction in strong electrolytes is fabricated to produce the strong AC-EO flow with two separated stagnation lines, one aligned with the field minimum and one with the field maximum. The continuous wire design allows a large applied voltage without inducing Faradaic electrode reactions. Particles are trapped within seconds at one of the traps depending on whether they suffer negative or positive DEP. The particles can also be rapidly released from their respective traps by varying the frequency of the applied AC field below particle-distinct cross-over frequencies. Zwitterion addition to the buffer allows further geometric and frequency alignments of the AC-EO and DEP motions. The same device hence allows fast trapping, detection, sorting, and characterization on a sample with realistic conductivity, volume, and bacteria count.     

Bacterial chromosome extraction and isolation

We have used diffusive mixing and dielectrophoretic trapping to lyse Escherichia coli cells in a microfabricated environment and trap the E. coli chromosome. We characterize the conditions needed for efficient lysis of the cells, and conditions needed for the dielectrophoretic trapping of the chromatin without the trapping of cytoplasmic proteins.     

Cross-scale electric manipulations of cells and droplets by frequency-modulated dielectrophoresis and electrowetting

Two important electric forces, dielectrophoresis (DEP) and electrowetting-on-dielectric (EWOD), are demonstrated by dielectric-coated electrodes on a single chip to manipulate objects on different scales, which results in a dielectrophoretic concentrator in an EWOD-actuated droplet. By applying appropriate electric signals with different frequencies on identical electrodes, EWOD and DEP can be selectively generated on the proposed chip. At low frequencies, the applied voltage is consumed mostly in the dielectric layer and causes EWOD to pump liquid droplets on the millimetre scale. However, high frequency signals establish electric fields in the liquid and generate DEP forces to actuate cells or particles on the micrometre scale inside the droplet. For better performance of EWOD and DEP, square and strip electrodes are designed, respectively. Mammalian cells (Neuro-2a) and polystyrene beads are successfully actuated by a 2 MHz signal in a droplet by positive DEP and negative DEP, respectively. Droplet splitting is achieved by EWOD with a 1 kHz signal after moving cells or beads to one side of the droplet. Cell concentration, measured by a cell count chamber before and after experiments, increases 1.6 times from 8.6 x 10(5) cells ml(-1) to 1.4 x 10(6) cells ml(-1) with a single cycle of positive DEP attraction. By comparing the cutoff frequency of the voltage drop in the dielectric layer and the cross-over frequency of Re(fCM) of the suspended particles, we can estimate the frequency-modulated behaviors between EWOD, positive DEP, and negative DEP. A proposed weighted Re(fCM) facilitates analysis of the DEP phenomenon on dielectric-coated electrodes.     

Recent advances in dielectrophoresis-based cell viability assessment

Dielectrophoresis (DEP) is a non-destructive, accurate, and label-free cell manipulating technique and DEP applications have been found in various fields. Assessment of cell viability is one of the important applications and many investigations have been reported. In this paper, cell polarization and its modeling, some key parameters employed for living/dead cell separation, as well as electrode configurations are reviewed. Focus is given to the latest development of DEP devices employed for the assessment of cell viability. Experimentally determined factors for separating living/dead cells, such as the conductivity of suspending medium and the frequency of applied electric field, are summarized. The future directions and potential challenges in this field are also outlined.     

Dynamically controlled dielectrophoresis using resonant tuning

Electrically polarizable micro- and nanoparticles and droplets can be trapped using the gradient electric field of electrodes. But the spatial profile of the resultant dielectrophoretic force is fixed once the electrode structure is defined. To change the force profile, entire complex lab-on-a-chip systems must be re-fabricated with modified electrode structures. To overcome this problem, we propose an approach for the dynamic control of the spatial profile of the dielectrophoretic force by interfacing the trap electrodes with a resistor and an inductor to form a resonant resistor–inductor–capacitor (RLC) circuit. Using a dielectrophoretically trapped water droplet suspended in silicone oil, we show that the resonator amplitude, detuning, and linewidth can be continuously varied by changing the supply voltage, supply frequency, and the circuit resistance to obtain the desired trap depth, range, and stiffness. We show that by proper tuning of the resonator, the trap range can be extended without increasing the supply voltage, thus preventing sensitive samples from exposure to high electric fields at the stable trapping position. Such unprecedented dynamic control of dielectrophoretic forces opens avenues for the tunable active manipulation of sensitive biological and biochemical specimen in droplet microfluidic devices used for single-cell and biochemical reaction analysis.     

Dielectrophoretic trapping of DNA-coated gold nanoparticles on silicon based vertical nanogap devices

We report on the successful dielectrophoretic trapping and electrical characterization of DNA-coated gold nanoparticles on vertical nanogap devices (VNDs). The nanogap devices with an electrode distance of 13 nm were fabricated from Silicon-on-Insulator (SOI) material using a combination of anisotropic reactive ion etching (RIE), selective wet chemical etching and metal thin-film deposition. Au nanoparticles (diameter 40 nm) coated with a monolayer of dithiolated 8 base pairs double stranded DNA were dielectrophoretically trapped into the nanogap from electrolyte buffer solution at MHz frequencies as verified by scanning and transmission electron microscopy (SEM/TEM) analysis. First electrical transport measurements through the formed DNA-Au-DNA junctions partially revealed an approximately linear current-voltage characteristic with resistance in the range of 2-4 G? when measured in solution. Our findings point to the importance of strong covalent bonding to the electrodes in order to observe DNA conductance, both in solution and in the dry state. We propose our setup for novel applications in biosensing, addressing the direct interaction of biomolecular species with DNA in aqueous electrolyte media.     

Multi-step dielectrophoresis for separation of particles

A new concept for separation of particles based on repetitive dielectrophoretic trapping and release in a flow system is proposed. Calculations using the finite element method have been performed to envision the particle behavior and the separation effectiveness of the proposed method. As a model system, polystyrene beads in deionized water and a micro-flow channel with arrays of interdigited electrodes have been used. Results show that the resolution increases as a direct function of the number of trap-and-release steps, and that a difference in size will have a larger influence on the separation than a difference in other dielectrophoretic properties. About 200 trap-and-release steps would be required to separate particles with a size difference of 0.2%. The enhanced separation power of dielectrophoresis with multiple steps could be of great importance, not only for fractionation of particles with small differences in size, but also for measuring changes in surface conductivity, or for separations based on combinations of difference in size and dielectric properties.     

Polarization behavior of polystyrene particles under direct current and low‐frequency (<1 kHz) electric fields in dielectrophoretic systems

The relative polarization behavior of micron and submicron polystyrene particles was investigated under direct current and very low frequency (<1 kHz) alternating current electric fields. Relative polarization of particles with respect to the suspending medium is expressed in terms of the Clausius–Mossotti factor, a parameter of crucial importance in dielectrophoretic‐based operations. Particle relative polarization was studied by employing insulator‐based dielectrophoretic (iDEP) devices. The effects of particle size, medium conductivity, and frequency (10–1000 Hz) of the applied electric potential on particle response were assessed through experiments and mathematical modeling with COMSOL Multiphysics^®. Particles of different sizes (100–1000 nm diameters) were introduced into iDEP devices fabricated from polydimethylsiloxane (PDMS) and their dielectrophoretic responses under direct and alternating current electric fields were recorded and analyzed in the form of images and videos. The results illustrated that particle polarizability and dielectrophoretic response depend greatly on particle size and the frequency of the electric field. Small particles tend to exhibit positive DEP at higher frequencies (200–1000 Hz), while large particles exhibit negative DEP at lower frequencies (20–200 Hz). These differences in relative polarization can be used for the design of iDEP‐based separations and analysis of particle mixtures.     

Electronic sorting and recovery of single live cells from microlitre sized samples

Sorting and recovering specific live cells from samples containing less than a few thousand cells have become major hurdles in rare cell exploration such as stem cell research, cell therapy and cell based diagnostics. We describe here a new technology based on a microelectronic chip integrating an array of over 100,000 independent electrodes and sensors which allow individual and parallel single cell manipulation of up to 10,000 cells while maintaining viability and proliferation capabilities. Manipulation is carried out using dynamic dielectrophoretic traps controlled by an electronic interface. We also demonstrate the capabilities of the chip by sorting and recovering individual live fluorescent cells from an unlabeled population.     

Dielectrophoretic characterization and separation of monocytes and macrophages using 3D carbon‐electrodes

Monocyte heterogeneity and its prevalence are revealed as indicator of several human diseases ranking from cardiovascular diseases to rheumatoid arthritis, chronic kidney diseases, autoimmune multiple sclerosis, and stroke injuries. When monocytes and macrophages are characterized and isolated with preserved genetic, phenotypic and functional properties, they can be used as label‐free biomarkers for precise diagnostics and treatment of various diseases. Here, the dielectrophoretic responses of the monocytes and macrophages were examined. We present 3D carbon‐electrode dielectrophoresis (carbon‐DEP) as a separation tool for U937 monocytes and U937 monocyte‐differentiated macrophages. The carbon‐electrodes advanced the usability and throughput of DEP separation, presented wider electrochemical stability. Using the 3D carbon‐DEP chip, we first identified the selective positive and negative DEP responses and specific crossover frequencies of monocytes and macrophages as their signatures for separation. The crossover frequency of monocytes and macrophages was 17 and 30 kHz, respectively. Next, we separated monocyte and macrophage subpopulations using their specific dielectrophoretic responses. Afterward, we used a fluorescence‐activated cell sorter to confirm our results. Finally, we enriched 70% of monocyte cells from the mixed cell population, in other words, concentration of monocyte cells to macrophage cells was five times increased, using the 30‐kHz, 10‐Vpp electric field and 1 μL/min flow rate.     

Lateral separation of colloids or cells by dielectrophoresis augmented by AC electroosmosis

Colloidal particles and biological cells are patterned and separated laterally adjacent to a micropatterned electrode array by applying AC electric fields that are principally oriented normally to the electrode array. This is demonstrated for yeast cells, red blood cells, and colloidal polystyrene particles of different sizes and zeta-potentials. The separation mechanism is observed experimentally to depend on the applied field frequency and voltage. At high frequencies, particles position themselves in a manner that is consistent with dielectrophoresis, while at low frequencies, the positioning is explained in terms of a strong coupling between gravity, the vertical component of the dielectrophoretic force, and the Stokes drag on particles induced by AC electroosmotic flow. Compared to high frequency dielectrophoretic separations, the low frequency separations are faster and require lower applied voltages. Furthermore, the AC electroosmosis coupling with dielectrophoresis may enable cell separations that are not feasible based on dielectrophoresis alone.     

DC insulator dielectrophoretic applications in microdevice technology: a review

Dielectrophoresis is a noninvasive, nondestructive, inexpensive, and fast technique for the manipulation of bioparticles. Recent advances in the field of dielectrophoresis (DEP) have resulted in new approaches for characterizing the behavior of particles and cells using direct current (DC) electric fields. In such approaches, spatial nonuniformities are created in the channel by embedding insulating obstacles in the channel or flow field in order to perform separation or trapping. This emerging field of dielectrophoresis is commonly termed DC insulator dielectrophoresis (DC-iDEP), insulator-based dielectrophoresis (iDEP), or electrodeless dielectrophoresis (eDEP). In many microdevices, this form of dielectrophoresis has advantages over traditional AC-DEP, including single material microfabrication, remotely positioned electrodes, and reduced fouling of the test region. DC-iDEP applications have included disease detection, separation of cancerous cells from normal cells, and separation of live from dead bacteria. However, there is a need for a critical report to integrate these important research findings. The aim of this review is to provide an overview of the current state-of-art technology in the field of DC-iDEP for the separation and trapping of inert particles and cells. In this article, a review of the concepts and theory leading to the manipulation of particles via DC-iDEP is given, and insulating obstacle geometry designs and the characterization of device performance are discussed. This review compiles and compares the significant findings obtained by researchers in handling and manipulating particles.     

Numerical comparison between Maxwell stress method and equivalent multipole approach for calculation of the dielectrophoretic force in single-cell traps

This paper presents detailed numerical calculations of the dielectrophoretic force in traps designed for single-cell trapping. A trap with eight planar electrodes is studied for spherical and ellipsoidal particles using the boundary element method (BEM). Multipolar approximations of orders one to three are compared with the full Maxwell stress tensor (MST) calculation of the electrical force on spherical particles. Ellipsoidal particles are also studied, but in their case only the dipolar approximation is available for comparison with the MST solution. The results show that a small number of multipolar terms need to be considered in order to obtain accurate results for spheres, even in the proximity of the electrodes, and that the full MST calculation is only required in the study of non-spherical particles.