Dielectrophoresis Experiments Research Articles

A dielectrophoresis proof of concept of polystyrene particles and in‐vitro human epidermal keratinocytes migration for wound rejuvenation

AbstractDiabetes affects approximately 170 million people worldwide, is expected to double by 2030, and is a severe problem. Electrical stimulation (ES) via dielectrophoresis (DEP) technique may be an effective alternative in enhancing healing rates in diabetic patients with open ulcers. This research used DEP force (FDEP) to manipulate 3.2, 4.8, 10, and 15 μm polystyrene (PS) particles to predict the migration capability of human epidermal keratinocytes (HEKs). A numerical modeling method, MyDEP, was used to predict the interpretation of Clausius–Mossotti factors of PS particles and HEKs. The finite element method computes the electric field intensity and particle trajectory based on DEP and drag forces in their respective medium. DEP experiments on numerous size PS particles and alive HEKs were carried out in a tapered aluminium microelectrode array using a non‐uniform electric field. The distinct PS particles exhibit positive DEP (PDEP), crossover frequency (fXO), and negative DEP (NDEP), whereas the HEKs experience, only NDEP due to its high conductive medium in frequency ranges from 100 kHz to 1 MHz. Finally, the DIPP‐MotionV analysis shows that particle mobility between speed and acceleration is statistically considerable. When an appropriate frequency is applied to HEKs in random locations, the FDEP aligns at the desired target position based on its dielectric properties, which accelerates wound healing in in‐vivo conditions.

Dielectrophoresis Prototypic Polystyrene Particle Synchronization toward Alive Keratinocyte Cells for Rapid Chronic Wound Healing.

Diabetes patients are at risk of having chronic wounds, which would take months to years to resolve naturally. Chronic wounds can be countered using the electrical stimulation technique (EST) by dielectrophoresis (DEP), which is label-free, highly sensitive, and selective for particle trajectory. In this study, we focus on the validation of polystyrene particles of 3.2 and 4.8 μm to predict the behavior of keratinocytes to estimate their crossover frequency (fXO) using the DEP force (FDEP) for particle manipulation. MyDEP is a piece of java-based stand-alone software used to consider the dielectric particle response to AC electric fields and analyzes the electrical properties of biological cells. The prototypic 3.2 and 4.8 μm polystyrene particles have fXO values from MyDEP of 425.02 and 275.37 kHz, respectively. Fibroblast cells were also subjected to numerical analysis because the interaction of keratinocytes and fibroblast cells is essential for wound healing. Consequently, the predicted fXO from the MyDEP plot for keratinocyte and fibroblast cells are 510.53 and 28.10 MHz, respectively. The finite element method (FEM) is utilized to compute the electric field intensity and particle trajectory based on DEP and drag forces. Moreover, the particle trajectories are quantified in a high and low conductive medium. To justify the simulation, further DEP experiments are carried out by applying a non-uniform electric field to a mixture of different sizes of polystyrene particles and keratinocyte cells, and these results are well agreed. The alive keratinocyte cells exhibit NDEP force in a highly conductive medium from 100 kHz to 25 MHz. 2D/3D motion analysis software (DIPP-MotionV) can also perform image analysis of keratinocyte cells and evaluate the average speed, acceleration, and trajectory position. The resultant NDEP force can align the keratinocyte cells in the wound site upon suitable applied frequency. Thus, MyDEP estimates the Clausius–Mossotti factors (CMF), FEM computes the cell trajectory, and the experimental results of prototypic polystyrene particles are well correlated and provide an optimistic response towards keratinocyte cells for rapid wound healing applications.

Protein Dielectrophoresis: I. Status of Experiments and an Empirical Theory.

The dielectrophoresis (DEP) data reported in the literature since 1994 for 22 different globular proteins is examined in detail. Apart from three cases, all of the reported protein DEP experiments employed a gradient field factor that is much smaller (in some instances by many orders of magnitude) than the ~4 × 1021 V2/m3 required, according to current DEP theory, to overcome the dispersive forces associated with Brownian motion. This failing results from the macroscopic Clausius–Mossotti (CM) factor being restricted to the range 1.0 > CM > −0.5. Current DEP theory precludes the protein’s permanent dipole moment (rather than the induced moment) from contributing to the DEP force. Based on the magnitude of the β-dispersion exhibited by globular proteins in the frequency range 1 kHz–50 MHz, an empirically derived molecular version of CM is obtained. This factor varies greatly in magnitude from protein to protein (e.g., ~37,000 for carboxypeptidase; ~190 for phospholipase) and when incorporated into the basic expression for the DEP force brings most of the reported protein DEP above the minimum required to overcome dispersive Brownian thermal effects. We believe this empirically-derived finding validates the theories currently being advanced by Matyushov and co-workers.

A mathematical model of dielectrophoretic data to connect measurements with cell properties.

Dielectrophoresis (DEP) brings about the high-resolution separations of cells and other bioparticles arising from very subtle differences in their properties. However, an unanticipated limitation has arisen: difficulty in assignment of specific biological features which vary between two cell populations. This hampers the ability to interpret the significance of the variations. To realize the opportunities made possible by dielectrophoresis, the data and the diversity of structures found in cells and bioparticles must be linked. While the crossover frequency in DEP has been studied in-depth and exploited in applications using AC fields, less attention has been given when a DC field is present. Here, a new mathematical model of dielectrophoretic data is introduced which connects the physical properties of cells to specific elements of the data from potential- or time-varied DEP experiments. The slope of the data in either analysis is related to the electrokinetic mobility, while the potential at which capture initiates in potential-based analysis is related to both the electrokinetic and dielectrophoretic mobilities. These mobilities can be assigned to cellular properties for which values appear in the literature. Representative examples of high and low values of properties such as conductivity, zeta potential, and surface charge density for bacteria including Streptococcus mutans, Rhodococcus erythropolis, Pasteurella multocida, Escherichia coli, and Staphylococcus aureus are considered. While the many properties of a cell collapse into one or two features of data, for a well-vetted system the model can indicate the extent of dissimilarity. The influence of individual properties on the features of dielectrophoretic data is summarized, allowing for further interpretation of data. Graphical abstract.

MyDEP: A New Computational Tool for Dielectric Modeling of Particles and Cells

Dielectrophoresis (DEP) and electrorotation (ROT) are two electrokinetic phenomena exploiting nonuniform electric fields to exert a force or torque on biological particles suspended in liquid media. They are widely used in lab-on-chip devices for the manipulation, trapping, separation, and characterization of cells, microorganisms, and other particles. The DEP force and ROT torque depend on the respective polarizabilities of the particle and medium, which in turn depend on their dielectric properties and on the field frequency. In this work, we present a new software, MyDEP, which implements several particle models based on concentric shells with adjustable dielectric properties. This tool enables the study of the variation in DEP and ROT spectra according to different parameters, such as the field frequency and medium conductivity. Such predictions of particle behavior are very useful for choosing appropriate parameters in DEP experiments. The software also enables the study of the homogenized properties of spherical or ellipsoidal multishell particles and provides a database containing published cell properties. Equivalent electrical conductivity and relative permittivity of the cell alone and in suspension can be calculated. The software also offers the ability to create graphs of the evolution of the crossover frequencies with the electric field frequency. These graphs can be directly exported from the software.

Dielectrophoretic sensitivity analysis of cell characterization

Dielectrophoresis (DEP) has been intensively studied to differentiate cells treated with heat or drugs. To estimate cell properties from DEP experiments, the multi-dimensional relationship between the electrical polarization and the electrical properties of cell components should be understood. However, the limited information extracted from DEP experiments has imposed the burden of approximation of multiple cell properties. In this paper, we present a novel sensitivity analysis to predict cell properties from DEP experimental data. We generated virtual sample cells of various properties in given ranges. Then, the experimental conditions to achieve a higher sensitivity of a polarization factor to cell properties were found through a regressive analysis. Based on the sampling results, cell properties were estimated from the experimental data using a conventional frequency method and a new transition condition method. The estimation predicts the possible properties of cells, which benefits rigorous analysis of cell properties.

A Novel Approach to the Sustainable Synthesis of Carbon Fibers

Presented here are preliminary experiments facilitating the use of the cellulose-extruding bacteria Gluconoacetobacter xylinus as a sustainable tool for the production of carbon fibers. G. xylinus has the natural ability to convert different kinds of sugars from its environment into highly crystalline cellulose nanofibrils. When grown in culture, the random movement of the bacteria leads to a porous cellulosic scaffold with no apparent order. Compared to the cellulose extracted from plants, bacterial cellulose has higher purity and exhibits superior mechanical properties [1]. Results presented last year in this same conference showed how heat treatment of these cellulose fibers that were functionalized with metal precursors yielded a porous tungsten carbide. Here, we focus on the positioning of G. xylinus cells using Dielectrophoresis (DEP) to grow cellulose fibers from specific locations, which we then carbonize by pyrolysis. DEP refers to the movement of electrically polarized cells in response to a non-uniform electric field of specific frequency and magnitude. Specifically, positive DEP will be used here to position the G. xylinus cells on the electrodes while they are continuously perfused with nutrient-rich media, allowing for the continuous extrusion of straight fibers. The ultimate goal is to develop a scalable platform for the continuous production of cellulose nanofibers that can then be converted into carbon. This can become a more sustainable alternative to current practices of cellulose production by eliminating the cost and energy required for forest management and cellulose extraction and purification. Most importantly, it eliminates the use of oil in the production of carbon fibers. In this work, we present initialcharacterization of the response of G. xylinus to electrical stimuli of varying frequencies as well as the optimized protocol to convert bacterial cellulose into carbon. Dielectrophoresis experiments were completed using AC signals of 1, 5, 10, 15, and 20MHz frequencies at a constant voltage of 20Vpp. The results indicate a positive DEP response across the spectrum of 1-20MHz with the highest trapping at a frequency in the range of 15-20 MHz (figure 1 and 2). Additional work focused on the carbonization of bacterial cellulose nanofibers. Amorphous carbon was successfully achieved (figure 3) through heat treatment of bacterial cellulose to 900 ºC in a nitrogen atmosphere following a 5 ºC/min heating ramp. A dwell time of 30 minutes at a temperature of 300 ºC was included in the heat treatment to induce as much oxygen release from the sample as possible to prevent burning the piece when increasing the temperature. Such step is also necessary because the removal of the hydroxyl groups stabilizes the carbon polymer by producing conjugated double bonds, forming an aromatic structure [2]. An optimal heating rate was determined to be 5 ºC/min to achieve high carbon yield without making the process prohibitively long. Although we observed cellulose to carbonize at around 400 ºC, 900 ºC was used as the final temperature to ensure a high degree of carbonization [3]. Ongoing work is on characterizing the fiber properties and dimensions depending on the suspending media and its perfusion rate over the cells. The goal is to understand how the conditions in the microreactor lead to the tailoring of the fiber properties and a maximized throughput. ******INSERT FIGURE 1 HERE******* *****CAPTION****Figure 1: Characterizing the response of G. xylinus to electrical stimuli using Carbon DEP ******INSERT FIGURE 2 HERE******* *****CAPTION****Figure 2: A 2D carbon electrode showing indications of inducing a positive dielectrophoretic force on the G. xylinus bacterium (A.) Electrode is electrically polarized (B.) Same electrode immediately after electric field is switched off ******INSERT FIGURE 3 HERE******* *****CAPTION****Figure 3: Carbonized Cellulose Nanofibrils

Ultralow-Power Electronic Trapping of Nanoparticles with Sub-10 nm Gold Nanogap Electrodes.

We demonstrate nanogap electrodes for rapid, parallel, and ultralow-power trapping of nanoparticles. Our device pushes the limit of dielectrophoresis by shrinking the separation between gold electrodes to sub-10 nm, thereby creating strong trapping forces at biases as low as the 100 mV ranges. Using high-throughput atomic layer lithography, we manufacture sub-10 nm gaps between 0.8 mm long gold electrodes and pattern them into individually addressable parallel electronic traps. Unlike pointlike junctions made by electron-beam lithography or larger micron-gap electrodes that are used for conventional dielectrophoresis, our sub-10 nm gold nanogap electrodes provide strong trapping forces over a mm-scale trapping zone. Importantly, our technology solves the key challenges associated with traditional dielectrophoresis experiments, such as high voltages that cause heat generation, bubble formation, and unwanted electrochemical reactions. The strongly enhanced fields around the nanogap induce particle-transport speed exceeding 10 μm/s and enable the trapping of 30 nm polystyrene nanoparticles using an ultralow bias of 200 mV. We also demonstrate rapid electronic trapping of quantum dots and nanodiamond particles on arrays of parallel traps. Our sub-10 nm gold nanogap electrodes can be combined with plasmonic sensors or nanophotonic circuitry, and their low-power electronic operation can potentially enable high-density integration on a chip as well as portable biosensing.

Discriminating dengue-infected hepatic cells (WRL-68) using dielectrophoresis.

Dielectrophoresis (DEP), the induced movement of dielectric particles placed in a nonuniform electric field, has been used as a potential technique for manipulation and separation of many biological samples without destructive consequences to the cell. Cells of the same genotype in different physiological and pathological states have unique morphological and structural features, therefore, it is possible to differentiate between them using their DEP responses. This paper reports the experimental discrimination of normal and dengue-infected human hepatic fetal epithelial cells (WRL-68 cells) based on their DEP crossover frequency, at which no resultant movement occurs in the cells in response to the DEP force. A microarray dot electrode was used to conduct the DEP experiments. The DEP forces applied to the cells were quantified by analyzing the light intensity shift within the electrode's dot region based on the Cumulative Modal Intensity Shift image analysis technique. The differences in dielectric properties between infected and uninfected cells were exploited by plotting a unique DEP spectrum for each set of cells. We observed that the crossover frequency decreased from 220 kHz for the normal WRL-68 cells to 140 kHz after infection with the dengue virus in a medium conductivity of 100 μS/cm. We conclude that the change in the DEP crossover frequency between dengue-infected cells and their healthy counterparts should allow direct characterization of these cell types by exploiting their electrophysiological properties.

Protein dielectrophoresis and the link to dielectric properties.

There is a growing interest in protein dielectrophoresis (DEP) for biotechnological and pharmaceutical applications. However, the DEP behavior of proteins is still not well understood which is important for successful protein manipulation. In this paper, we elucidate the information gained in dielectric spectroscopy (DS) and electrochemical impedance spectroscopy (EIS) and how these techniques may be of importance for future protein DEP manipulation. EIS and DS can be used to determine the dielectric properties of proteins predicting their DEP behavior. Basic principles of EIS and DS are discussed and related to protein DEP through examples from previous studies. Challenges of performing DS measurements as well as potential designs to incorporate EIS and DS measurements in DEP experiments are also discussed.

Preparation and Experimental Study on Dielectrophoresis-Based Microfluidic Chip for Cell Patterning

Abstract A dielectrophoresis (DEP)-based microfluidic chip for cell patterning was designed and fabricated to achieve non-contact and batch manipulation of cells. The microfluidic chip employed a polydimethylsiloxane (PDMS) microchannel and two indium tin oxide (ITO) electrodes which were designed as a “step” structure. The distribution of the electric field caused by the microelectrodes was simulated by finite element simulation software COMSOL. The position of the maximum intensity of the electric field was also determined. The ITO microelectrodes and PDMS microchannel were fabricated using the micro-electro-mechanical system (MEMS) technique. After oxygen plasma surface treatment, the PDMS microchannel and glass substrate with the ITO microelectrodes were aligned and bonded to form the experimental microfluidic chip. Through DEP experiments with varying frequencies, the DEP behavior of yeast cells was examined, and the electric field frequency of both positive and negative DEP responses were determined. The results showed that yeast cells with solution conductivity of 60 μS cm−1 exhibited negative DEP movement in a frequency range of 1–10 kHz, positive DEP movement from 100 kHz to 10 MHz, and no DEP movement at 50 kHz. Under a sinusoidal potential of 8Vp-p with frequency of 5 MHz, the yeast cells were aligned into chains along the “step” edge of microelectrodes.

Cytoplasm Resistivity of Mammalian Atrial Myocardium Determined by Dielectrophoresis and Impedance Methods

Many cardiac arrhythmias are caused by slowed conduction of action potentials, which in turn can be due to an abnormal increase of intracellular myocardial resistance. Intracellular resistivity is a linear sum of that offered by gap junctions between contiguous cells and the cytoplasm of the myocytes themselves. However, the relative contribution of the two components is unclear, especially in atrial myocardium, as there are no precise measurements of cytoplasmic resistivity, Rc. In this study, Rc was measured in atrial tissue using several methods: a dielectrophoresis technique with isolated cells and impedance measurements with both isolated cells and multicellular preparations. All methods yielded similar values for Rc, with a mean of 138 ± 5 Ω·cm at 23°C, and a Q10 value of 1.20. This value is about half that of total intracellular resistivity and thus will be a significant determinant of the actual value of action potential conduction velocity. The dielectrophoresis experiments demonstrated the importance of including divalent cations (Ca2+ and Mg2+) in the suspension medium, as their omission reduced cell integrity by lowering membrane resistivity and increasing cytoplasm resistivity. Accurate measurement of Rc is essential to develop quantitative computational models that determine the key factors contributing to the development of cardiac arrhythmias.

Interpretation of ac dielectrophoretic behavior of tin oxide nanobelts using Maxwell stress tensor approach modeling

In order to explain the dielectrophoretic behavior of ethanol suspended tin oxide (SnO 2) nanobelts observed experimentally, modeling of the ac dielectrophoresis (DEP) phenomenon is carried out. Nanobelts are a type of ribbon like semiconducting oxide nanowire with rectangular cross-sections. To calculate the DEP induced forces and torques the Maxwell stress tensor (MST) approach is used. DEP experiments have indicated negative DEP (repulsion) in the low frequency range (<100 kHz) and positive DEP (attraction) in the high frequency range (∼1–10 MHz) of the applied ac electric field. The experimentally observed DEP characteristic is unusual if the nanobelt is treated as possessing uniform bulk material electrical properties of tin oxide. Several different nanobelt models are studied using simulations to explain the unusual behavior. The model providing the best explanation is the one where the nanobelt has an electrically conductive interior and a non-conductive outer layer, depleted of charge carriers. This is consistent with the surface depletion phenomenon commonly observed in SnO 2 and other semiconducting metal oxide materials. This work demonstrates the importance of surface dominant properties of nanostructures in DEP manipulation and assembly.

Electrorotation of Metallic Microspheres

Electrorotation (ER) experiments of gold-coated micrometer-sized spheres suspended in an electrolyte are presented for three different ionic conductivities over the frequency range of 100 Hz to 40 MHz. The direction of rotation was observed to be counter-field (opposite to the rotating field vector) with a single rotation peak. The maximum in rotation occurs for a frequency on the order of the reciprocal RC time constant for charging the double layer at the gold surface. Dielectrophoresis (DEP) experiments showed that the gold-coated particles undergo negative DEP at low frequencies and positive DEP at high frequencies with the same relaxation frequency as electrorotation. No induced charge electrophoresis was noticeable in ER or DEP experiments.

Dielectrophoretic manipulation of human chromosomes in microfluidic channels: extracting chromosome dielectric properties

An investigation of the dielectric properties of polyamine buffer prepared human chromosomes is presented in this paper. Chromosomes prepared in this buffer are only a few micrometers in size and shaped roughly like spherical discs. Dielectrophoresis was therefore chosen as the method of manipulation combined with a custom designed microfluidic system containing the required electrodes for dielectrophoresis experiments. Our results show that although this system is presently not able to distinguish between the different chromosomes, it can provide average data for the dielectric properties of human chromosomes in polyamine buffer. These can then be used to optimize system designs for further characterization and even sorting. The experimental data from the dielectrophoretic manipulation were combined with theoretical calculations to extract a range of values for the permittivity and conductivity of human polyamine buffer prepared chromosomes.

Application of capillary electrophoresis to predict crossover frequency of polystyrene particles in dielectrophoresis

CE is used to calculate the zeta potential of polystyrene particles in order to estimate the surface conductivity of the particles. These values are used to predict the dielectrophoretic behavior, including the crossover frequency of particles under the influence of an AC electric field. Predictions for fluorescent polystyrene particles that are unmodified, carboxylate-modified, or streptavidin-modified are tested using miniaturized dielectrophoresis cells containing patterned gold quadrupole electrodes. The particle surface conductivities derived from CE separations differ from those derived from dielectrophoresis experiments by < or =0.003 S/m and serve as a guide to effectively select the suspending medium to generate a crossover frequency approximately 0.5 MHz. Prediction of dielectrophoresis crossover frequency is limited by several factors, especially the accuracy and precision of the conductivity measurements for the particle and suspending medium. A rapid analysis of particles by CE is a viable alternative to determine zeta potential.

Ac Dielectrophoresis of Tin Oxide Nanobelts Suspended in Ethanol: Manipulation and Visualization

This article presents results of detailed and direct real-time observations of the wide variety of SnO(2) nanobelt motions induced by ac dielectrophoresis (DEP) in an innovative microfluidic setup. High ac electric fields were generated on a gold microelectrode (approximately 20 microm electrode gap) array, patterned on a glass substrate and covered by a approximately 10 microm tall polydimethylsiloxane (PDMS) microchannel. Ethanol suspended SnO(2) nanobelts were introduced into the microchannel, and the DEP experiments were performed. Negative DEP (repulsion) of the nanobelts was observed in the low-frequency range (<100 kHz) of the applied electric field, which caused rigid body motion as well as deformation of the nanobelts. The negative DEP effect observed in ethanol is unusual and contrary to what is predicted by the Clausius-Mossotti factor (using bulk SnO(2) conductivity and permittivity values) of the dipole approximation theory. In the high-frequency range (approximately 1-10 MHz), positive DEP (attraction) of the nanobelts was observed. Pearl chain formation involving short nanobelts and particles was also observed in the two DEP regimes.

Bulk electrical properties of single‐walled carbon nanotubes immobilized by dielectrophoresis: Evidence of metallic or semiconductor behavior

We report the electrical characterization of single-walled carbon nanotubes (SWCNTs) trapped between two electrodes by dielectrophoresis (DEP). At high frequency, SWCNTs collected by DEP are expected to be of metallic type. Indeed current-voltage (I-V) measurements for devices made at 10 MHz show high values of conductivity and exhibit metallic behavior with linear and symmetric electrical features attributed to ohmic conduction. At low frequency, SWCNTs attracted by DEP are expected to be of semiconducting nature. Devices made at 10 kHz behave as semiconductors and demonstrate nonlinear and rectifying electrical characteristics with conductivities many orders of magnitude below the sample resulting from high-frequency immobilization of SWCNTs. Conducting atomic force microscopy (C-AFM) and current density calculation results are presented to reinforce results obtained by I-V measurements which clearly show type separation of SWCNTs after DEP experiments.

Electrokinetic measurements of membrane capacitance and conductance for pancreatic -cells

Membrane capacitance and membrane conductance values are reported for insulin secreting cells (primary -cells and INS-1 insulinoma cells), determined using the methods of dielectrophoresis and electrorotation. The membrane capacitance value of 12.57 (+/-1.46) mFm(-2), obtained for -cells, and the values from 9.96 (+/-1.89) mFm(-2) to 10.65 (+/-2.1) mFm(-2), obtained for INS-1 cells, fall within the range expected for mammalian cells. The electrorotation results for the INS-1 cells lead to a value of 36 (+/-22) Sm(-2) for the membrane conductance associated with ion channels, if values in the range 2-3 nS are assumed for the membrane surface conductance. This membrane conductance value falls within the range reported for INS cells obtained using the whole-cell patch-clamp technique. However, the total 'effective' membrane conductance value of 601 (+/-182) Sm(-2) obtained for the INS-1 cells by dielectrophoresis is significantly larger (by a factor of around three) than the values obtained by electrorotation. This could result from an increased membrane surface conductance, or increased passive conduction of ions through membrane pores, induced by the larger electric field stresses experienced by cells in the dielectrophoresis experiments.