Dielectrophoresis Response Research Articles

Multi-Frequency DEP Cytometer Employing a Microwave Sensor for Dielectric Analysis of Single Cells

We present a microfluidic device for in-flow dielectric characterization of single biological cells. The dielectric spectrum is obtained by measuring the multiple-frequency dielectrophoresis (DEP) response of individual cells as they travel over an array of sensing and actuating electrodes. The DEP induced translation of each cell is detected by measuring the differential impedance of the array using a microwave interferometer, which is capable of sub-attofarad sensitivity, and is coupled to the sensing electrodes. The DEP response of a cell at multiple frequencies in the beta-dispersion region is chosen to discern particular cell dielectric properties as it travels along the array—such as cytoplasm conductivity and membrane capacitance. The Clausius–Mossotti factor of the cell is determined from the measured response signal in conjunction with numerical simulation of its trajectory. The approach is validated through measuring polystyrene microspheres. The DEP response of Chinese hamster ovary cells using two simultaneous frequencies is demonstrated.

Dielectric model for Chinese hamster ovary cells obtained by dielectrophoresis cytometry.

We present a dielectric model and its parameters for Chinese hamster ovary (CHO) cells based on a double-shell structure which includes the cell membrane, cytoplasm, nuclear envelope, and nucleoplasm. Employing a dielectrophoresis (DEP) based technique and a microfluidic system, the DEP response of many single CHO cells is measured and the spectrum of the Clausius-Mossotti factor is obtained. The dielectric parameters of the model are then extracted by curve-fitting to the measured spectral data. Using this approach over the 0.6-10 MHz frequency range, we report the values for CHO cells' membrane permittivity, membrane thickness, cytoplasm conductivity, nuclear envelope permittivity, and nucleoplasm conductivity. The size of the cell and its nuclei are obtained using optical techniques.

Differential dielectric responses of chondrocyte and Jurkat cells in electromanipulation buffers.

Electromanipulation of cells as a label-free cell manipulation and characterization tool has gained particular interest recently. However, the applicability of electromanipulation, particularly dielectrophoresis (DEP), to biological cells is limited to cells suspended in buffers containing lower amounts of salts relative to the physiological buffers. One might question the use of low conductivity buffers (LCBs) for DEP separation, as cells are stressed in buffers lacking physiological levels of salt. In LCB, cells leak ions and undergo volume regulation. Therefore, cells exhibit time-dependent DEP response in LCB. In this work, cellular changes in LCB are assessed by dielectric spectroscopy, cell viability assay, and gene expression of chondrocytes and Jurkats. Results indicate leakage of ions from cells, increases in cytoplasmic conductivity, membrane capacitance, and conductance. Separability factor, which defines optimum conditions for DEP cell separation, for the two cell types is calculated using the cellular dielectric data. Optimum DEP separation conditions change as cellular dielectric properties evolve in LCB. Genetic analyses indicate no changes in expression of ionic channel proteins for chondrocytes suspended in LCB. Retaining cellular viability might be important during dielectrophoretic separation, especially when cells are to be biologically tested at a downstream microfluidic component.

Liposomes as a model for the study of high frequency dielectrophoresis.

Liposomes were used as a physical model to study the dielectrophoretic response of single-shelled particles at high frequencies. For a typical particle, the single-shelled theoretical model predicts a lower cross-over frequency that depends upon the dielectric properties of the shell and an upper crossover frequency that depends upon the dielectric properties of the interior. Dried liposomes were rehydrated in media with conductivity ranging from 100 to 2000 μS/cm. The high frequency dielectrophoresis response of the liposomes was observed in the range of 1-80 MHz at 30 volts peak-to-peak, and the upper cross-over frequency was recorded. The experimental results closely matched the theoretical expectations. In particular, the upper cross-over frequency ranged from 9 to 60 MHz and was found to depend linearly on the interior conductivity of the liposome. These results further confirm the single-shell model at high-frequencies. Moreover, they suggest liposomes may be a useful model particle for use during the development of dielectrophoresis-based devices.

Enhancing sensitivity and specificity in rare cell capture microdevices with dielectrophoresis.

The capture and subsequent analysis of rare cells, such as circulating tumor cells from a peripheral blood sample, has the potential to advance our understanding and treatment of a wide range of diseases. There is a particular need for high purity (i.e., high specificity) techniques to isolate these cells, reducing the time and cost required for single-cell genetic analyses by decreasing the number of contaminating cells analyzed. Previous work has shown that antibody-based immunocapture can be combined with dielectrophoresis (DEP) to differentially isolate cancer cells from leukocytes in a characterization device. Here, we build on that work by developing numerical simulations that identify microfluidic obstacle array geometries where DEP-immunocapture can be used to maximize the capture of target rare cells, while minimizing the capture of contaminating cells. We consider geometries with electrodes offset from the array and parallel to the fluid flow, maximizing the magnitude of the resulting electric field at the obstacles' leading and trailing edges, and minimizing it at the obstacles' shoulders. This configuration attracts cells with a positive DEP (pDEP) response to the leading edge, where the shear stress is low and residence time is long, resulting in a high capture probability; although these cells are also repelled from the shoulder region, the high local fluid velocity at the shoulder minimizes the impact on the overall transport and capture. Likewise, cells undergoing negative DEP (nDEP) are repelled from regions of high capture probability and attracted to regions where capture is unlikely. These simulations predict that DEP can be used to reduce the probability of capturing contaminating peripheral blood mononuclear cells (using nDEP) from 0.16 to 0.01 while simultaneously increasing the capture of several pancreatic cancer cell lines from 0.03-0.10 to 0.14-0.55, laying the groundwork for the experimental study of hybrid DEP-immunocapture obstacle array microdevices.

Theoretical and experimental examination of particle–particle interaction effects on induced dipole moments and dielectrophoretic responses of multiple particle chains

Dielectrophoresis (DEP), an electrokinetic phenomenon based on particle polarizations in nonuniform electric fields, is increasingly employed for particle and cell characterizations and manipulations in microdevices. However, particle number densities are rarely varied and particle-particle interactions are largely overlooked, but both affect particle's effective polarizations by changing the local electric field, which directly impacts particle assembly into chains. This work examines theoretical and experimental particle-particle interactions and dielectrophoretic responses in nonuniform electric fields, then presents individual and chain velocities of spherical polystyrene microparticles and red blood cells (RBCs) under DEP forces in a modified quadruple electrode microdevice. Velocities are independently compared between 1, 2, 3, and 4 polystyrene beads and RBCs assembled into chains aligned with the electric field. Simulations compared induced dipole moments for particles experiencing the same (single point) and changing (multiple points) electric fields. Experiments and simulations are compared by plotting DEP velocities versus applied signal frequency from 1 kHz to 80 MHz. Simulations indicate differences in the DEP force exerted on each particle according to chain position. Simulations and experiments show excellent qualitative agreement; chains with more particles experienced a decrease in the DEP response for both polystyrene beads and RBCs. These results advance understanding of the extent that induced dipole polarizations with multiple particle chains affect observed behaviors in electrokinetic cellular diagnostic systems.

Dielectrophoresis Response and Manipulation of Silica and its Application to Micro/Nanosensors

In this paper, silica microspheres and several novel μm scale DEP (dielectrophoresis) micro electrode array chips were prepared. A DEP experimental system was assembled to investigate the the DEP response of silica microspheres and micrometer particles. The effects of pattern of electrodes and frequency of AC field on the manipulation of silica microspheres were analyzed. The results showed that silica microspheres were manipulated by negative DEP (nDEP) in all types of electrodes, the position of silica microspheres in the electrodes changed when the frequency was changed and the phenomenon of nDEP was more obvious at higher frequency. A gas sensor was fabricated by dielectrophoretically assembling SiO2 micrometer particles. The sensor showed good response to SO2 and NH3 gas. The measurement results confirmed DEP could be a quicker method for constructing gas sensor.

Frequency sweep rate dependence on the dielectrophoretic response of polystyrene beads and red blood cells

Alternating current (AC) dielectrophoresis (DEP) experiments for biological particles in microdevices are typically done at a fixed frequency. Reconstructing the DEP response curve from static frequency experiments is laborious, but essential to ascertain differences in dielectric properties of biological particles. Our lab explored the concept of sweeping the frequency as a function of time to rapidly determine the DEP response curve from fewer experiments. For the purpose of determining an ideal sweep rate, homogeneous 6.08 μm polystyrene (PS) beads were used as a model system. Translatability of the sweep rate approach to ∼7 μm red blood cells (RBC) was then verified. An Au/Ti quadrapole electrode microfluidic device was used to separately subject particles and cells to 10Vpp AC electric fields at frequencies ranging from 0.010 to 2.0 MHz over sweep rates from 0.00080 to 0.17 MHz/s. PS beads exhibited negative DEP assembly over the frequencies explored due to Maxwell-Wagner interfacial polarizations. Results demonstrate that frequency sweep rates must be slower than particle polarization timescales to achieve reliable incremental polarizations; sweep rates near 0.00080 MHz/s yielded DEP behaviors very consistent with static frequency DEP responses for both PS beads and RBCs.

Protein dielectrophoresis: advances, challenges, and applications.

Protein dielectrophoresis (DEP) has the potential to play an important role as a manipulation, fractionation, preconcentration, and separation method in bioanalysis and as manipulation tool for nanotechnological applications. The first demonstrations of protein DEP have been reported almost 20 years ago. Since then various experimental realizations to manipulate proteins by DEP as well as more targeted applications employing protein DEP have been demonstrated. This review summarizes the experimental studies in the field of protein DEP trapping and focusing as well as specific applications in separation, molecular patterning, on bioprobes and biosensors. While a comprehensive theoretical model describing protein DEP is still lacking we also attempt to provide an overview of the factors influencing protein DEP and relate to currently available theoretical models. We further point out the variations in experimental conditions used in the past to study the somewhat 20 proteins as well as the implications of protein molecular structure to the DEP response.

Alternating current-dielectrophoresis driven on-chip collection and chaining of green microalgae in freshwaters

The capability of the AC dielectrophoresis (DEP) for on-chip capture and chaining of microalgae suspended in freshwaters was evaluated. The effects of freshwater composition as well as the electric field voltage, frequency, and duration, on the dielectrophoretic response of microalga Chlamydomonas reinhardtii were characterized systematically. Highest efficiency of cell alignment in one-dimensional arrays, determined by the percentage of cells in chain and the chain length, was obtained at AC-field of 20 V mm(-1) and 1 kHz applied for 600 s. The DEP response and cell alignment of C. reinhardtii in water sampled from lake, pond, and river, as well as model media were affected by the chemical composition of the media. In the model media, the efficiency of DEP chaining was negatively correlated to the conductivity of the cell suspensions, being higher in suspensions with low conductivity. The cells suspended in freshwaters, however, showed anomalously high chaining at long exposure times. High concentrations of nitrate and dissolved organic matter decrease cell chaining efficiency, while phosphate and citrate concentrations increase it and favor formation of longer chains. Importantly, the application of AC-field had no effect on algal autofluorescence, cell membrane damage, or oxidative stress damages in C. reinhardtii.

Biomarker-free dielectrophoretic sorting of differentiating myoblast multipotent progenitor cells and their membrane analysis by Raman spectroscopy

Myoblasts are muscle derived mesenchymal stem cell progenitors that have great potential for use in regenerative medicine, especially for cardiomyogenesis grafts and intracardiac cell transplantation. To utilise such cells for pre-clinical and clinical applications, and especially for personalized medicine, it is essential to generate a synchronised, homogenous, population of cells that display phenotypic and genotypic homogeneity within a population of cells. We demonstrate that the biomarker-free technique of dielectrophoresis (DEP) can be used to discriminate cells between stages of differentiation in the C2C12 myoblast multipotent mouse model. Terminally differentiated myotubes were separated from C2C12 myoblasts to better than 96% purity, a result validated by flow cytometry and Western blotting. To determine the extent to which cell membrane capacitance, rather than cell size, determined the DEP response of a cell, C2C12 myoblasts were co-cultured with GFP-expressing MRC-5 fibroblasts of comparable size distributions (mean diameter ∼10 μm). A DEP sorting efficiency greater than 98% was achieved for these two cell types, a result concluded to arise from the fibroblasts possessing a larger membrane capacitance than the myoblasts. It is currently assumed that differences in membrane capacitance primarily reflect differences in the extent of folding or surface features of the membrane. However, our finding by Raman spectroscopy that the fibroblast membranes contained a smaller proportion of saturated lipids than those of the myoblasts suggests that the membrane chemistry should also be taken into account.

Real‐time cell electrophysiology using a multi‐channel dielectrophoretic‐dot microelectrode array

Dielectrophoresis (DEP) has been used for many years for the analysis of the electrophysiological properties of cells. However, such analyses have in the past been time-consuming, such that it can take 30 min or more to collect sufficient data to make valid interpretations from a single DEP spectrum. This has limited the application of the technology to a rapid tool for non-invasive, label-free research in areas from drug discovery to diagnostics. In this paper we present the development of a programmable, multi-channel DEP system for rapid biophysical assessment of populations of biological cells. A new assay format has been developed for continuous near-real-time monitoring, using simultaneous application of up to eight alternating current electrical signals to independently addressable dot microelectrodes in an array format, allowing a DEP spectrum to be measured in 20 s, with a total cycle time between measurements of 90 s. To demonstrate the system, human leukaemic K562 cells were monitored after exposure to staurosporine and valinomycin. The DEP response curves showed the timing and manner in which the membrane properties changed for the actions of these two drugs at the early phase of induction. This technology shows the great potential for increasing our understanding of the role of electrophysiology in drug action, by observing the changes in electrical characteristics as they occur.

Three‐dimensional focusing of particles using negative dielectrophoretic force in a microfluidic chip with insulating microstructures and dual planar microelectrodes

The focusing of biological and synthetic particles in microfluidic devices is a prerequisite for the construction of microstructured materials, as well as for medical applications. In the present study, a microdevice that can effectively focus particles in three dimensions using a combination of insulator-based and metal-electrode dielectrophoresis (DEP) has been designed and fabricated. The DEP force is employed to confine the particles using a negative DEP response. Four insulating microstructures, which form an X-pattern in the microchannel, were employed to distort the electric field between the insulators in a conducting solution, thereby generating regions with a high electric-field gradient. Two strips of microelectrodes on the top and bottom surfaces were placed in the middle of the microchannel and connected to an electric pole. Two sets of dual-planar electrodes connected to the opposite pole were placed at the sides of the microchannel at the top and bottom surfaces. The results of a transient simulation of tracks of polystyrene particles, which was performed using the commercial software package CFD-ACE⁺ (ESI Group, France), demonstrate that the three-dimensional focusing of particles was achieved when the applied voltage was larger than 35 V at a frequency of 1 MHz. Furthermore, the focusing performance increased with the increased strength of the applied electric field and decreased inlet flow rate. Experiments on particle focusing, employing polystyrene particles 10 μm in diameter, were conducted to demonstrate the feasibility of the proposed design; the results agree with the trend predicted by numerical simulations.

Dielectrophoretic manipulation of ribosomal RNA

The manipulation of ribosomal RNA (rRNA) extracted from E. coli cells by dielectrophoresis (DEP) has been demonstrated over the range of 3 kHz-50 MHz using interdigitated microelectrodes. Quantitative measurement using total internal reflection fluorescence microscopy of the time dependent collection indicated a positive DEP response characterized by a plateau between 3 kHz and 1 MHz followed by a decrease in response at higher frequencies. Negative DEP was observed above 9 MHz. The positive DEP response below 1 MHz is described by the Clausius-Mossotti model and corresponds to an induced dipole moment of 3300 D with a polarizability of 7.8×10(-32) F m(2). The negative DEP response above 9 MHz indicates that the rRNA molecules exhibit a net moment of -250 D, to give an effective permittivity value of 78.5 ε(0), close to that of the aqueous suspending medium, and a relatively small surface conductance value of ∼0.1 nS. This suggests that our rRNA samples have a fairly open structure accessible to the surrounding water molecules, with counterions strongly bound to the charged phosphate groups in the rRNA backbone. These results are the first demonstration of DEP for fast capture and release of rRNA units, opening new opportunities for rRNA-based biosensing devices.

Dielectrophoretic separation of mouse melanoma clones

Dielectrophoresis (DEP) is employed to differentiate clones of mouse melanoma B16F10 cells. Five clones were tested on microelectrodes. At a specific excitation frequency, clone 1 showed a different DEP response than the other four. Growth rate, melanin content, recovery from cryopreservation, and in vitro invasive studies were performed. Clone 1 is shown to have significantly different melanin content and recovery rate from cryopreservation. This paper reports the ability of DEP to differentiate between two malignant cells of the same origin. Different DEP responses of the two clones could be linked to their melanin content.

Preimplantation Mouse Embryo Selection Guided by Light-Induced Dielectrophoresis

Selection of optimal quality embryos for in vitro fertilization (IVF) transfer is critical to successful live birth outcomes. Currently, embryos are chosen based on subjective assessment of morphologic developmental maturity. A non-invasive means to quantitatively measure an embryo's developmental maturity would reduce the variability introduced by the current standard. We present a method that exploits the scaling electrical properties of pre-transfer embryos to quantitatively discern embryo developmental maturity using light-induced dielectrophoresis (DEP). We show that an embryo's DEP response is highly correlated with its developmental stage. Uniquely, this technique allows one to select, in sequence and under blinded conditions, the most developmentally mature embryos among a mixed cohort of morphologically indistinguishable embryos cultured in optimized and sub-optimal culture media. Following assay, embryos continue to develop normally in vitro. Light-induced dielectrophoresis provides a non-invasive, quantitative, and reproducible means to select embryos for applications including IVF transfer and embryonic stem cell harvest.

A Polymer Microfluidic Chip With Interdigitated Electrodes Arrays for Simultaneous Dielectrophoretic Manipulation and Impedimetric Detection of Microparticles

This paper presents the design, fabrication, and characterization of a polymer microfluidic biochip with integrated interdigitated electrodes arrays (IDAs) used to simultaneously separate, manipulate, and detect microparticles using dielectrophoresis (DEP) and electrochemical impedance spectroscopy (EIS) methods. The DEP response of silica microspheres has been characterized, and microspheres of different sizes (1.8 and 3.5 in diameter) have been DEP flow separated and individually trapped in different microchambers by IDAs in a single run. Simultaneously, the impedance change caused by microspheres captured on IDAs has been analyzed for quantification. High-throughput polymer microfabrication techniques such as micro injection molding were used in this work, so that the polymer microfluidic chip can be produced in a low-cost, disposable platform. This low-cost microfluidic chip provides a generic platform for developing multifunctional lab-on-a-chip devices that require the ability to handle and sense microparticles.