Dielectrophoretic Capture Research Articles

Interference-Free Detection of Genetic Biomarkers Using Synthetic Dipole-Facilitated Nanopore Dielectrophoresis.

The motion of polarizable particles in a nonuniform electric field (i.e., dielectrophoresis) has been extensively used for concentration, separation, sorting, and transport of biological particles from cancer cells and viruses to biomolecules such as DNAs and proteins. However, current approaches to dielectrophoretic manipulation are not sensitive enough to selectively target individual molecular species. Here, we describe the application of the dielectrophoretic principle for selective detection of DNA and RNA molecules using an engineered biological nanopore. The key element of our approach is a synthetic polycationic nanocarrier that selectively binds to the target biomolecules, dramatically increasing their dielectrophoretic response to the electric field gradient generated by the nanopore. The dielectrophoretic capture of the nanocarrier-target complexes is detected as a transient blockade of the nanopore ionic current, while any nontarget nucleic acids are repelled from the nanopore by electrophoresis and thus do not interfere with the signal produced by the target's capture. Strikingly, we show that even modestly charged nanocarriers can be used to capture DNA or RNA molecules of any length or secondary structure and simultaneously detect several molecular targets. Such selective, multiplex molecular detection technology would be highly desirable for real-time analysis of complex clinical samples.

Enumeration, Dielectrophoretic Capture, and Molecular Analysis of Circulating Tumor Cells.

The identification of therapeutically targetable mutations in circulating tumor cells (CTCs) from cancer patient blood is increasingly used to personalize patient care. Here, we describe a novel approach for the enumeration, capture, and molecular analysis of CTCs from blood using an FDA-approved CTC enrichment and enumeration platform followed by dielectrophoretic capture and next-generation sequencing.

Separation of Particles of Rare Earth Oxides by Dielectrophoresis

A major challenge in chemical engineering is the separation and purification of materials, especially of rare earth compounds. It has been reported that a dielectrophoretic (DEP) apparatus can be used for separating rare earth oxides. The DEP capture of REOs on screen electrode was observed with microscope. The concentration of ROEs was determined using a nephelometer and ICP-OES before and after injection into the DEP apparatus. The results show that three types of REOs, La2O3, CeO2 and Dy2O3, generate nDEP in the DEP capture apparatus. There are two factors: voltage and suspension concentration, that affect the capture rate. The capture rate increases as voltage rises within certain limits. The capture rate also increases with increase in the original suspension concentration. Different REOs show marked differences in the DEP capture conditions. Consequently, factors such as voltage and flow rate can be controlled to achieve separation of different REOs.

Negative dielectrophoretic capture and repulsion of single cells at a bipolar electrode: the impact of faradaic ion enrichment and depletion.

This paper describes the dielectrophoretic (DEP) forces generated by a bipolar electrode (BPE) in a microfluidic device and elucidates the impact of faradaic ion enrichment and depletion (FIE and FID) on electric field gradients. DEP technologies for manipulating biological cells provide several distinct advantages over other cell-handling techniques including label-free selectivity, inexpensive device components, and amenability to single-cell and array-based applications. However, extension to the array format is nontrivial, and DEP forces are notoriously short-range, limiting device dimensions and throughput. BPEs present an attractive option for DEP because of the ease with which they can be arrayed. Here, we present experimental results demonstrating both negative DEP (nDEP) attraction and repulsion of B-cells from each a BPE cathode and anode. The direction of nDEP force in each case was determined by whether the conditions for FIE or FID were chosen in the experimental design. We conclude that FIE and FID zones generated by BPEs can be exploited to shape and extend the electric field gradients that are responsible for DEP force.

Dielectrophoretic capture and genetic analysis of single neuroblastoma tumor cells.

Our understanding of the diversity of cells that escape the primary tumor and seed micrometastases remains rudimentary, and approaches for studying circulating and disseminated tumor cells have been limited by low throughput and sensitivity, reliance on single parameter sorting, and a focus on enumeration rather than phenotypic and genetic characterization. Here, we utilize a highly sensitive microfluidic and dielectrophoretic approach for the isolation and genetic analysis of individual tumor cells. We employed fluorescence labeling to isolate 208 single cells from spiking experiments conducted with 11 cell lines, including 8 neuroblastoma cell lines, and achieved a capture sensitivity of 1 tumor cell per 106 white blood cells (WBCs). Sample fixation or freezing had no detectable effect on cell capture. Point mutations were accurately detected in the whole genome amplification product of captured single tumor cells but not in negative control WBCs. We applied this approach to capture 144 single tumor cells from 10 bone marrow samples of patients suffering from neuroblastoma. In this pediatric malignancy, high-risk patients often exhibit wide-spread hematogenous metastasis, but access to primary tumor can be difficult or impossible. Here, we used flow-based sorting to pre-enrich samples with tumor involvement below 0.02%. For all patients for whom a mutation in the Anaplastic Lymphoma Kinase gene had already been detected in their primary tumor, the same mutation was detected in single cells from their marrow. These findings demonstrate a novel, non-invasive, and adaptable method for the capture and genetic analysis of single tumor cells from cancer patients.

Fabrication of carbon nanotube film directly grown on conductive stainless steel film and application to dielectrophoretic nanoparticle capture

Multi-walled carbon nanotubes (CNTs) were synthesized directly on stainless steel film thermally deposited on an alumina plate. To activate the growth of CNTs, the stainless steel film was reduced in H2 stream without oxidation step. The electrical resistivity of the CNT film synthesized by this way turned to be 1/190 of CNT film synthesized by a conventional way using catalyst preparation method with magnetron sputtering. Dielectrophoretic (DEP) particle capture was demonstrated using the patterned CNT film synthesized on stainless steel film, and it was observed that carbon nanohorns (CNHs) dispersed with Pd nanoparticles (Pd-CNHs) and Pd-Au alloy nanoparticles (Pd/Au-CNHs) were captured at the CNT electrodes due to the high electric field strength there. In this DEP capture, Pd-CNHs were enriched in the present condition. The temperature to deposit stainless steel film and the influence of oxidation step were also investigated for the growth of CNTs.

Dielectrophoretic trapping of nanoparticles with an electrokinetic nanoprobe

A high aspect ratio 3D electrokinetic nanoprobe is used to trap polystyrene particles (200 nm), gold nanoshells (120 nm), and gold nanoparticles (mean diameter 35 nm) at low voltages (<1 V(rms)). The nanoprobe is fabricated using room temperature self-assembly methods, without the need for nanoresolution lithography. The nanoprobe (150-500 nm in diameter, 2-150 μm in length) is mounted on the end of a glass micropipette, enabling user-specified positioning. The nanoprobe is one electrode within a point-and-plate configuration, with an indium-tin oxide cover slip serving as the planar electrode. The 3D structure of the nanoprobe enhances dielectrophoretic capture; further, electro-hydrodynamic flow enhances trapping, increasing the effective trapping region. Numerical simulations show low heating (1 K), even in biological media of moderate conductivity (1 S/m).

Abstract 3512: Dielectrophoretic capture and genetic analysis of individual disseminated solid tumor cells.

Abstract Background: The purpose of this study was to develop a novel approach for isolation and genetic analysis of individual circulating and disseminated tumor cells (CTC, DTC) from patient samples. This work is motivated by our desire to develop non-invasive molecular diagnostics and understand single tumor cell heterogeneity. Methods: Here we used the DEPArray microelectronics and microfluidics platform for the manipulation and capture of individual tumor cells. A suspension of live or fixed cells, prelabeled with fluorochrome-conjugated antibodies, was loaded into the DEPArray cartridge, and application of non-uniform electrical fields (dielectrophoresis) was employed to trap individual cells into movable micro-cages. Cell images were acquired via the incorporated fluorescence microscope and CCD camera, and cells were selected based on morphology and expression of tumor antigens. To determine sensitivity and specificity of the capture process, cell lines were spiked into normal donor white blood cells (WBC) over a 5-log range. Pre-enrichment by FACSAria was employed when the starting tumor cell concentration was below 1 in 1,000 WBC. The whole genome amplification (WGA) product of single cells was then subjected to targeted sequencing. To apply this approach to patient samples, we obtained blood and bone marrow samples from patients with refractory high-risk neuroblastoma, a cancer with a high rate and quantity of distant metastatic disease. Results: Single tumor cells were recovered from all spiking experiments, including spiked samples with a tumor cell concentration as low as 1 in 106 WBC. Recovery of EpCAM-negative single cells was accomplished with the neuroblastoma cell lines SY5Y, NB1643, and Kelly labeled with an antibody for the disialoganglioside GD2, the breast cancer cell line MCF-7 with a HER2 antibody, and for the lung cancer cell line H292 with EGFR labeling. Recovery of EpCAM-positive cells was achieved with the colon carcinoma cell line SW480. Captured single cells were WGA'd and specificity was verified using Sanger sequencing. For the 2 neuroblastoma spiking experiments for which sequencing has been completed, the known ALK mutation (F1174L) was detected in 100% of cells. The known KRAS mutation (G12V) was detected in 100% of single SW480 cells sequenced to date. No mutations were detected in isolated WBC. We next analyzed the post-Ficoll fraction of 12 patient bone marrow and 1 patient blood sample, with a median tumor cell concentration of 0.119% (range 0.003 - 80.0%). A minimum of 10 single tumor cells were captured per patient. Genome-wide analysis of DNA and RNA copy number and mutation status from these samples is ongoing. Conclusion: These data demonstrate the development of a novel workflow for the reliable capture and genetic analysis of rare CTC. Efforts are ongoing to join this approach with a more comprehensive genome-wide analysis of single cells from neuroblastoma and other cancers. Citation Format: Erica L. Carpenter, JulieAnn Rader, Kateryna Krytska, Jacob Ruden, Eric Rappaport, Yael Mosse, Francesca Fontana, Manuela Banzi, Gianni Medoro, Nicolo Manaresi, Peter O'Dwyer, John Maris. Dielectrophoretic capture and genetic analysis of individual disseminated solid tumor cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3512. doi:10.1158/1538-7445.AM2013-3512

Microfluidic-Based Amplification-Free Bacterial DNA Detection by Dielectrophoretic Concentration and Fluorescent Resonance Energy Transfer Assisted in Situ Hybridization (FRET-ISH).

Although real-time PCR (RT-PCR) has become a diagnostic standard for rapid identification of bacterial species, typical methods remain time-intensive due to sample preparation and amplification cycle times. The assay described in this work incorporates on-chip dielectrophoretic capture and concentration of bacterial cells, thermal lysis, cell permeabilization, and nucleic acid denaturation and fluorescence resonance energy transfer assisted in situ hybridization (FRET-ISH) species identification. Combining these techniques leverages the benefits of all of them, allowing identification to be accomplished completely on chip less than thirty minutes after receipt of sample, compared to multiple hours required by traditional RT-PCR and its requisite sample preparation.

Dielectrophoretic Cell Capture on Polyester Membranes

A new system for dielectrophoretic cell capture on permeable polyester membranes is presented. Conventional photolithographic techniques were used to fabricate gold microelectrodes on a polyester membrane. The characterization of the microelectrodes showed that there were no differences regarding roughness, permeability, and hydrophilicity of the membrane before and after processing. Finally, dielectrophoretic cell capture and viability in a microfluidic device was demonstrated on the patterned membrane. These membranes could ultimately be combined with multilayer microfluidic devices to form a powerful tool for studies of cell-cell interactions in coculture, whereby spatial separation of different cell types and/or microenvironments are required.

Dielectrophoretic capture voltage spectrum for measurement of dielectric properties and separation of cancer cells

In this paper, a new dielectrophoresis (DEP) method based on capture voltage spectrum is proposed for measuring dielectric properties of biological cells. The capture voltage spectrum can be obtained from the balance of dielectrophoretic force and Stokes drag force acting on the cell in a microfluidic device with fluid flow and strip electrodes. The method was demonstrated with the measurement of dielectric properties of human colon cancer cells (HT-29 cells). From the capture voltage spectrum, the real part of Clausius-Mossotti factor of HT-29 cells for different frequencies of applied electric field was obtained. The dielectric properties of cell interior and plasma membrane were then estimated by using single-shell dielectric model. The cell interior permittivity and conductivity were found to be insensitive to changes in the conductivity of the medium in which the cells are suspended, but the measured permittivity and conductivity of cell membrane were found to increase with the increase of medium conductivity. In addition, the measurement of capture voltage spectrum was found to be useful in providing the optimum operating conditions for separating HT-29 cells from other cells (such as red blood cells) using dielectrophoresis.

Dielectrophoretic capture of E. coli cells at micropatterned nanoelectrode arrays

This paper reports capture and detection of pathogenic bacteria based on AC dielectrophoresis (DEP) and electrochemical impedance spectroscopy (EIS) employing an embedded vertically aligned carbon nanofiber (VACNF) nanoelectrode array (NEA) versus a macroscopic indium-tin-oxide (ITO) transparent electrode in "points-and-lid" configuration. The nano-DEP device was fabricated using photolithography processes to define an exposed active region on a randomly distributed NEA and a microfluidic channel on ITO to guide the flow of labeled Escherichia coli cells, respectively, and then bond them into a fluidic chip. A high-frequency (100 kHz) AC field was applied to generate positive DEP at the tips of exposed CNFs. Enhanced electric field gradient was achieved due to reduction in electrode size down to nanometer scale which helped to overcome the large hydrodynamic drag force experienced by E. coli cells at high flow velocities (up to 1.6 mm/s). This DEP device was able to effectively capture a significant number of E. coli cells. Significant decrease in the absolute impedance at the NEA was observed in EIS experiments. The results obtained in this study suggest the possibility of integration of a fully functional electronic device for rapid, reversible and label-free capture and detection of pathogenic bacteria.

Negative dielectrophoretic capture of bacterial spores in food matrices

A microfluidic device with planar square electrodes is developed for capturing particles from high conductivity media using negative dielectrophoresis (n-DEP). Specifically, Bacillus subtilis and Clostridium sporogenes spores, and polystyrene particles are tested in NaCl solution (0.05 and 0.225 S∕m), apple juice (0.225 S∕m), and milk (0.525 S∕m). Depending on the conductivity of the medium, the Joule heating produces electrothermal flow (ETF), which continuously circulates and transports the particles to the DEP capture sites. Combination of the ETF and n-DEP results in different particle capture efficiencies as a function of the conductivity. Utilizing 20 μm height DEP chambers, "almost complete" and rapid particle capture from lower conductivity (0.05 S∕m) medium is observed. Using DEP chambers above 150 μm in height, the onset of a global fluid motion for high conductivity media is observed. This motion enhances particle capture on the electrodes at the center of the DEP chamber. The n-DEP electrodes are designed to have well defined electric field minima, enabling sample concentration at 1000 distinct locations within the chip. The electrode design also facilitates integration of immunoassay and other surface sensors onto the particle capture sites for rapid detection of target micro-organisms in the future.

Magnetically controlled dielectrophoresis of metallic colloids

We present a finite-element/discrete-element numerical model for calculating full trajectories of cylindrical metallic colloids in liquid flows and subjected to non-uniform electric fields. The effect of the particle orientation relative to the liquid flow is investigated by considering barcode magnetic nanowires pinned in different directions by applying uniform magnetic fields. We compare the motion of free as well as vertically and horizontally pinned nanowires and demonstrate that their nanoassembly may accurately be tuned by magnetically controlling the orientation during the dielectrophoretic capture.

Aerosol penetration properties of an electret filter with submicron aerosols with various operating factors

This study was undertaken to determine the effects of using an electret filter on aerosol penetration. Various factors, including particle size (0.05 to 0.5 μ m), aerosol charge state (neutral and single charge), face velocity (0.1, 0.3, 0.5 and 1.0 m/s), and relative humidity (RH 30% and RH 70%), were examined to assess their effects on aerosol collection characteristics. The results presented here demonstrate that the electric fields of the electret and discharged filter were −1.53 × 104 and −1.3 × 102 (V/m). The penetration through the electret filter with singly charged aerosol and neutral aerosol ranged from 0.4% to 13% and 14% to 29%, respectively. According to these results, the coulombic capture force was dominant for the smaller aerosol and the dielectrophoretic capture mechanism was considered important for the larger aerosol. The level of penetration through the electret filter increased with increasing face velocity and relative humidity. The temperature did not affect the penetration through the electret. Furthermore, from the regression analysis conducted during the operating conditions of this work, the aerosol charge was shown to exert the greatest influence on aerosol penetration.

Filtration characteristics of a fibrous filter pretreated with anionic surfactants for monodisperse solid aerosols

This study elucidates the effects of using an anionic surfactant-pretreated filter as an electret filter on the aerosol penetration. Anionic surfactants (sodium oleate, SO; sodium dodecyl sulfate, SDS) were used to pretreat polypropylene fibrous filters to make them negatively charged. Various factors, including the particle size (0.05– 0.5 μ m ), the aerosol charge state (Boltzmann-equilibrium charge, neutral and singly charge), the face velocity (0.1, 0.3, 0.5 and 1.0 m / s ), the species of aerosol (sodium chloride, NaCl and aluminum oxide, Al 2 O 3 ), the relative humidity (RH 30% and 70%) and the concentration of surfactant (0.01, 0.05 and 0.08 M) were considered to evaluate their effects on the aerosol collection characteristics. Experimental results from our study demonstrate that the aerosol penetrations through anionic surfactant-pretreated filters (ASPFs) were lower than through the untreated filter. The 0.3 μ m -aerosol penetrations through an untreated, 0.01, 0.05 and 0.08 M SO-pretreated filters with Boltzmann-equilibrium charged aerosol were about 78%, 53%, 45% and 40%. The penetration of 0.3- μ m aerosol of 0.01, 0.05 and 0.08 M SDS-pretreated filters with Boltzmann-equilibrium charged aerosol were, 62%, 58% and 51%. Pretreating the filter with anionic surfactant did not change the structure of the filter and the mechanical capture force. The electric field measured by an electrofieldmeter of the ASPFs was larger than that of untreated filter obviously. These findings imply that pretreatment with anionic surfactant made the filters charged. The surface charge of the ASPFs increased with the anionic-surfactant concentration. The increase of the collection efficiency of the ASPFs is in the range of 5–30%, and that depends on the aerosol size and surfactant concentration. Comparing between the aerosol penetration through the ASPFs with singly charged aerosol and that through the untreated filter with neutral aerosol indicates that the penetration through the ASPFs with singly charged aerosol decreases by a factor of 1.3–12.5. In contrast with the penetration through the ASPFs to that through the untreated filter with neutral aerosol, the penetration reduction factor of the ASPFs is in the range of 1.1–1.9. As the results shown, the Coulombic capture force is dominant for the smaller aerosol ( < 0.1 μ m ) and the dielectrophoretic capture mechanism works at larger aerosol size ( > 0.2 μ m ) . Additionally, the penetrations through the ASPFs increased with the face velocity. ASPFs performed better when the tested aerosol had a larger dielectric constant ( k ) such as the penetration through 0.05 M SO-pretreated filter with 0.3 μ m Al 2 O 3 aerosol ( k = 9.6 ) was 47% and that with NaCl aerosol ( k = 5.9 ) was about 53%. RH has no effect on the aerosol penetration through the ASPFs.

Electrostatic precipitation of charged particles from a turbulent flow between coaxial cylinders

Transport and deposition of charged particles from a turbulent flow within a cylindrical precipitator are analyzed theoretically. Particle collection efficiency is calculated for different diffusivity models. The effect of particle inertial properties upon the collection rate is studied. It is shown that the Deutsch formula for particle efficiency is valid for moderate values of the applied electrostatic field. The performances of parallel plate and cylindrical precipitators are compared on the basis of the calculated collection efficiencies. The effect of the dielectrophoretic capture mechanism upon the particle collection rate is evaluated.

FREQUENCY DEPENDENCE IN DIELECTROPHORESIS: TRAJECTORIES AND PARTICLE BUILDUP

A study of the effect of frequency of a sinusoidal field on dielectrophoretic capture is presented, using numerical analysis to compute particle trajectories. It is shown that at certain critical frequencies the trapping efficiency drops sharply due to weakening of the dielectrophoretic pull. Experiments in which the capture performance was visually recorded by photographing the particle buildup on a single rod at increasing frequencies are reported. Results are discussed in the light of the numerical analysis.

Dielectrophoretic capture on single rods in creeping flow: Theory, trajectories and experimental data

The theoretical aspects of dielectrophoresis with a rod matrix are presented briefly with emphasis on the creeping flow regime. Particle trajectories are computed numerically in the general case and the critical trajectory ordinates in the numerical solution are compared to their analytical counterpart. Experimental results obtained with a high-frequency field provide qualitative verification of the theory.

Influence of the sinusoidal field frequency on dielectrophoretic capture of a particle on a rod

Using a model describing dielectrophoretic capture of a single spherical particle on a single cylindrical rod, it is shown that the time taken by the particle to be collected is a function of the frequency of the applied sinusoidal field. For certain critical frequencies, a kind of anti-resonance is discerned with the time required for collection reaching a maximum. A parametric study of this phenomenon involving the competing forces (drag, gravity) was carried out using a numerical analysis. The phase of the sinusoidal field is also shown to be of great importance.