Adrienne R. Minerick

DC insulator dielectrophoretic applications in microdevice technology: a review

AbstractDielectrophoresis 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. FigureCommon insulating obstacle geometries reported in the literature. Red zones indicate where the particles experience the maximum dielectrophoretic effect under DC or DC plus AC-biased electric field conditions

Dielectrophoretic characterization of erythrocytes: Positive ABO blood types

Dielectrophoretic manipulation of erythrocytes/red blood cells is investigated as a tool to identify blood type for medical diagnostic applications. Positive blood types of the ABO typing system (A+, B+, AB+ and O+) were tested and cell responses quantified. The dielectrophoretic response of each blood type was observed in a platinum electrode microdevice, delivering a field of 0.025Vpp/μm at 1 MHz. Responses were recorded via video microscopy for 120 s and erythrocyte positions were tabulated at 20–30 s intervals. Both vertical and horizontal motions of erythrocytes were quantified via image object recognition, object tracking in MATLAB, binning into appropriate electric field contoured regions (wedges) and statistical analysis. Cells of O+ type showed relatively attenuated response to the dielectrophoretic field and were distinguished with greater than 95% confidence from all the other three blood types. AB+ cell responses differed from A+ and B+ blood types likely because AB+ erythrocytes express both the A and B glycoforms on their membrane. This research suggests that dielectrophoresis of untreated erythrocytes beyond simple dilution depends on blood type and could be used in portable blood typing devices.

A continuous DC-insulator dielectrophoretic sorter of microparticles

A lab-on-a-chip device is described for continuous sorting of fluorescent polystyrene microparticles utilizing direct current insulating dielectrophoresis (DC-iDEP) at lower voltages than previously reported. Particles were sorted by combining electrokinetics and dielectrophoresis in a 250 μm wide PDMS microchannel containing a rectangular insulating obstacle and four outlet channels. The DC-iDEP particle flow behaviors were investigated with 3.18, 6.20 and 10 μm fluorescent polystyrene particles which experience negative DEP forces depending on particle size, DC electric field magnitude and medium conductivity. Due to negative DEP effects, particles are deflected into different outlet streams as they pass the region of high electric field density around the obstacle. Particles suspended in dextrose added phosphate buffer saline (PBS) at conductivities ranging from 0.50 to 8.50 mS/cm at pH 7.0 were compared at 6.85 and 17.1V/cm. Simulations of electrokinetic and dielectrophoretic forces were conducted with COMSOL Multiphysics® to predict particle pathlines. Experimental and simulation results show the effect of medium and voltage operating conditions on particle sorting. Further, smaller particles experience smaller iDEP forces and are more susceptible to competing nonlinear electrostatic effects, whereas larger particles experience greater iDEP forces and prefer channels 1 and 2. This work demonstrates that 6.20 and 10 μm particles can be independently sorted into specific outlet streams by tuning medium conductivity even at low operating voltages. This work is an essential step forward in employing DC-iDEP for multiparticle sorting in a continuous flow, multiple outlet lab-on-a-chip device.

Direct current insulator-based dielectrophoretic characterization of erythrocytes: ABO-Rh human blood typing

A microfluidic platform developed for quantifying the dependence of erythrocyte (red blood cell, RBC) responses by ABO‐Rh blood type via direct current insulator dielectrophoresis (DC‐iDEP) is presented. The PDMS DC‐iDEP device utilized a 400×170 μm2 rectangular insulating obstacle embedded in a 1.46‐cm long, 200‐μm wide inlet channel to create spatial non‐uniformities in direct current (DC) electric field density realized by separation into four outlet channels. The DC‐iDEP flow behaviors were investigated for all eight blood types (A+, A−, B+, B−, AB+, AB−, O+, O−) in the human ABO‐Rh blood typing system. Three independent donors of each blood type, same donor reproducibility, different conductivity buffers (0.52–9.1 mS/cm), and DC electric fields (17.1–68.5 V/cm) were tested to investigate separation dependencies. The data analysis was conducted from image intensity profiles across inlet and outlet channels in the device. Individual channel fractions suggest that the dielectrophoretic force experienced by the cells is dependent on erythrocyte antigen expression. Two different statistical analysis methods were conducted to determine how distinguishable a single blood type was from the others. Results indicate that channel fraction distributions differ by ABO‐Rh blood types suggesting that antigens present on the erythrocyte membrane polarize differently in DC‐iDEP fields. Under optimized conductivity and field conditions, certain blind blood samples could be sorted with low misclassification rates.

Quantification of pH gradients and implications in insulator-based dielectrophoresis of biomolecules.

Direct current (DC) insulator‐based dielectrophoretic (iDEP) microdevices have the potential to replace traditional alternating current dielectrophoretic devices for many cellular and biomolecular separation applications. The use of large DC fields suggest that electrode reactions and ion transport mechanisms can become important and impact ion distributions in the nanoliters of fluid in iDEP microchannels. This work tracked natural pH gradient formation in a 100 μm wide, 1 cm‐long microchannel under applicable iDEP protein manipulation conditions. Using fluorescence microscopy with the pH‐sensitive dye FITC Isomer I and the pH‐insensitive dye TRITC as a reference, pH was observed to drop drastically in the microchannels within 1 min in a 3000 V/cm electric field; pH drops were observed in the range of 6–10 min within a 100 V/cm electric field and varied based on the buffer conductivity. To address concerns of dye transport impacting intensity data, electrokinetic mobilities of FITC were carefully examined and found to be (i) toward the anode and (ii) 1 to 2 orders of magnitude smaller than H+ transport which is responsible for pH drops from the anode toward the cathode. COMSOL simulations of ion transport showed qualitative agreement with experimental results. The results indicate that pH changes are severe enough and rapid enough to influence the net charge of a protein or cause aggregation during iDEP experiments. The results also elucidate reasonable time periods over which the phosphate buffering capacity can counter increases in H+ and OH− for unperturbed iDEP manipulations.

Explorations of ABO‐Rh antigen expressions on erythrocyte dielectrophoresis: Changes in cross‐over frequency

A quadrupole dielectrophoretic microdevice was utilized to examine the ABO‐Rh dependencies on erythrocyte polarizations. This important step toward medical microdevice technology would transform key clinical blood tests from the laboratory into the field. Previous work in dielectrophoretic microdevices demonstrated that the large number of ABO antigens on erythrocyte membranes impacts their dielectrophoretic signature at 1 MHz. This work explores the dielectrophoretic behavior of native human erythrocytes categorized by their ABO‐Rh blood types and directly compares these responses to the same erythrocyte sample modified to remove the A and B antigens. A β(1–3)‐galactosidase enzyme was utilized to cleave the ABO polysaccharide backbone at the galactosidase bonds. The enzymatic reaction was optimized by comparing agglutination of the native and modified blood cells in addition to UV–Vis and HPLC analysis of the reaction effluent for saccharide residues. Next, the dielectrophoretic behaviors of the native and modified erythrocytes were visually verified in a quadrupole electrode microdevice over a frequency range from 100 kHz to 80 MHz. The lower cross‐over frequency (COF), which transitions from negative to positive dielectrophoresis, for ABO blood types tested (A+, A−, B+, B−, AB+, O+ and O−) differed over the range from 17 to 47 MHz. The COFs of the corresponding enzyme‐modified erythrocytes were also determined and the range narrowed to 29–41 MHz. A second COF in the 70–80 MHz range was observed and was reduced in the presence of the transmembrane Rhesus factor. These results suggest that antigen expression on erythrocyte membrane surfaces influence cell polarizations in nonuniform AC fields.

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 particles 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.

Electrochemical hematocrit determination in a direct current microfluidic device

Hematocrit (HCT) tests are widely performed to screen blood donors and to diagnose medical conditions. Current HCT test methods include conventional microhematocrit, Coulter counter, CuSO4 specific gravity, and conductivity‐based point‐of‐care (POC) HCT devices, which can be either expensive, environmentally inadvisable, or complicated. In the present work, we introduce a new and simple microfluidic system for a POC HCT determination. HCT was determined by measuring current responses of blood under 100 V DC for 1 min in a microfluidic device containing a single microchannel with dimensions of 180 μm by 70 μm and 10 mm long. Current responses of red blood cell (RBC) suspensions in PBS or separately plasma at HCT concentrations of 10, 20, 25, 30, 35, 40, 45, 50, 55, 60, and 70 vol% were measured to show feasibility of the microfluidic system for HCT determination. Key parameters affecting current responses included electrolysis bubbles and irreversible RBC adsorption; parameters were optimized via addition of nonionic surfactant Triton X‐100 into sample solution and carbonizing electrode surfaces. The linear trend line of current responses over a range of RBC concentrations were obtained in both PBS and plasma. This work suggested that a simple microfluidic device could be a promising platform for a new POC HCT device.

Improving electrokinetic microdevice stability by controlling electrolysis bubbles

The voltage‐operating window for many electrokinetic microdevices is limited by electrolysis gas bubbles that destabilize microfluidic system causing noise and irreproducible responses above ∼3 V DC and less than ∼1 kHz AC at 3 Vpp. Surfactant additives, SDS and Triton X‐100, and an integrated semipermeable SnakeSkin® membrane were employed to control and assess electrolysis bubbles from platinum electrodes in a 180 by 70 μm, 10 mm long microchannel. Stabilized current responses at 100 V DC were observed with surfactant additives or SnakeSkin® barriers. Electrolysis bubble behaviors, visualized via video microscopy at the electrode surface and in the microchannels, were found to be influenced by surfactant function and SnakeSkin® barriers. Both SDS and Triton X‐100 surfactants promoted smaller bubble diameters and faster bubble detachment from electrode surfaces via increasing gas solubility. In contrast, SnakeSkin® membranes enhanced natural convection and blocked bubbles from entering the microchannels and thus reduced current disturbances in the electric field. This data illustrated that electrode surface behaviors had substantially greater impacts on current stability than microbubbles within microchannels. Thus, physically blocking bubbles from microchannels is less effective than electrode functionalization approaches to stabilize electrokinetic microfluidic systems.

Characterizing the dielectric properties of human mesenchymal stem cells and the effects of charged elastin-like polypeptide copolymer treatment

HUMAN MESENCHYMAL STEM CELLS (HMSCS) HAVE THREE KEY PROPERTIES THAT MAKE THEM DESIRABLE FOR STEM CELL THERAPEUTICS: differentiation capacity, trophic activity, and ability to self-renew. However, current separation techniques are inefficient, time consuming, expensive, and, in some cases, alter hMSCs cellular function and viability. Dielectrophoresis (DEP) is a technique that uses alternating current electric fields to spatially separate biological cells based on the dielectric properties of their membrane and cytoplasm. This work implements the first steps toward the development of a continuous cell sorting microfluidic device by characterizing native hMSCs dielectric signatures and comparing them to hMSCs morphologically standardized with a polymer. A quadrapole Ti-Au electrode microdevice was used to observe hMSC DEP behaviors, and quantify frequency spectra and cross-over frequency of hMSCs from 0.010-35 MHz in dextrose buffer solutions (0.030 S/m and 0.10 S/m). This combined approach included a systematic parametric study to fit a core-shell model to the DEP spectra over the entire tested frequency range, adding robustness to the analysis technique. The membrane capacitance and permittivity were found to be 2.2 pF and 2.0 in 0.030 S/m and 4.5 pF and 4.1 in 0.10 S/m, respectively. Elastin-like polypeptide (ELP-) polyethyleneimine (PEI) copolymer was used to control hMSCs morphology to spheroidal cells and aggregates. Results demonstrated that ELP-PEI treatment controlled hMSCs morphology, increased experiment reproducibility, and concurrently increased hMSCs membrane permittivity to shift the cross-over frequency above 35 MHz. Therefore, ELP-PEI treatment may serve as a tool for the eventual determination of biosurface marker-dependent DEP signatures and hMSCs purification.