Over the past decade, dielectrophoresis (DEP) has evolved into a powerful, robust and flexible method for cellular characterization, manipulation, separation and cell patterning. It is a field with widely varying disciplines, as it is quite common to see DEP integrated with a host of applications including microfluidics, impedance spectroscopy, tissue engineering, real-time PCR, immunoassays, stem-cell characterization, gene transfection and electroporation, just to name a few. The field is finally at the point where analytical and numerical polarization models can be used to adequately describe and characterize the dielectrophoretic behavior of cells, and there is ever increasing evidence demonstrating that electric fields can safely be used to manipulate cells without harm. As such, DEP is slowly making its way into the biological sciences. Today, DEP is being used to manipulate individual cells to specific regions of space for single-cell assays. DEP is able to separate rare cells from a heterogeneous cell suspension, where isolated cells can then be characterized and dynamically studied using nothing more than electric fields. However, there is need for a critical report to integrate the many new features of DEP for cellular applications. Here, a review of the basic theory and current applications of DEP, specifically for cells, is presented.

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

We report an interesting buffer electric relaxation time tuning technique, coupled with a glutaraldehyde cross-linking cell fixation reaction, which allows for sensitive dielectrophoretic analysis and discrimination of bovine red blood cell ͑bRBC͒ starvation age. The buffer composition is selected such that two easily accessible dielectrophoretic crossover frequencies ͑cof͒ exist. Low concentration glutaraldehyde fixation was observed to produce a threefold decrease in the higher cof with a comparable increase in the lower cof also witnessed. More importantly, increased glutaraldehyde fixation concentration significantly increased the higher cof by a factor found to be sensitive to the bRBC starvation age.

We report a novel buffer electric and dielectric relaxation time tuning technique, coupled with a glutaraldehyde (Glt.) cross-linking cell fixation reaction that allows for sensitive dielectrophoretic analysis and discrimination of bovine red blood cells of different starvation age. Guided by a single-shell oblate spheroid model, a zwitterion buffer composition is selected to ensure that two measurable crossover frequencies (cof's) near 500 kHz exist for dielectrophoresis (DEP) within a small range of each other. It is shown that the low cof is sensitive to changes in the cell membrane dielectric constant, in which cross-linking by Glt. reduces the dielectric constant of the cell membrane from 10.5 to 3.8, while the high cof is sensitive to cell cytoplasm conductivity changes. We speculate that this enhanced particle polarizability that results from the cross-linking reaction is because younger (reduced starvation time) cells possess more amino groups that the reaction can release to enhance the cell interior ionic strength. Such sensitive discrimination of cells with different age (surface protein density) by DEP is not possible without the zwitterion buffer and cleavage by Glt. treatment. It is then expected that rapid identification and sorting of healthy from diseased cells can be similarly sensitized.

DNA-DNA hybridization reactions on 100 nm oligonucleotide-functionalized silica nanoparticles are found to sensitively affect the amplitude and direction of the dielectrophoretic mobility of the particles at nanomolar target ssDNA concentrations. Such sensitivity permits visual detection of the hybridization event without fluorescent labeling and confocal microscopy by imaging the cross-over frequency (cof) of the particle suspension on a quadrupole electrode array. Strong correlation with effective particle radius and zeta-potential measurements suggests that the dielectrophoretic cof offers not just sensitive signatures for successful functionalization and hybridization but also those for three distinct DNA surface conformations that appear at different surface densities of hybridized DNA. A properly normalized cof calibration chart allows simplified quantification of the target ssDNA concentrations. These results provide a simple, rapid and portable genetic detection method compatible for use outside the laboratory.

We introduce a method for improved dielectrophoretic ͑DEP͒ discrimination and separation of viable and nonviable yeast cells. Due to the higher cell wall permeability of nonviable yeast cells compared with their viable counterpart, the crosslinking agent glutaraldehyde ͑GLT͒ is shown to selectively cross-link nonviable cells to a much greater extent than viable yeast. The DEP crossover frequency ͑cof͒ of both viable and nonviable yeast cells was measured over a large range of buffer conductivities ͑22 S / cm-400 S / cm͒ in order to study this effect. The results indicate that due to selective nonviable cell cross-linking, GLT modifies the DEP cof of nonviable cells, while viable cell cof remains relatively unaffected. To investigate this in more detail, a dual-shelled oblate spheroid model was evoked and fitted to the cof data to study cell electrical properties. GLT treatment is shown to minimize ion leakage out of the nonviable yeast cells by minimizing changes in cytoplasm conductivity over a large range of ionic concentrations. This effect is only observable in nonviable cells where GLT treatment serves to stabilize the cell cytoplasm conductivity over a large range of buffer conductivity and allow for much greater differences between viable and nonviable cell cofs. As such, by taking advantage of differences in cell wall permeability GLT magnifies the effect DEP has on the field induced separation of viable and nonviable yeasts.

Over the last decade, microfluidics has become increasingly popular in biology and bioengineering. While lab-on-a-chip fabrication costs have continued to decrease, the hardware required for delivering controllable fluid flows to the microfluidic devices themselves remains expensive and often cost prohibitive for researchers interested in starting a microfluidics project. Typically, microfluidic experiments require precise and tunable flow rates from a system that is simple to operate. While many labs use commercial platforms or syringe pumps, these solutions can cost thousands of dollars and can be cost prohibitive. Here, we present an inexpensive and easy-to-use constant pressure system for delivering flows to microfluidic devices. The controller costs less than half the price of a single syringe pump but can independently switch and deliver fluid through up to four separate fluidic inlets at known flow rates with significantly faster fluid response times. It is constructed of readily available pressure regulators, gauges, plastic connectors and adapters, and tubing. Flow rate is easily predicted and calibrated using hydraulic circuit analysis and capillary tubing resistors. Finally, we demonstrate the capabilities of the flow system by performing well-known microfluidic experiments for chemical gradient generation and emulsion droplet production.

Two ac polarization mechanisms, charge accumulation due to electrode double layer charging and bulk permittivity/conductivity gradients generated by Joule heating, are combined in the double layer by introducing zwitterions to produce a new ac electrokinetic pump with the largest velocity ͑Ͼ1 mm/ s͒ and flow penetration depth ͑100 m͒ reported for low-conductivity fluids. The large fluid velocity is due to a quartic scaling with respect to voltage, as is true of electrothermal flow, but exhibits a clear maximum at a frequency corresponding to the electrode double layer inverse RC time.The ability to direct fluid flow at length scales of the order of tens of microns is an essential requirement in labon-a-chip devices. Conventional solutions often involve scaling down peristaltic, diaphragm, or other popular macroscale mechanical pumps to microscale dimensions. 1 Most such pumps contain moving parts, which can lead to clogging and cell lyses, and render them unsuitable for biological applications. Due to such drawbacks, the use of electrokinetic ͑EK͒ micropumps with no moving parts to transport and mix fluid has attracted considerable attention. 2,3 The most popular include dc and ac micropumps. In a dc pump electro-osmotic ͑EO͒ flow is induced by an external tangential electric field E t produced by pair of electrodes placed on the inlet and outlet of a microchannel. 2 The applied field within the microchannel produces a tangential Maxwell force within the naturally polarized Debye layer on the channel wall, and through viscous dissipation drives a characteristic Smoluchowski slip velocity U s =−E t / , where , , and are the medium dielectric constant, zeta potential, and viscosity, respectively.A popular alternative to the dc pump is an ac pump, as the resulting acEO flow is driven by microelectrodes that can be embedded within the chip to affect more precise flow control. The ac field across the microelectrodes is sustained at such a high frequency ͑100-500 kHz͒ that Faradaic generation of air bubbles and ionic contaminants is avoided even at high voltages. In contrast, dc pumps necessarily involve a charge transfer reaction at the electrode and are not embeddable. That a zero-mean ac field can induce a nonzero time-averaged tangential Maxwell stress along a polarized electrode surface to drive the acEO flow is quite counterintuitive and was only recently understood. 3 The electric field induced electrode polarization occurs because the applied field sustains a time-varying charging current within the working electrolyte solution, charging the double layer on the electrode surface much like a capacitor. The accumulation of charge must partially, but not entirely, screen the external electric field such that there is still a tangential component of the electric field on the induced charge layer on each electrode to generate flow. As such, acEO flow has a distinct maximum at a frequency corresponding to the inverse RC time scale for the electrode-electrolyte circuit ͑about 100-500 kHz for most conditions͒ that scal...