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Microscopic objects change the apparent permittivity and conductivity of aqueous systems and thus their overall polarizability. In inhomogeneous fields, dielectrophoresis (DEP) increases the overall polarizability of the system by moving more highly polarizable objects or media to locations with a higher field. The DEP force is usually calculated from the object’s point of view using the interaction of the object’s induced dipole or multipole moments with the inducing field. Recently, we were able to derive the DEP force from the work required to charge suspension volumes with a single object moving in an inhomogeneous field. The capacitance of the volumes was described using Maxwell–Wagner’s mixing equation. Here, we generalize this system’s-point-of-view approach describing the overall polarizability of the whole DEP system as a function of the position of the object with a numerical “conductance field”. As an example, we consider high- and low conductive 200 µm 2D spheres in a square 1 × 1 mm chamber with plain-versus-pointed electrode configuration. For given starting points, the trajectories of the sphere and the corresponding DEP forces were calculated from the conductance gradients. The model describes watersheds; saddle points; attractive and repulsive forces in front of the pointed electrode, increased by factors >600 compared to forces in the chamber volume where the classical dipole approach remains applicable; and DEP motions with and against the field gradient under “positive DEP” conditions. We believe that our approach can explain experimental findings such as the accumulation of viruses and proteins, where the dipole approach cannot account for sufficiently high holding forces to defeat Brownian motion.

The new findings on Spinosaurus’ swim tail strongly suggest that Spinosaurus was a specialized deep-water predator. However, the tail must be seen in the context of the propelled body. The comparison of the flow characteristics of Spinosaurus with geometrically similar animals and their swimming abilities under water must take their Reynolds numbers into account and provide a common context for the properties of Spinosaurus’ tail and dorsal sail. Head shape adaptations such as the head crest reduced hydrodynamic disturbance and facilitated stealthy advance, especially when hunting without visual contact, when Spinosaurus could have used its rostral integumentary mechanoreceptors for prey detection. The muscular neck permitted ‘pivot’ feeding, where the prey’s escape abilities were overcome by rapid dorsoventral head movement, facilitated by crest-mediated lower friction.

The impedance of electrodes with adherent biological cells correlates with cell viability and proliferation. To model this correlation, we exploited the idea that the introduction of a highly conductive layer into the equatorial equipotential slice of a system with an oriented, freely suspended, single ellipsoidal cell may split the system into mirror-symmetrical halves without changing the field distribution. Each half possesses half of the system's impedance and contains a hemiellipsoidal cell attached to the conductive layer, which can be considered a bottom electrode. For a hemiellipsoidal adherent cell model (ACM) with standard electrical properties for the external and cellular media, the assumption of a bottom membrane and a subcellular cleft in the 100 nm range, as found in adherent cells, changed the potential distribution over a one-% range up to frequencies of 1 MHz. For simplicity, potential distributions for slices of spheroidal objects can be numerically calculated in 2D. The 2D distributions can be converted into three dimensions using simplified equations for the influential radii of spheroids. After the ACM approach was expanded to adherent cell patch models (APMs), the feasibility of our model modifications was tested using two criteria: the constancy of the equipotential plane touching the poles of ACMs or APMs and a comparison of the impedance, which could be numerically calculated from the overall current between the bottom electrode and a plane-parallel counter-electrode, with the impedance of the suspension obtained from Maxwell-Wagner's mixing equation applied to hemiellipsoidal cells.

The ubiquitous molecule spermidine is known for its pivotal roles in the contact mediation, fusion, and reorganization of biological membranes and DNA. In our model system, borosilicate beads were attached to atomic force microscopy cantilevers and used to probe mica surfaces to study the details of the spermidine-induced attractions. The negative surface charges of both materials were largely constant over the measured pH range of pH 7.8 to 12. The repulsion observed between the surfaces turned into attraction after the addition of spermidine. The attractive force was correlated with the degree of spermidine protonation, which changed from +3 to +1 over the measured pH range. The force was maximal at pH 7.8. To explain the observed pH and spermidine concentration dependence, two different theoretical approaches were used: a chemical model of the charge equilibrium of spermidine and Monte-Carlo simulations of the orientation of the rodlike spermidine molecules in the gap between the borosilicate and mica surfaces. Monte-Carlo simulations of the orientational ordering of the rodlike spermidine molecules suggested the induction of attractive interactions between the surfaces if the gap was bridged by the molecules. For larger gaps, the orientational distribution function of the spermidine molecules predicted a considerable degree of parallel attachment of the molecules to the surfaces, resulting in reduced effective surface charge densities of both surfaces, which reduced their electrostatic repulsion.

For ellipsoidal objects, the complex conductivity of the suspension depends on the objects' axis ratio and orientation. It can be described by analytical equations that were derived by combining the influential radius approach with the mixing equation of Maxwell and Wagner. Here, we consider conductive or insulating homogeneous spheroids, with their symmetry axes being oriented in parallel, in perpendicular or at random with respect to the external field. Considerations show that the field-induced orientations of both nonconductive and conductive objects will result in a reduction of the suspension's impedance and an increased dissipation of electrical energy.

The data presented in this article are related to the research article entitled “Increased osteoblast viability at alkaline pH in vitro provides a new perspective on bone regeneration” (doi: 10.1016/j.bbrep.2017.02.001; (Galow et al., 2017) [1]). The water soluble tetrazolium (WST) proliferation assay detects the metabolic activity of the respiratory chain of cultured cells. The assay is based on changes in the light absorbance resulting from the metabolism of WST-1 into formazane by mitochondrial succinate reductase. We present data of three different tests that were carried out to check whether WST assay readouts are pH-dependent. In a first test, a possible pH effect on the photometric measurements, for example by shifting the absorbance spectrum of the pH indicator of the cell culture medium, was excluded. Because the second test revealed a significant pH-dependence of the activity of the mitochondrial succinate reductase, a third long-term test was conducted to analyze possible changes of the pH dependence over time. The higher absorbance per one million cells at alkaline pH, which was approximately four-fold at pH 8.4 compared to the pH-7.4 reference on day one decayed gradually, with the pH-differences equilibrating over six days.

An animal model of deep brain stimulation (DBS) was used in in vivo studies of the encapsulation process of custom-made platinum/iridium microelectrodes in the subthalamic nucleus of hemiparkinsonian rats via electrical impedance spectroscopy. Two electrode types with 100-μm bared tips were used: i) a unipolar electrode with a 200-μm diameter and a subcutaneous gold wire counter electrode and ii) a bipolar electrode with two parallelshifted 125-μm wires. Miniaturized current-controlled pulse generators (130 Hz, 200 μA, 60 μs) enabled chronic DBS of the freely moving animals. A phenomenological electrical model enabled recalculation of the resistivity of the wound tissue around the electrodes from daily in vivo recordings of the electrode impedance over two weeks. In contrast to the commonly used 1 kHz impedance, the resistivity is independent of frequency, electrode properties, and current density. It represents the ionic DC properties of the tissue. Significant resistivity changes were detected with a characteristic decrease at approximately the 2nd day after implantation. The maximum resistivity was reached before electrical stimulation was initiated on the 8th day, which resulted in a decrease in resistivity. Compared with the unipolar electrodes, the bipolar electrodes exhibited an increased sensitivity for the tissue resistivity.

We investigated the effects of acute valproate (VPA) on mouse embryonic primary cortex cells (MEPCs). Intracellular ATP concentrations were compared with changes in the mean action potential (AP) frequencies of MEPC networks growing on microelectrode arrays. Our data implies biphasic reactions towards increasing VPA concentrations for both parameters. Intracellular ATP and mean AP frequencies increased around characteristic concentrations of 0.15 and 0.07mM to hormetic plateaus of approx. 120% and 160% of their controls, before fading around 17 and 1.7 mM, respectively. The biphasic in vitro behavior of the two parameters hinders a simple extraction of IC50 and Hillslope values. Different ways of data-fitting with single and double logistic functions are discussed. For a typical hormetic increase of 60% above control, IC50 and Hillslope were decreased by 37% and 15%, respectively. Despite these marginal effects at a logarithmic concentration scale, the hormetic and double logistic behavior of parameters may provide information on the mode of action of toxic compounds. Comparison of our values with the LD50 of mice, recalculated by normalization to body mass, suggests that a neurotoxic rather than a cytotoxic mechanism is killing the animals. The future use of cellular microsystems to replace animal experiments will motivate the development of new microsensors, as well as the consideration of newly accessible parameters in systems biology models.

Spinosaurus aegyptiacus was probably the largest predatory dinosaur of the Cretaceous period. A new study shows that it was a semiaquatic hunter. The function of Spinosaurus’ huge dorsal ‘sail’ remains unsolved, however. Three hypotheses have been proposed: (1) thermoregulation; (2) humpback storage; or (3) display. According to our alternative hypothesis, the submerged sail would have improved manoeuvrability and provided the hydrodynamic fulcrum for powerful neck and tail movements such as those made by sailfish or thresher sharks when stunning or injuring prey. Finally, it could have been employed as a screen for encircling prey underwater.

Breathing hyperbaric air or gas mixtures, for example during diving or when working underwater is known to alter the electrophysiological behavior of neuronal cells, which may lead to restricted cognition. During the last few decades, only very few studies into hyperbaric effects have been published, especially for the most relevant pressure range of up to 10 bar. We designed a pressurized measuring chamber to record pressure effects on the electrical activity of neuronal networks formed by primary cells of the frontal cortex of NMRI mice. Electrical activity was recorded with multi-electrode arrays (MEAs) of glass neuro chips while subjected to a step-by-step pressure increase from atmospheric pressure (1 bar) to 2 and 4 bar, followed by a decompression to 1 bar, in order to record recovery effects. The effects of pressure on the total spike rates (TSRs), which were averaged from at least 45 chips, were detected in two cell culture media with different compositions. In a DMEM medium with 6% horse serum, the TSR was increased by 19% after a pressure increase to 2 bar and remained stable at 4 bar. In NMEM medium with 2% B27, the TSR was not altered by a pressure increase to 2 bar but increased by 9% at 4 bar. After decompression to 1 bar, the activities decreased to 76% and 101% of their respective control levels in the two media. MEA recordings from neuronal networks in miniaturized hyperbaric measuring chambers provide new access for exploring the neuronal effects of hyperbaric breathing gases.

Dielectrophoresis (DEP), electrorotation (ROT), and electro-orientation were used for the dielectric spectroscopy of nucleated three-axial chicken red blood cells (CRBCs). Because the different AC-electrokinetic effects are not mutually independent, their DEP and ROT spectra were combined in ranges separated by the reorientation of the CRBCs in the inhomogeneous linear DEP and circular ROT fields. This behavior can be qualitatively described by a single-shell ellipsoidal model. Whereas in linear fields, the maximum of the Clausius-Mossotti factor along the three axes determines the orientated axis, in circular fields, the minimum of the factor determines the axis perpendicularly orientated to the field plane. Quantitatively, it has not been possible to find a consistent parameter set for fitting the DEP and ROT spectra, as well as the reorientation frequencies. Our ellipsoidal CRBC standard model had semiaxes of a = 7.7 μm, b = 4.0 μm, and c = 1.85 μm, a relative permittivity of 35 to 45 and conductivity of 0.36 to 0.04 S/m for the cytoplasm, combined with a specific capacitance of 10 to 14 mF/m2 and a conductivity of 3500 S/m2 for the cell membrane. The fits in different external conductivity ranges between external conductivities of 0.015 and 1.0 S/m were improved when the membrane capacitance was changed between 4 to 25 mF/m2 depending on the method used. A similar transition was reflected in the effective properties of a three-shell spherical model containing an internal membranous sphere with the geometry of the CRBC nucleus. Our findings suggest that the simultaneous interpretation of various AC-electrokinetic spectra is a step toward the dielectric fingerprinting of biological cells.

AC fields induce charges at the structural interfaces of particles or biological cells. The interaction of these charges with the field generates frequency-dependent forces that are the basis for AC-electrokinetic effects such as dielectrophoresis (DEP), electrorotation (ROT), electro-orientation, and electro-deformation. The effects can be used for the manipulation or dielectric single-particle spectroscopy. The observation of a particular effect depends on the spatial and temporal field distributions, as well as on the shape and the dielectric and viscoelastic properties of the object. Because the effects are not mutually independent, combined frequency spectra are obtained, for example, discontinuous DEP and ROT spectra with ranges separated by the reorientation of nonspherical objects in the linearly and circularly polarized DEP and ROT fields, respectively. As an example, the AC electrokinetic behavior of a three-axial ellipsoidal single-shell model with the geometry of chicken-red blood cells is considered. The geometric and electric problems were separated using the influential-radius approach. The obtained finite-element model can be electrically interpreted by an RC model leading to an expression for the Clausius-Mossotti factor, which permits the derivation of force, torque, and orientation spectra, as well as of equations for the critical frequencies and force plateaus in DEP and of the characteristic frequencies and peak heights in ROT. Expressions for the orientation in linearly and circularly polarized fields, as well as for the reorientation frequencies were also derived. The considerations suggested that the simultaneous registration of various AC-electrokinetic spectra is a step towards the dielectric fingerprinting of single objects.

Immune competence can be tested by challenging organisms with a set of infectious agents. However, disease control requirements impose restrictions on the infliction of infections upon domestic pigs. Alternatively, vaccinations induce detectable immune responses that reflect immune competence. Here, we tested this approach with tetanus toxoid (TT) in young domestic pigs. To optimize the vaccination protocol, we immunized the pigs with a commercial TT vaccine at the age of 21 or 35 days. Booster immunizations were performed either 14 or 21 days later. TT-specific antibodies in plasma as well as lymphoproliferative responses were determined both 7 and 14 days after booster immunization using ELISA and lymphocyte transformation tests, respectively. In addition, general IgG and IgM plasma concentrations and mitogen-induced proliferation were measured. The highest TT-specific antibody responses were detected when blood samples were collected 1 week after a booster immunization conducted 21 days after primary immunization. The pigs' age at primary immunization did not have a significant influence on TT-specific antibody responses. Similarly, the TT-specific proliferative responses were highest when blood samples were collected 1 week after booster immunization, while age and time of primary and booster immunization were irrelevant in our setup. While general IgG and IgM plasma levels were highly age dependent, there were no significant age effects for TT-specific immune responses. In addition, mitogen-induced proliferation was independent of immunization as well as blood sampling protocols. In summary, our model of TT vaccination provides an interesting approach for the assessment of immune competence in young pigs. The detected vaccination effects were not biased by age, even though our data were acquired from immune systems that were under development during our tests.

Human induced pluripotent stem cells can be differentiated into dopaminergic neurons (Dopa.4U). Dopa.4U neurons expressed voltage-gated NaV and KV channels and showed neuron-like spontaneous electrical activity. In automated patch clamp measurements with suspended Dopa.4U neurons, delayed rectifier K+ current (delayed KV) and rapidly inactivating A-type K+ current (fast KV) were identified. Examination of the fast KV current with inhibitors yielded IC50 values of 0.4 mM (4-aminopyridine) and 0.1 mM (tetraethylammonium). In manual patch clamp measurements with adherent Dopa.4U neurons, fast KV current could not be detected, while the delayed KV current showed an IC50 of 2 mM for 4-aminopyridine. The NaV channels in adherent and suspended Dopa.4U neurons showed IC50 values for tetrodotoxin of 27 and 2.9 nM, respectively. GABA-induced currents that could be observed in adherent Dopa.4U neurons could not be detected in suspended cells. Application of current pulses induced action potentials in approx. 70 % of the cells. Our results proved the feasibility of automated electrophysiological characterization of neuronal cells.

This article considers the electric and magnetic properties of biological matter for frequencies ranging from DC up to GHz, mainly focusing on electric phenomena, since the interaction with magnetic fields is usually weak for the generally low-magnetic susceptibility of biological matter. Impedance and AC electrokinetic techniques as methods that apply fields for exploring the passive properties of tissues and suspended cells are also considered.

Electrothermal micropumps (ETμPs) use local heating to create conductivity and permittivity gradients in the pump medium. In the presence of such gradients, an external AC electric field influences smeared spatial charges in the bulk of the medium. When there is also a symmetry break, the field-charge interaction results in an effective volumetric force resulting in medium pumping. The advantages of the ETμP principle are the absence of moving parts, the opportunity to passivate all the pump structures, homogeneous pump-channel cross-sections, as well as force plateaus in broad frequency ranges. The ETμPs consisted of a DC-heating element and AC field electrodes arranged in a 1000 μm × 250 μm × 60 μm (length × width × height) channel. They were processed as platinum structures on glass carriers. An equivalent-circuit diagram allowed us to model the frequency-dependent pumping velocities of passivated and nonpassivated ETμPs, which were measured at medium conductivities up to 1.0 S/m in the 300 kHz to 52 MHz frequency range. The temperature distributions within the pumps were controlled by thermochromic beads. Under resonance conditions, an additional inductance induced a tenfold pump-velocity increase to more than 50 μm/s at driving voltages of 5 V(rms). A further miniaturization of the pumps is viewed as quite feasible.

We developed different types of glass cell-culture chips (GC3s) for culturing cells for microscopic observation in open media-containing troughs or in microfluidic structures. Platinum sensor and manipulation structures were used to monitor physiological parameters and to allocate and permeabilize cells. Electro-thermal micro pumps distributed chemical compounds in the microfluidic systems. The integrated temperature sensors showed a linear, Pt1000-like behavior. Cell adhesion and proliferation were monitored using interdigitated electrode structures (IDESs). The cell-doubling times of primary murine embryonic neuronal cells (PNCs) were determined based on the IDES capacitance-peak shifts. The electrical activity of PNC networks was detected using multi-electrode arrays (MEAs). During seeding, the cells were dielectrophoretically allocated to individual MEAs to improve network structures. MEA pads with diameters of 15, 20, 25, and 35 µm were tested. After 3 weeks, the magnitudes of the determined action potentials were highest for pads of 25 µm in diameter and did not differ when the inter-pad distances were 100 or 170 µm. Using 25-µm diameter circular oxygen electrodes, the signal currents in the cell-culture media were found to range from approximately −0.08 nA (0% O2) to −2.35 nA (21% O2). It was observed that 60-nm thick silicon nitride-sensor layers were stable potentiometric pH sensors under cell-culture conditions for periods of days. Their sensitivity between pH 5 and 9 was as high as 45 mV per pH step. We concluded that sensorized GC3s are potential animal replacement systems for purposes such as toxicity pre-screening. For example, the effect of mefloquine, a medication used to treat malaria, on the electrical activity of neuronal cells was determined in this study using a GC3 system.

We present a label-free in vitro method for testing the toxic potentials of chemical substances using primary neuronal cells. The cells were prepared from 16-day-old NMRI mouse embryos and cultured on silicon chips ( www.bionas.de ) under the influence of different parathion concentrations with sensors for respiration (Clark-type oxygen electrodes), acidification (pH-ISFETs) and cell adhesion (interdigitated electrode structures, IDES). After 12 days in vitro , the sensor readouts were simultaneously recorded for 350 min in the presence of parathion applying a serial 1:3 dilution. The parathion-dependent data was fitted by logistic functions. IC 50 values of approximately 105 μM, 65 μM, and 54 μM were found for respiration, acidification, and adhesion, respectively. An IC 50 value of approximately 36 μM was determined from the intracellular ATP-levels of cells, which were detected by an ATP-luminescence assay using micro-well plates. While the intracellular ATP level and cell adhesion showed no deviation from a simple logistic decay, increases of approximately 29% in the respiration and 15% in the acidification rates above the control values were found at low parathion concentrations, indicating hormesis. These increases could be fitted by a modified logistic function. We believe that the label-free, continuous, multi-parametric monitoring of cell-metabolic processes may have applications in systems-biology and biomedical research, as well as in environmental monitoring. The parallel characterization of IC 50 values and hormetic effects may provide new insights into the metabolic mechanisms of toxic challenges to the cell.

Previous studies have shown that cellular function depends on rod-like membrane proteins, among them Bin/Amphiphysin/Rvs (BAR) proteins may curve the membrane leading to physiologically important membrane invaginations and membrane protrusions. The membrane shaping induced by BAR proteins has a major role in various biological processes such as cell motility and cell growth. Different models of binding of BAR domains to the lipid bilayer are described. The binding includes hydrophobic insertion loops and electrostatic interactions between basic amino acids at the concave region of the BAR domain and negatively charged lipids. To shed light on the elusive binding dynamics, a novel experiment is proposed to expand the technique of single-molecule AFM for the traction of binding energy of a single BAR domain.

An AC-field will influence smeared spatial charges in the bulk of a polarizable medium in the presence of conductivity and permittivity gradients. Such gradients can be generated by a temperature gradient induced by Joule heating by the AC-field itself or a separate heating element. Finally, the interaction of the AC-field with the spatial charges may drive an effective medium flow in asymmetric systems. The induced pumping force depends on the temperature and field gradients, as well as on the frequency of the driving AC-field. We present various designs for new AC electro-thermal micropumps (ETμPs) that exploit the spatially smeared bulk polarization principle. In some designs, an integrated heating element allows the temperature gradient to be adjusted. Advantages of the ETμP principle are the absence of any moving parts, force plateaus in broad frequency ranges, and a reversal of the pump direction in dependence on the driving frequency. The operating frequencies ranging from kHz to GHz avoid electrolytic electrode processes and electrode deterioration. Designs with evenly shaped cross-sections of the pump channel are possible. Maximum pumping velocities above 1 mm s−1 were observed in channels with a 120 × 60 μm2 cross-section at an induced temperature increase below 10 °C. For pump volumes of approximately 100 nl the usable conductivity range for aqueous media spans from the extremely low to above physiological values. A further miniaturization of the pumps is viewed as quite feasible.

This short communication reports on the innovative method of the local micro-invasive needle electroporation (LOMINE) of single adherent cells. The investigation of cellular reactions in living cell cultures represents a fundamental method, e.g. for drug development and environmental monitoring. Existing classical methods for intracellular measurements using, e.g. patch clamp techniques are time-consuming and complex. Present patch-on-chip systems are limited to the investigation of single cells in suspension. Nevertheless, the most part of the cells of the human body is adherently growing. Therefore, we develop a new chip system for the growth of adherent cells with 64 micro-structured needle electrodes as well as 128 dielectrophoretic electrodes, located within a measuring area of 1 mm 2 . With this analytical chip, the intracellular investigation of electro-chemical changes and processes in adherently growing cells will become possible in the near future. Here, we present first intracellular measurements with this chip system.

Gold nanostructuring of smooth gold disk-electrodes has been performed by electrodeposition. FE-SEM images indicate a strongly enlarged electrochemically active surface of the modified electrodes. This could be approved by the gold oxide reduction peak in cyclic voltammetry. The electrodes were also applied for sequence-specific DNA detection. For this, complementary single stranded capture probes were immobilized on the electrodes and consecutively hybridized with complementary and noncomplementary [OsO4(bipy)]-labeled DNA targets. We found that intermediate dehybridization steps did not decrease the sensitivity for the specific target. Moreover, the surface modification increased the voltammetric signal by an approx. factor of 9. Further, an enhanced linear calibration range was observed. The presented simple nanostructuring process holds great promise for many electrochemical sensor applications.

Our new type of on-chip micro-pump exploits the ac electro-kinetic forces acting in the volume of a fluid in the presence of a temperature gradient. No mechanically movable parts are used. The velocity of the pump flow observed depends on the frequency and strength of the driving ac field and on the temperature gradient across the pump channel. An integrated heating element allows the temperature gradient to be adjusted. Both ac field electrodes and heating element are platinum structures processed on a glass chip. The pump-channel walls and cover are made from polymer and thin-glass, respectively. In this paper, we present measurements of the fluid velocity as functions of the medium conductivity (0.1–1.3 S m−1) and field frequency (300 kHz–52 MHz), voltage across the field-electrode voltage (0–35 Vrms) and the heating element (1.1–3.6 V). Velocities of up to 120 µm s−1 were observed in the pump channel. The advantage of our new design is an evenly shaped cross-section of the pump channel, which reduces the risk of the channel becoming clogged by debris. Ac-electro-osmosis is not a predominant effect in our structures. Pumping could only be observed when the heating current and ac-pump field were applied simultaneously. The effects observed were simulated with the COMSOL Multiphysics program.

We describe a novel instrument and electrical circuit for sensitive electrochemical measurements at simultaneous direct electrode heating. The new measuring principle can be applied to working electrodes of various designs featuring two end contacts. In our experiments, the contacts were connected to a 100 kHz AC heating power supply and the potentiostat via the new inductor bridge circuit. A compact heating-generator housing contains all components necessary for sine wave generation as well as amplification and transformation of the heating power. The new arrangement yields high temperature cyclovoltammetric signals for the [Fe(CN)6]3−/4− redox system with a noise level superior to the earlier symmetrically branched wire electrode designs. Noise and disturbances are dramatically suppressed especially for high resistance electrodes such as glassy carbon electrodes. Without a center contact, the working electrode design is greatly simplified. This opens new opportunities for the design of a great variety of heated electrodes that may be arranged in arrays or consist of materials with relatively high resistivity such as carbon and conducting polymers.

The induction of transmembrane potentials (Δ) by an external field is the basis of numerous applications in biotechnology, cell-technology and medicine. We have developed new, simplified analytical equations that avoid the complicated description by the depolarizing factors. The equations apply to the Δ induced in cells resembling ellipsoids of rotation, i.e. spheroids by homogeneous dc or ac fields. They will be especially useful for experimental scientists. The equations describe the dependence of the Δ on the electric media properties, the field frequency and the axis ratio for oblate and prolate spheroids for which the symmetry semiaxis (c) is shorter and longer than the other two semiaxes (a and b, with a = b), respectively. According to the Schwan equation, an electric field E may induce a maximum Δ of 1.5aE at the poles of a spherical cell. For the poles of spheroidal cells, the maxima can be easily described by Δ = (a + 2c)E/2 and Δ = a(a + 2c)E/(a + c) for fields oriented along and perpendicular to the symmetry axis, respectively. For practically important shapes the error in the magnitude of Δ is smaller than 5% except along the c-axis for axis ratios larger than 2. Nevertheless, the errors vanish for the three limiting shapes of infinitely thin disc, sphere and cylinder.

The possible effects of high-frequency electromagnetic fields (EMF) on biological systems are a subject of public concern and scientific discussion. It is generally accepted that the absorption of part of the field energy may cause a temperature rise in biological tissue. Nevertheless, our setup aims to detect possible athermal effects on the electric activity of neuronal in vitro networks. Such networks were formed by primary neurons derived from the murine frontal cortex and cultivated on micro-sensor chips. The action potentials of the neurons were detected in real time by an integrated, electrically passive microelectrode array. For EMF exposure, the chips were introduced into a rectangular wave-guide that could be operated in the propagating or standing wave modes. The drive signals were either continuous waves (1.9–2.2 GHz) or a generic mobile phone signal (UMTS-standard) of up to approximately 8 W. An on-chip sensor allowed the temperature progression to be recorded. In addition, ISFETs and Clark-like electrodes were integrated for the on-chip detection of pH and O 2 , respectively.

Electrorotation (ER) spectra of human red blood cells (HRBCs) have been recorded in the frequency range from 10 kHz to 250 MHz in a 4-electrode microchip chamber. The cells were suspended at conductivities in the range from 0.02 to 3.00 S/m (corresponding to an ionic strength range from 1.6 to 343 mM) at temperatures between 10 degrees C and 35 degrees C. Generally, the characteristic frequencies as well as the rotation speeds of the first (membrane-dispersion) and second ER peaks increased with temperature. The rotation speed increase was largely correlated to the temperature dependence of the medium viscosity. Standard temperature dependencies were assumed for the conductivities and permittivities of cytoplasm, membrane, and external solution to explain the frequency shifts, starting from the cell parameters of Gimsa et al. [Gimsa et al., 1996, Biophys. J. 71: 495-506.]. The membrane capacitance was assumed to be temperature independent, based on the permittivity of alkyl-chains. Under these assumptions, the spectra could be well fitted only in a narrow temperature range around 20 degrees C. The temperature dependence of the first characteristic frequency was much stronger than predicted. In addition, around 15 degrees C, an anomalously high rotation speed was observed for the first peak at low external conductivities. Interestingly, this finding corresponds to the change in the chloride transport rate described by Brahm [Brahm, 1977, J. Gen. Physiol. 70: 283-306.].

The electrorotation method can be applied to characterize the passive electric properties of individual cells or particles. For this, their frequency-dependent speed of rotation is registered microscopically. Commonly, object rotation is induced in harmonic rotating fields which are generated in four electrode chambers. The rotation speed of the objects is proportional to the square of the field strength. In this study, we calculated the two-dimensional electric field distribution in electrorotation chips using the FEMLAB finite element program (Comsol, Sweden). For reasons of symmetry, a perfectly circular field is generated at the centers of the four electrode chambers. Nevertheless, the field strength is reduced with respect to the quotient of electrode voltage and distance for any electrode shape. Distant from the center, the field polarization is elliptical with an eccentricity increasing with the distance to the center. For optimizing the electrode shape, the deviations in torques for given distances from the center have been calculated. These deviations have been compared for various electrode shapes in order to find an optimal chip design. Another criterion was a large electrode distance in the corners of the electrode arrangement in order to avoid field hot spots and to minimize dielectrophoretic particle collection.

Deep brain stimulation, the electric stimulation of basal ganglia nuclei, is a treatment for movement disorders such as Parkinson's disease. The underlying mechanisms are studied in animals, e.g. rodents. Designs and materials of commercially available microelectrodes, as well as experimentally applied driving signals vary tremendously. We used finite integration modeling to compare the electric field and current density distributions induced by various electrodes. Current density or field strength "hot spots", which are located particularly at sites of high curvature and material interfaces coincided with corrosion and erosion at poles and insulation, respectively, as shown by scanning electron microscopy of stainless steel electrodes. Cell constants, i.e. geometry factors relating the electrode impedance to the specific medium conductivity, were calculated to determine the electrode voltage for a given stimulation current. Nevertheless, for electrodes of the same cell constant but of different geometry, current and field distributions may be very dissimilar. We found geometry-dependent limiting values of the stimulation current, above which electric tissue damage may occur. These values limit the reach of the stimulation signal for a given electrode geometry. Also, electrode geometries determine the shape of the stimulated tissue volume. This study provides tools for choosing the most appropriate geometry for targeting different-sized brain areas.

We consider the influence of the molecular structure of phospholipid membranes on their dielectric properties in the radio frequency range. Membranes have a stratified dielectric structure on the submolecular level, with the lipid chains forming a central hydrophobic layer enclosed by the polar headgroups (HGs) and bound water layers. In our numerical model, isotropic permittivities of 2.2 and 48.8 were assigned to the lipid chain and bound water layers, respectively. The HG region was assumed to possess an anisotropic static permittivity with 142.2 and 30.2 in the tangential and normal directions, respectively. The permittivities of the HG and bound water regions have been assumed to disperse at frequencies around 51 and 345 MHz to become 2.2 and 1.8, respectively, in both the normal and tangential directions. Electric field distribution and absorption were calculated for phospholipid vesicles with 75 nm radius as an example. Significant absorption has been obtained in the HG and bound water regions. Averaging the membrane absorption over the layers resulted in a decreased absorption below 1 GHz but a more than 10-fold increase above 1 GHz, compared to a model with a homogeneous membrane of averaged properties. We propose single particle dielectric spectroscopy by AC electrokinetics at low-bulk medium conductivities for an experimental verification of our model.

Potentially, lipid membranes possess a high tangential conductivity and permittivity due to their surface charges and the in-plane orientation of the headgroup dipoles. Electrically, membranes exhibit a sandwich structure with a largely isotropic centre formed by the fatty acid chains and confined by two anisotropic headgroup layers. Accordingly, we described spherical vesicles by an aqueous core covered by three shells. For a theoretical comparison, models with an anisotropic single shell and anisotropic homogeneous spheres were also considered. Two effects can be clearly demonstrated. (1) High tangential conductivities or permittivities may lead to cyclic variations in the phase of the electric potential in the radial direction, resulting in a hemi-shell structure of the electric potential inside the objects with oppositely charged facets. The thickness of the anisotropic shell restricts the number of phase oscillations. (2) Despite the strong local field inhomogeneities, an isotropic homogeneous Maxwellian equivalent body with an identical external field distribution exists for any of the anisotropic models. Its properties can be found from a comparison of the numerically calculated surface potential and the classical expression of the Clausius–Mossotti factor at any given frequency. The permittivity conductivity pairs obtained exhibit a sigmoidal-like frequency dependence.

Deep brain stimulation (DBS) is a therapy of movement disorders including Parkinson's disease (PD). Commercially available electrodes for animal models of Parkinson's disease vary in geometry and material. We characterized such electrodes and found a drift in their properties within minutes and up to about 60 h after immersion in cell culture medium, both with and without a stimulation signal. Electrode properties could largely be restored by proteolytic treatment for platinum/iridium electrodes but not for stainless steel ones. Short-term drift and irreversible aging could be followed by impedance measurements. Aging was accompanied by metal corrosion and erosion of the plastic insulation. For both materials, the degradation rates depended on the current density at the electrode surfaces. Fourier analysis of the DBS pulse (60 micros, repetition rate 130 Hz) revealed harmonic frequencies spanning a band of more than three decades, with significant harmonics up to the MHz range. The band is located in a window imposed by electrode processes and capacitive cell membrane bridging at the low and high frequency ends, respectively. Even though electrode processes are reduced at higher frequencies they only vanish above 1 MHz and cannot be avoided. Therefore, the use of inert electrode materials is of special importance. The neurotoxicity of iron makes avoiding stainless steel electrodes imperative. Future developments need to avoid the use of corrosive materials and current density hot spots at the electrode surface, and to reduce low frequency components in the DBS pulses in order to diminish electrode processes.

We present an analytical equation for the transmembrane voltage (Deltaphi) induced by a homogeneous AC field on arbitrarily oriented cells of the general ellipsoidal shape. The equation generalizes the Schwan equation for spherical cells and describes the dependence of Deltaphi on field frequency, cell size and shape, membrane capacitance, conductivities of cytoplasm, membrane and external medium, the location of the membrane site under consideration, and on the orientation of the cell with respect to the field. The derivation is based on the fact that the cytoplasm and the Maxwellian equivalent body of the whole cell are both of a general ellipsoidal shape and must thus exhibit constant local fields. The constant fields allow for a relatively simple description of the potentials on the internal and external membrane sides, leading to Deltaphi. For this, the properties of cytoplasm, membrane, and external medium have been introduced into a special, finite element model. We found that Deltaphi can be unambiguously defined for non-spherical cells, provided that the membrane thickness is thin in comparison to the cell dimensions.

Suspended cells may respond to AC polarization by orienting, deforming, moving or rotating. For modeling of ellipsoidal cells, a new dipole approach is proposed. Along each of the principal axis of the model, three finite elements of arbitrary but equal cross-sectional area for the interior, low conductive membrane shell and exterior are assumed. The length of the external medium elements is defined by influential radii which are related to the depolarizing factors. The model predicts the potential at the ellipsoid's surface leading to the induced dipole moment. The moment obtained is identical to the Laplace approach for homogeneous ellipsoids; in the single-shell case, it is slightly different. The reason is the constant shell thickness which overcomes the confocal thickness necessary for the Laplace solution. Expressions for electro-orientation, deformation, dielectrophoresis, and electrorotation are derived. In linearly and circularly polarized fields, different orientation spectra are predicted to occur. While in linearly polarized AC fields, particles are oriented along their axis of highest polarizability, in circularly polarized fields, the axis of lowest polarizability is oriented perpendicular to the plane of field rotation. Based on this finding, a new electro-orientation method is proposed. In dielectrophoresis and electrorotation, reorientations are predicted which lead to discontinuous spectra.

We present a new model for a variety of electric polarization effects on oblate and prolate homogeneous and single-shell spheroids. For homogeneous spheroids the model is identical to the Laplace model. For single-shell spheres of cell-like geometry the calculated difference of the induced dipole moments is in the thousandths range. To solve Laplace's equation for nonspherical single-shell objects it is necessary to assume a confocal shell, which results in different cell membrane properties in the pole and equator regions, respectively. Our alternative model addresses this drawback. It assumes that the disturbance of the external field due to polarization may project into the medium to a characteristic distance, the influential radius. This parameter is related to the axis ratio of the spheroid over the depolarizing factors and allows us to determine the geometry for a finite resistor-capacitor model. From this model the potential at the spheroid's surface is obtained and, consequently, the local field inside a homogeneous spheroid is determined. In the single-shell case, this is the effective local field of an equivalent homogeneous spheroid. Finally, integration over the volume yields the frequency-dependent induced dipole moment. The resistor-capacitor approach allowed us to find simple equations for the critical and characteristic frequencies, force plateaus and peak heights of deformation, dielectrophoresis and electrorotation for homogeneous and single-shell spheroids, and a more generalized equation for the induced transmembrane potential of spheroidal cells.

Particle polarization in AC fields depends on the temporal and spatial properties of the inducing field and on the dielectric properties of the suspension medium and the particles. In inhomogeneous fields the interaction of the induced dipole moment with the inducing field may give rise to particle translation (dielectrophoresis, DP). The frequency dependence of DP is mediated by the dispersions of different interfacial polarization processes. Consequently, DP spectroscopy yields information on internal particle structures. Common methods microscopically analyze the frequency behavior of the induced motion. Phase analysis light scattering (PALS) is a suitable light scattering method for the registration of extremely slow particle translation. We used PALS to analyze the DP behavior of polystyrene Latex particles in an optical two-electrode chamber. The dielectric particle data obtained are in correspondence to those of the literature. Our method offers statistical significance and computerized operation for dielectric particle spectroscopy.

When the response of a particle suspension to an external electric field is analyzed, measurement of the frequency dependence widens the parameter space and enables access to the inherent electric properties of the particles or, for known electric properties, to layer thicknesses, etc. The common impedance method registers the direct electric response to an applied field, whereas dielectrophoresis (DP) and electrorotation (ER) analyze the electrokinetic response of single particles. Since all responses are based on the same principles of interfacial polarization they can generally be described by a unified theory. This is demonstrated by a simple resistor–capacitor model describing the interdependence of the three methods. While the methods generally yield the same information, the particle properties are more clearly reflected in DP and ER spectra. The reason is that, in contrast to impedance, the latter methods are differential; i.e., they are based on the polarizability difference between the particles and the medium. Recently, a main disadvantage of the single particle methods, namely the limited external conductivity and frequency, was overcome by ultra-microelectrode chambers. Another drawback was the requirement of tedious microscopic measurements on single particles. Light scattering methods are proposed to improve the measuring procedure and statistical significance. The new methods, DP/phase analysis light scattering and ER/light scattering, are presented in detail in two additional manuscripts.

Dielectric dispersions of the polarizability of suspended particles can be directly measured by electrorotation (ER). The method records particle rotation spectra in rotating electric fields of the radio frequency range. The frequency dependence of ER spectra is characterized by individual rotation peaks which provide information on inherent electric particle structures. To avoid the inconveniences of microscopic registration of particle rotation we developed electrorotational light scattering (ERLS). Results on human red blood cells, platelets and conducting latices are presented. For all particles the obtained ERLS spectra corresponded to microscopic measurements. New methodological developments such as the introduction of optical microstructured chambers are discussed. These chambers possess a simple structure and expand the available frequency and conductivity range. A special measuring procedure which distinguishes between the rotation sense at the measuring frequency relative to the sense at a reference frequency is presented. The procedure is also useful to reject the contributions of field-induced particle motions other than ER and to enhance the sensitivity of ERLS in frequency ranges of smaller particle rotations.

Common dynamic light scattering (DLS) methods determine the size and zeta-potential of particles by analyzing the motion resulting from thermal noise or electrophoretic force. Dielectric particle spectroscopy by common microscopic electrorotation (ER) measures the frequency dependence of field-induced rotation of single particles to analyze their inherent dielectric structure. We propose a new technique, electrorotational light scattering (ERLS). It measures ER in a particle ensemble by a homodyne DLS setup. ER-induced particle rotation is extracted from the initial decorrelation of the intensity autocorrelation function (ACF) by a simple optical particle model. Human red blood cells were used as test particles, and changes of the characteristic frequency of membrane dispersion induced by the ionophore nystatin were monitored by ERLS. For untreated control cells, a rotation frequency of 2 s-1 was induced at the membrane peak frequency of 150 kHz and a field strength of 12 kV/m. This rotation led to a decorrelation of the ACF about 10 times steeper than that of the field free control. For deduction of ERLS frequency spectra, different criteria are discussed. Particle shape and additional field-induced motions like dielectrophoresis and particle-particle attraction do not significantly influence the criteria. For nystatin-treated cells, recalculation of dielectric cell properties revealed an ionophore-induced decrease in the internal conductivity. Although the absolute rotation speed and the rotation sense are not yet directly accessible, ERLS eliminates the tedious microscopic measurements. It offers computerized, statistically significant measurements of dielectric particle properties that are especially suitable for nonbiological applications, e.g., the study of colloidal particles.

Deep brain stimulation (DBS) is an invasive therapeutic option for patients with Parkinson’s disease (PD) but the mechanisms behind it are not yet fully understood. Animal models are essential for basic DBS research, because cell based in-vitro techniques are not complex enough. However, the geometry difference between rodents and humans implicates transfer problems of the stimulation conditions. For rodents, the development of miniaturized mobile stimulators and adapted electrodes are desirable. We implanted uni- and bipolar platinum/iridium electrodes in rats and were able to establish chronical instrumentation of freely moving rats (3 weeks). We measured the impedance of unipolar electrodes in-vivo to characterize the influence of electrochemical processes at the electrode-tissue interface. During the encapsulation process, the real part of the electrode impedance at 10 kHz doubled after 12 days and increased almost 10 times after 22 days. An outlook is given on the quantification of the DBS effect by sensorimotor behavioral tests.

Usually dielectrophoretic and electrorotation measurements are carried out at low ionic strength to reduce electrolysis and heat production. Such problems are minimized in microelectrode chambers. In a planar ultramicroelectrode chamber fabricated by semiconductor technology, we were able to measure the dielectric properties of human red blood cells in the frequency range from 2 kHz to 200 MHz up to physiological ion concentrations. At low ionic strength, red cells exhibit a typical electrorotation spectrum with an antifield rotation peak at low frequencies and a cofield rotation peak at higher ones. With increasing medium conductivity, both electrorotational peaks shift toward higher frequencies. The cofield peak becomes antifield for conductivities higher than 0.5 S/m. Because the polarizability of the external medium at these ionic strengths becomes similar to that of the cytoplasm, properties can be measured more sensitively. The critical dielectrophoretic frequencies were also determined. From our measurements, in the wide conductivity range from 2 mS/m to 1.5 S/m we propose a single-shell erythrocyte model. This pictures the cell as an oblate spheroid with a long semiaxis of 3.3 microns and an axial ratio of 1:2. Its membrane exhibits a capacitance of 0.997 x 10(-2) F/m2 and a specific conductance of 480 S/m2. The cytoplasmic parameters, a conductivity of 0.4 S/m at a dielectric constant of 212, disperse around 15 MHz to become 0.535 S/m and 50, respectively. We attribute this cytoplasmic dispersion to hemoglobin and cytoplasmic ion properties. In electrorotation measurements at about 60 MHz, an unexpectedly low rotation speed was observed. Around 180 MHz, the speed increased dramatically. By analysis of the electric chamber circuit properties, we were able to show that these effects are not due to cell polarization but are instead caused by a dramatic increase in the chamber field strength around 180 MHz. Although the chamber exhibits a resonance around 180 MHz, the harmonic content of the square-topped driving signals generates distortions of electrorotational spectra at far lower frequencies. Possible technological applications of chamber resonances are mentioned.

For human erythrocytes the bilayer-couple model predicts a reversible crenation for increasing ratios of external to internal membrane monolayer area. This can be proven experimentally by changing the ratio of lipids in the two monolayers. Under physiological conditions with no addition of membrane-active compounds, active lipid translocation and passive flip-flop are comparatively slow. Nevertheless, individual erythrocytes underwent dramatic shape changes within seconds when the suspension medium was changed. After cell suspension in low ionic strength solutions a rapid formation of echinocytes was observed. This suspension effect was also found at high ionic concentrations when Cl− was replaced by SO42−, an ion transported very slowly by band 3. The band-3 anion-exchange protein occupies about 10% of the total membrane area. Band-3 conformers are asymmetrically distributed with about 90% of the transport sites facing the inside. Specific band-3 inhibitors can induce echinocytes. These inhibitors are known to recruit the band-3 conformation with the transport site exposed to the external medium, e.g. 4,4′-diisothio-cyanatostilbene-2,2′-disulfonic acid, 4-acet-amido-4′-isothiocyanatostilbene-2,2′-disulfonic acid, flufenamic acid, furosemide and salicylic acid. Other inhibitors, e.g. phenylglyoxal and niflumic acid, which do not recruit a certain conformation, are not echinocytogenic. Physiologically, band 3 ensures the equilibration of internal and external anions like Cl− and HCO3− which are distributed according to the transmembrane potential (TMP). In the literature a correlation of the TMP and the band-3 conformation, as well as a correlation of the TMP and the erythrocyte shape, is described. In the proposed model, conformational changes may significantly alter the monolayer area ratio. Low external Cl− concentrations, negative TMPs or inhibitors may recruit the band-3 transport site towards the outside solution. It is proposed that this increases the bulk of the protein in the external monolayer and subsequently induces an echinocytogenic shape transformation according to the bilayer couple model.

Electrorotation provides information about the dielectric and conductive properties of particles over a large frequency range, which is not easily available from other techniques. Numerous applications of the dielectric particle spectroscopy technique have already been demonstrated by microscopic measurements in the field of biology. Aware of the serious limitations of microscopy for applications in colloid science, we demonstrate that dynamic light scattering can be used to measure electrorotation spectra. A suitable chamber was designed which fits into a commercial correlation spectrometer. Measurements on human red blood cells showed that the apparent diameter of the particles recorded as a function of the frequency of the rotating field matches the microscopically measured electrorotation spectrum. However, the applicability of conventional dynamic light scattering sizing-devices to the recording of electrorotation spectra is limited, since the autocorrelation function in the presence of particle rotation is a superposition of zero-order Bessel functions. Consequently, new mathematical techniques have to be developed. Another limitation is that the direction of rotation is not yet detectable. Nevertheless, since the new method overcomes the restrictions of microscopic observations it offers a potentially wide range of applications for colloidal particles, vesicles, microemulsions, subcellular particles, and cells.

Circular spatial filtering velocimetry (CSFV) was tested during the microscopic registration of the individual rotations of baker's yeast cells. Their frequency-dependent rotation (electrorotation; ER) was induced in rotating electric fields, which were generated in a glass chip chamber with four electrodes (600 μm tip-to-tip distance). The electrodes were driven with sinusoidal quadrature signals of 5 or 8 V PP with frequencies up to 3 MHz. The observed cell rotation was of the order of 1–100 s per revolution. At each measuring frequency, the independent rotations of up to 20 cells were simultaneously recorded with a high-speed camera. CSFV was software-implemented using circular spatial filters with harmonic gratings. ER was proportional to the phase shift between the values of the spatial filtering signal of consecutive frames. ER spectra obtained by CSFV from the rotation velocities at different ER-field frequencies agreed well with manual measurements and theoretical spectra. Oscillations in the rotation velocity of a single cell in the elliptically polarized field near an electrode, which were resolved by CSFV, could not be visually discerned. ER step responses after field-on were recorded at 2500 frames per second. Analysis proved the high temporal resolution of CSFV and revealed a largely linear torque-friction relation during the acceleration phase of ER. Future applications of CSFV will allow for the simple and cheap automated high-resolution analysis of rotational movements where mechanical detection has too low a resolution or is not possible, e.g. in polluted environments or for gas and fluid vortices, microscopic objects, etc.

Electrothermal micropumps (ETμPs) use local heating to create conductivity and permittivity gradients in the pump medium. In the presence of such gradients, an external AC electric field influences smeared spatial charges in the bulk of the medium. When there is also a symmetry break, the field-charge interaction results in an effective volumetric force resulting in medium pumping. The advantages of the ETμP principle are the absence of moving parts, the opportunity to passivate all the pump structures, homogeneous pump-channel cross-sections, as well as force plateaus in broad frequency ranges. The ETμPs consisted of a DC-heating element and AC field electrodes arranged in a 1000 μm × 250 μm × 60 μm (length × width × height) channel. They were processed as platinum structures on glass carriers. An equivalent-circuit diagram allowed us to model the frequency-dependent pumping velocities of passivated and nonpassivated ETμPs, which were measured at medium conductivities up to 1.0 S/m in the 300 kHz to 52 MHz frequency range. The temperature distributions within the pumps were controlled by thermochromic beads. Under resonance conditions, an additional inductance induced a tenfold pump-velocity increase to more than 50 μm/s at driving voltages of 5 V(rms). A further miniaturization of the pumps is viewed as quite feasible.