G. E. Bridges

Microwave frequency sensor for detection of biological cells in microfluidic channels

We present details of an apparatus for capacitive detection of biomaterials in microfluidic channels operating at microwave frequencies where dielectric effects due to interfacial polarization are minimal. A circuit model is presented, which can be used to adapt this detection system for use in other microfluidic applications and to identify ones where it would not be suitable. The detection system is based on a microwave coupled transmission line resonator integrated into an interferometer. At 1.5 GHz the system is capable of detecting changes in capacitance of 650 zF with a 50 Hz bandwidth. This system is well suited to the detection of biomaterials in a variety of suspending fluids, including phosphate-buffered saline. Applications involving both model particles (polystyrene microspheres) and living cells-bakers yeast (Saccharomyces cerevisiae) and Chinese hamster ovary cells-are presented.

Microfluidic electromanipulation with capacitive detection for the mechanical analysis of cells

The mechanical behavior of cells offers insight into many aspects of their properties. We propose an approach to the mechanical analysis of cells that uses a combination of electromanipulation for stimulus and capacitance for sensing. To demonstrate this approach, polystyrene spheres and yeast cells flowing in a 25 mumx100 mum microfluidic channel were detected by a perpendicular pair of gold thin film electrodes in the channel, spaced 25 mum apart. The presence of cells was detected by capacitance changes between the gold electrodes. The capacitance sensor was a resonant coaxial radio frequency cavity (2.3 GHz) coupled to the electrodes. The presence of yeast cells (Saccharomyces cerevisiae) and polystyrene spheres resulted in capacitance changes of approximately 10 and 100 attoFarad (aF), respectively, with an achieved capacitance resolution of less than 2 aF in a 30 Hz bandwidth. The resolution is better than previously reported in the literature, and the capacitance changes are in agreement with values estimated by finite element simulations. Yeast cells were trapped using dielectrophoretic forces by applying a 3 V signal at 1 MHz between the electrodes. After trapping, the cells were displaced using amplitude and frequency modulated voltages to produce modulated dielectrophoretic forces. Repetitive displacement and relaxation of these cells was observed using both capacitance and video microscopy.

Parallel pseudorandom number generation in GaAs cellular automata for high speed circuit testing

To facilitate test vector generation for high-speed circuits, we present the design and circuit simulation of parallel pseudorandom number generators in GaAs technology. These PRNGs are based on hybrid cellular automata (CA) in which mixtures of local rules are employed in one dimensional arrays, with minimal delay due to having only local wiring between neighboring cells. HSPICE simulations of these circuits demonstrate that they operate at a clock frequency above 1 GHz. Delay simulations indicate that GaAs PRNGs based upon linear feedback shift registers, in contrast with hybrid CAs, exhibit a degradation in clock frequency due to the effects of global interconnects, and that this degradation increases with the register length.

Torque-mixing magnetic resonance spectroscopy

Mechanically detected spin resonances The interaction of spins in a sample with a magnetic field can generate forces that can be sensed with cantilever probes. Losby et al. measured the resonance signals at room temperature with a micromechanical torque magnetometer. The difference between two applied radio-frequency signals corresponded to the mechanical frequency of the resonator. This approach revealed the vortex core dynamics of the ferri-toferro–magnetic transition in a micrometer-sized yttrium-iron-garnet single-crystal disk. Science, this issue p. 798 Electronic spin resonances of a magnetic single crystal are measured with a mechanical torque sensor. A universal, torque-mixing method for magnetic resonance spectroscopy is presented. In analogy to resonance detection by magnetic induction, the transverse component of a precessing dipole moment can be measured in sensitive broadband spectroscopy, here using a resonant mechanical torque sensor. Unlike induction, the torque amplitude allows equilibrium magnetic properties to be monitored simultaneously with the spin dynamics. Comprehensive electron spin resonance spectra of a single-crystal, mesoscopic yttrium iron garnet disk at room temperature reveal assisted switching between magnetization states and mode-dependent spin resonance interactions with nanoscale surface imperfections. The rich detail allows analysis of even complex three-dimensional spin textures. The flexibility of microelectromechanical and optomechanical devices combined with broad generality and capabilities of torque-mixing magnetic resonance spectroscopy offers great opportunities for development of integrated devices.

Dielectrophoresis study of temporal change in internal conductivity of single CHO cells after electroporation by pulsed electric fields

Applying sufficiently strong pulsed electric fields to a cell can permeabilize the membrane and subsequently affect its dielectric properties. In this study, we employ a microfluidic dielectrophoresis cytometry technique to simultaneously electroporate and measure the time-dependent dielectric response of single Chinese hamster ovary cells. Using experimental measurements along with numerical simulations, we present quantitative results for the changes in the cytoplasm conductivity of single cells within seconds after exposure to 100 μs duration pulsed electric fields with various intensities. It is shown that, for electroporation in a medium with conductivity lower than that of the cells cytoplasm, the internal conductivity of the cell decreases after the electroporation on a time scale of seconds and stronger pulses cause a larger and more rapid decrease. We also observe that, after the electroporation, the cells internal conductivity is constrained to a threshold. This implies that the cell prevents some of the ions in its cytoplasm from diffusing through the created pores to the external medium. The temporal change in the dielectric response of each individual cell is continuously monitored over minutes after exposure to pulsed electric fields. A time constant associated with the cells internal conductivity change is observed, which ranges from seconds to tens of seconds depending on the applied pulse intensity. This experimental observation supports the results of numerical models reported in the literature.

Dielectric properties of CHO cells obtained using a microwave interferometer based dielectrophoresis cytometer

Studying biological phenomena at the cellular level allows understanding the basics of cellular function and the mechanism of their interactions with internal or external stimuli. Among available single cell analysis techniques such as, fluorescent labeling, magnetic labeling, mechanical, and electrical approaches, electrical ones are of special interest due to their minimum influence on the natural state of cells. Dielectrophoresis (DEP) cytometry- translation of cells in a non-uniform electric field- is an electrical based technique previously employed for differentiation and characterization of cells (R. Pethig, Biomicrofluidics, 2010). In this work we employ a microwave based DEP cytometry technique that we have developed to obtain dielectric parameters of Chinese Hamster Ovary (CHO) cells, a cell line with prominent pharmaceutical applications for production of therapeutic proteins. Cell electrical parameters are not well established. We present a model for CHO using a double shell model comprising the cell membrane, cytoplasm, nuclear envelope, and nucleus.

Monitoring the dielectric response of single cells following mitochondrial adenosine triphosphate synthase inhibition by oligomycin using a dielectrophoretic cytometer.

One of the main uses of adenosine triphosphate (ATP) within mammalian cells is powering the Na(+)/K(+) ATPase pumps used to maintain ion concentrations within the cell. Since ion concentrations determine the cytoplasm conductivity, ATP concentration is expected to play a key role in controlling the cytoplasm conductivity. The two major ATP production pathways within cells are via glycolysis within the cytoplasm and via the electron transport chain within the mitochondria. In this work, a differential detector combined with dielectrophoretic (DEP) translation in a microfluidic channel was employed to observe single cell changes in the cytoplasm conductivity. The DEP response was made sensitive to changes in cytoplasm conductivity by measuring DEP response versus media conductivity and using double shell models to choose appropriate frequencies and media conductivity. Dielectric response of Chinese hamster ovary (CHO) cells was monitored following inhibition of the mitochondria ATP production by treatment with oligomycin. We show that in CHO cells following exposure to oligomycin (8 μg/ml) the cytoplasm conductivity drops, with the majority of the change occurring within 50 min. This work demonstrates that dielectric effects due to changes in ATP production can be observed at the single cell level.

All-electronic detection and actuation of single biological cells for lab-on-a-chip applications

A device sensing the change in capacitance as single cells flow through a microfluidic channel past interdigitated electrodes is described. A 0.45 aF resolution within a 3 ms time constant has been achieved. A cellpsilas electrical properties can vary greatly due to ionic currents, double layer effects, and membrane potential gradients (particularly when operating below 200 MHz). Sensing at microwave frequencies (1.6 GHz) greatly reduces these effects. Cells are simultaneously actuated by superimposing a low frequency (LF) signal, producing a dielectrophoretic (DEP) force. As the signal varies with the cellpsilas elevation, the relative magnitude and sign of the DEP force is inferred, without optical sensing.

Statistical estimation of delay fault detectabilities and fault grading

In this paper we present a technique to statistically estimate transition delay and path delay fault coverage. The basic method is an extension of STAFAN to include delay faults. By partitioning a combinational circuit into non-overlapping fanout free logic cones, we accurately calculate the transition sensitization controllabilities of 0 ⇀ 1 and 1 ⇀ 0 transitions of the lines within a fanout free logic cone to the output of the fanout free logic cone for each fanout free logic cone. A strategy to calculate the transition observabilities of fanout stems is proposed. The detectability of a path delay fault is evaluated as the product of the observabilities of the input line to its head gate within each fanout free logic cone on the path multiplied by the transition controllability of the path. When compared with the fault simulations, the estimations of transition delay fault coverage are within 2.3%. Also, the technique gives reasonably good path delay fault coverage estimation for large fault set of the ISCAS85 benchmark circuits.

The effect of dielectric relaxation in nanosecond pulse electroporation of biological cells

In order to model nanosecond pulse electroporation of the cell membrane the effect of dielectric relaxation of membrane molecules has to be considered. Since the formation of pores is a nonlinear process the dielectric relaxation effects have to be incorporated as dispersion in the time-domain. This paper presents the time-domain implementation of a second-order Debye dispersion model for a single-shell cell structure in COMSOL Multiphysics. The model is validated by comparison with frequency-domain simulation. Applying a nanosecond rise-time electric pulse to the cell shows that the transmembrane voltage increases more rapidly at the beginning of the pulse due to the dielectric relaxation effect.