Droplet microfluidics for chip-based diagnostics.

Droplet microfluidics (DMF) is a fluidic handling technology that enables precision control over dispensing and subsequent manipulation of droplets in the volume range of microliters to picoliters, on a micro-fabricated device. There are several different droplet actuation methods, all of which can generate external stimuli, to either actively or passively control the shape and positioning of fluidic droplets over patterned substrates. In this review article, we focus on the operation and utility of electro-actuation-based DMF devices, which utilize one or more micro-/nano-patterned substrates to facilitate electric field-based handling of chemical and/or biological samples. The underlying theory of DMF actuations, device fabrication methods and integration of optical and opto-electronic detectors is discussed in this review. Example applications of such electro-actuation-based DMF devices have also been included, illustrating the various actuation methods and their utility in conducting chip-based laboratory and clinical diagnostic assays.

Liquid DEP actuation and precision dispensing of variable volume droplets.

Droplet based microfluidic systems, in recent years, have demonstrated numerous advantages and exciting potential for Lab-On-a Chip applications. In order to fully realize the potential benefits of this technology, one requires precision dispensing and manipulation of droplets of known volume and sample concentration, in a rapid and controlled manner. In this article, we demonstrate the rapid and controlled microactuation of aqueous samples and subsequent dispensing of variable volume droplets in nanolitre to picolitre regime by using a coplanar tapered electrode structure that leverages the phenomena of liquid dielectrophoresis (L-DEP). The transient behavior of the tapered liquid jet departs significantly from that of a uniform liquid jet case and is not adequately explained in terms of a simplified lumped capacitance model as in the case of the uniform jet, during the L-DEP actuation. A more generalized numerical model is developed for the tapered actuation scheme to account for the experimental observations. We furthermore demonstrate that the density of the dispensed droplets can be proactively controlled by the judicious placement of electrode bumps and pinches in the electrode structure thus overcoming the limitations imposed by Rayleigh's instability criterion. The proposed droplet dispensing schemes are superior to existing L-DEP based dispensing schemes which are restricted in size and spacing of the dispensed droplets by Rayleigh's instability criteria and furthermore mostly restricted to equi-volume droplets.

Liquid dielectrophoresis and surface microfluidics.

Liquid dielectrophoresis (L-DEP), when deployed at microscopic scales on top of hydrophobic surfaces, offers novel ways of rapid and automated manipulation of very small amounts of polar aqueous samples for microfluidic applications and development of laboratory-on-a-chip devices. In this article we highlight some of the more recent developments and applications of L-DEP in handling and processing of various types of aqueous samples and reagents of biological relevance including emulsions using such microchip based surface microfluidic (SMF) devices. We highlighted the utility of these devices for on-chip bioassays including nucleic acid analysis. Furthermore, the parallel sample processing capabilities of these SMF devices together with suitable on- or off-chip detection capabilities suggest numerous applications and utility in conducting automated multiplexed assays, a capability much sought after in the high throughput diagnostic and screening assays.

Polarizability of red blood cells with an anisotropic membrane.

We predict the complex polarizability of a realistic model of a red blood cell (RBC), with an inhomogeneous dispersive and anisotropic membrane. In this model, the frequency-dependent complex electrical parameters of the individual cell layers are described by the Debye equation while the dielectric anisotropy of the cell membrane is taken into account by the different permittivities along directions normal and tangential to the membrane surface. The realistic shape of the RBC is described in terms of the Jacobi elliptic functions. To calculate the polarizability, we evoke the effective dipole moment method to determine the cell internal electric field distribution, employing an adaptive finite-element numerical approach. We have furthermore investigated the influence of the anisotropic membrane and dispersive electrical parameters of each individual cell layer on the total complex polarizability. Our findings suggest that the individual layer contribution depends on two factors: the volume of the layer and the associated induced electric field, which in turn is influenced by other layers of the cell. These results further show that the average polarizability spectra of the cell are significantly impacted by the anisotropy and associated dispersion of the cellular compartments.

DEP actuation of emulsion jets and dispensing of sub-nanoliter emulsion droplets.

Liquid Dielectrophoresis (L-DEP) has been successfully leveraged at microscopic scales and shown to provide a controllable means of on-chip precision dispensing and manipulation of sub-nanoliter single emulsion droplets. In this paper, we report on the dynamics of a DEP actuated emulsion jet prior to break-up and compare its characteristic behavior based on the lumped parameter model of Jones et al. (R. Ahmed and T. B. Jones, J. Micromech. Microeng., 2007, 17, 1052). Furthermore, features and aspects of these emulsion jets, their break-up and formation of sub-nanoliter emulsion droplets is studied in further detail. Applications of the proposed scheme in dispensing encapsulated sub-nanoliter droplets is envisioned in various fields including microTAS, on-chip handling and storage of cells and other biological samples for longer duration in controlled environments as well as solving the more general encapsulation issues in surface microfluidic devices. Scalability of the proposed scheme is shown by producing controlled sample-oil single emulsion droplets (aqueous samples in oil) in the range of 50-400 picoliters.

Microfluidic device for dielectrophoresis manipulation and electrodisruption of respiratory pathogen Bordetella pertussis.

A miniaturized microfluidic device was developed to facilitate electromanipulation of bacterial respiratory pathogens. The device comprises a microchip with circular aluminum electrodes patterned on glass, which is housed in a microfluidic system fabricated utilizing polydimethylsiloxane. The device provides sample preparation capability by exploiting positive dielectrophoresis (DEP) in conjunction with pulsed voltage for manipulation and disruption of Bordetella pertussis bacterial cells. Positive DEP capture of B. pertussis was successfully demonstrated utilizing 10 Vrms and 1 MHz ac fields. Application of dc pulses (300 V amplitude and 50 micros pulsewidth applied 1 s apart) across the aluminum electrodes resulted in electrodisruption and lysis of B. pertussis bacterial cells. Real-time polymerase chain reaction, a 2(3) factorial experimental design and transmission electron microscopy were used to evaluate bacterial cell manipulation and factors affecting bacterial cell disruption. The main factors affecting bacterial cell disruption were electric field strength, the electrical conductivity of the cell suspension sample, and the combined effect of number of pulses and sample conductivity. The bacterial deoxyribonucleic acid target remained undamaged as a result of DEP and cell lysis experimentation. Our findings suggest that a simple miniaturized microfluidic device can achieve important steps in sample preparation on-chip involving respiratory bacterial pathogens.

Leveraging liquid dielectrophoresis for microfluidic applications.

Miniaturized fluidic systems have been developed in recent years and offer new and novel means of leveraging the domain of microfluidics for the development of micro-total analysis systems (microTAS). Initially, such systems employed closed microchannels in order to facilitate chip-based biochemical assays, requiring very small quantities of sample and/or reagents and furthermore providing rapid and low-cost analysis on a compact footprint. More recently, advancements in the domain of surface microfluidics have suggested that similar low volume sample handling and manipulation capabilities for bioassays can be attained by leveraging the phenomena of liquid dielectrophoresis and droplet dielectrophoresis (DEP), without the need for separate pumps or valves. Some of the key aspects of this surface microfluidic technology and its capabilities are discussed and highlighted in this paper. We, furthermore, examine the integration and utility of liquid DEP and droplet DEP in providing rapid and automated sample handling and manipulation capabilities on a compact chip-based platform.

Ingestible capsule for impedance and pH monitoring in the esophagus.

Twenty-four-hour ambulatory pH monitoring is an essential tool for diagnosing gastroesophageal reflux disease (GERD). Simultaneous impedance and pH monitoring of the esophagus improves the detection and characterization of GERD. Conventional catheter-based monitoring systems are uncomfortable and interfere with the normal activity of the patient. To overcome these disadvantages, different wireless esophageal monitoring systems have been proposed. A capsule containing sensors for impedance and pH monitoring with wireless communication capabilities is presented. A low cost miniature microcontroller was utilized for interfacing between the sensors and a wireless transmitter. The microcontroller program allowed efficient management of the electric power provided by a 3-V battery. Magnetic holding is proposed as an alternative to surgical affixation of the monitoring capsule. Permanent neodymium magnets separated by 27 cm successfully held the capsule in a test tube. Experimental results demonstrated that friction force can aid magnetic holding to overcome peristalsis. The proposed design efficiently detected acid and nonacid reflux. More research regarding the holding method and capsule packaging are necessary to optimize the mechanical performance of the proposed design in order to facilitate clinical testing on human subjects.

Integrated microfluidic systems for sample preparation and detection of respiratory pathogen Bordetella pertussis.

An integrated microfluidic system for combined manipulation, pre-concentration, and lysis of samples containing Bordetella pertussis by dielectrophoresis and electroporation has been developed and implemented. The microfluidic device was able to pre-concentrate the amount of B. pertussis cells present in 200 microl of a B. pertussis suspension stock into a 20 microl volume. The device exhibited optimal sample pre-concentration of 6.7x at a stock value of 10(3) cfu/ml and at a flow rate of 250 microl/h. Electro-disruption experiments showed that on-chip-based electroporation is an effective solution for lysis of B. pertussis cells that is easily integrated with dielectrophoresis assisted pre-concentration procedures. Pulsed voltage applied, number of pulses, and presence of potassium chloride in a B. pertussis suspension showed a reduction in B. pertussis cell viability by electroporation; and transmission electron microscopy confirmed B. pertussis cell disruption by electroporation. Genetic amplification and detection of the pre-concentrated sample employing an integrated chip-based system demonstrated a complete chip approach for pathogen detection.

Continuous dielectrophoretic cell separation microfluidic device.

We present a prototype microfluidic device developed for the continuous dielectrophoretic (DEP) fractionation and purification of sample suspensions of biological cells. The device integrates three fully functional and distinct units consisting of an injector, a fractionation region, and two outlets. In the sheath and sample injection ports, the cell sample are hydrodynamically focused into a stream of controlled width; in the DEP fractionation region, a specially shaped nonuniform (isomotive) electric field is synthesized and employed to facilitate the separation, and the sorted cells are then delivered to two sample collection ports. The microfluidic behavior of the injector region was simulated and then experimentally verified. The operation and performance of the device was evaluated using yeast cells as model biological particles. Issues relating to the fabrication and operation of the device are discussed in detail. Such a device takes a significant step towards an integrated lab-on-a-chip device, which could interface/integrate to a number of other on-chip components for the device to undertake the whole laboratory procedure.

Electro-disruption of Escherichia coli bacterial cells on a microfabricated chip.

A miniaturized system for sample preparation of relevant bacterial pathogens has been developed using a variety of microfabrication techniques. The system manipulates and disrupts Eschericha coli bacterial cells using dielectrophoresis, electroporation and enzymes. The microchip consisted of circular gold electrodes patterned on glass using standard photolithography housed in a PDMS chamber. The bacterial lysis efficiency by electroporation and enzymatic degradation was evaluated on the microchip. The miniaturized system was capable of concentrating and aligning bacterial cells in regions of high electric field by dielectrophoresis. The miniaturized sample preparation system had a lysis efficiency of 17% when the bacterial cells were suspended in a 0.25 M sucrose solution and increased to 80% when the bacterial cells were suspended in a solution containing 0.25 M sucrose and 10 KU/ml lysozyme. Sample preparation is a limiting factor for the successful application of molecular pathogen detection methods. Therefore, the development of miniaturized systems useful for sample preparation will improve molecular detection methods of bacterial pathogens.

A combined dielectrophoresis, traveling wave dielectrophoresis and electrorotation microchip for the manipulation and characterization of human malignant cells.

The study of the dielectric properties of micrometer- or nanometer-scale particles is of particular interest in present-day applications of biomedical engineering. Electrokinetics utilises electrically energised microelectrode structures within microfluidic chambers to noninvasively probe the physiological structure of live cancer cells. A system is described that combines the three complementary techniques of dielectrophoresis (DEP), travelling wave dielectrophoresis (TWD) and electrorotation (ROT) for the first time on a single, integrated chip (3 x 6 mm). The chip employs planar microelectrode arrays fabricated on a silicon substrate to facilitate the synthesis of the various nonuniform electric fields required for the controlled manipulation, measurement and characterization of mammalian cells. A study of the dielectric properties of human malignant cells (Daudi and NCI-H929) was performed to demonstrate the potential and the versatility of the system in providing a fully programmable microsystem.

Electrorotation of axolotl embryos.

The frequency dependent dielectric properties of individual axolotl embryos (Ambystoma mexicanum) were investigated experimentally utilizing the technique of electrorotation. Individual axolotl embryos, immersed in low conductivity media, were subjected to a known frequency and fixed amplitude rotating AC electric field and the ensuing rotational motion of the embryo was monitored using a conventional optical microscope. None of the embryos in the pregastrulation or neurulation stages of development exhibited any rotational motion over the field frequency range (10 Hz-5 MHz). Over the same frequency range, the embryos in the gastrulation stage of development exhibited both co-field and counterfield rotation over different ranges of the applied field frequency. Typically, the counterfield rotation exhibited a peak in the rotation spectrum at similar 1 KHz while the co-field peak was located at similar 1-2 MHz. The rotational spectral data was analyzed using a multishelled spherical embryo model to determine the electrical character of embryos during the early development stages (Stages 5-16; i.e., 16 cell through open neural plate stages).

Theoretical Model of Electrode Polarization and AC Electroosmotic Fluid Flow in Planar Electrode Arrays.

Strong frequency-dependent fluid flow has been observed near the surface of microelectrode arrays. Modeling this phenomenon has proven to be difficult, with existing theories unable to account for the qualitative trend observed in the frequency spectra of this flow. Using recent electrode polarization results, a more comprehensive model of the double layer on the electrode surface is used to obtain good theoretical agreement with experimental data. Copyright 2001 Academic Press.