A continuous droplet electroporation system for high throughput processing.

A continuous droplet electroporation (EP) system capable of handling a billion cells has been proposed and demonstrated using a proof-of-concept prototype design. Numerical simulations were conducted to design the new system and to compare the continuous droplet EP system with the previous single droplet EP system. Through parametric studies on the applied voltage and flow rate, a much higher cyan fluorescent protein transgene expression efficiency (38.8 ± 8.9%) was accomplished over that of the previous single droplet EP system. A parallel continuous droplet EP system is also demonstrated by introducing additional electrode pairs into the continuous droplet EP system to achieve ultrahigh throughput. Finally, the significance and meaning of the present work and future development direction have been discussed.

Digital Microfluidic Approach for Efficient Electroporation with High Productivity: Transgene Expression of Microalgae without Cell Wall Removal.

A unique digital microfluidic electroporation (EP) system successfully demonstrates higher transgene expression than that of conventional techniques, in addition to reliable productivity and feasible integrated processes. By systematic investigations into the effects of the droplet EP conditions for a wild-type microalgae, 1 order of magnitude higher transgene expression is accomplished without cell wall removal over the conventional bulk EP system. In addition, the newly proposed droplet EP method by a droplet contact charging phenomena shows a great potential for the integration of EP processes and on-chip cell culture providing easy controllability of each process. Finally, the implications of the accomplishments and future directions for development of the proposed technology are discussed.

Characterization of electrode alignment for optimal droplet charging and actuation in droplet-based microfluidic system.

The actuation method using electric force as a driving force is utilized widely in droplet-based microfluidic systems. In this work, the effects of charging electrode alignment on direct charging of a droplet on electrified electrodes and a subsequent electrophoretic control of the droplet are investigated. The charging characteristics of a droplet according to different electrode alignments are quantitatively examined through experiments and systematic numerical simulations with varying distances and angles between the two electrodes. The droplet charge acquired from the electrified electrode is directly proportional to the distance and barely affected by the angle between the two electrodes. This implies that the primary consideration of electrode alignment in microfluidic devices is the distance between electrodes and the insignificant effect of angle provides a great degree of freedom in designing such devices. Not only the droplet charge acquired from the electrode but also the force exerted on the droplet is analyzed. Finally, the implications and design guidance for microfluidic systems are discussed with an electrophoresis of a charged droplet method-based digital microfluidic device.

Interfacial tension of complex coacervated mussel adhesive protein according to the Hofmeister series.

Complex coacervation is a liquid-liquid phase separation in a colloidal system of two oppositely charged polyelectrolytes or colloids. The interfacial tension of the coacervate phase is the key parameter for micelle formation and interactions with the encapsulating material. However, the relationship between interfacial tensions and various salt solutions is poorly understood in complex coacervation. In the present work, the complex coacervate dynamics of recombinant mussel adhesive protein (MAP) with hyaluronic acid (HA) were determined in the presence of Hofmeister series salt ions. Using measurements of absorbance, hydrodynamic diameter, capillary force, and receding contact angle in the bulk phase, the interfacial tensions of complex coacervated MAP/HA were determined to be 0.236, 0.256, and 0.287 mN/m in 250 mM NaHCOO, NaCl, and NaNO3 solutions, respectively. The sequences of interfacial tensions and contact angles of the complex coacervates in the presence of three sodium salts with different anions were found to follow the Hofmeister ordering. The tendency of interfacial tension between the coacervate and dilute phases in the presence of different types of Hofmeister salt ions could provide a better understanding of Hofmeister effects on complex coacervated materials based on the protein-polysaccharide system. This information can also be utilized for microencapsulation and adsorption by controlling intramolecular interactions. In addition, the injection molding dynamics of mussel byssus formation was potentially explained based on the measured interfacial tension of coacervated MAP.

Electrowetting-induced droplet detachment from hydrophobic surfaces.

Detachment of droplets from solid surfaces is a basic and crucial process in practical applications such as heat transfer and digital microfluidics. In this study, electrowetting actuations with square pulse signals are employed to detach droplets from a hydrophobic surface. The threshold voltage for droplet detachment is obtained both experimentally and theoretically to find that it is almost constant for various droplet volumes ranging from 0.4 to 10 µL. It is also found that droplets can be detached more easily when the width of applied pulse is well-matched to the spreading time (i.e., the time to reach the maximum spread diameter). When the droplet is actuated by a double square pulse, the threshold voltage is reduced by ∼20% from that for a single square pulse actuation. Finally, by introducing an interdigitated electrode system, it is demonstrated that droplets can be detached from the solid bottom surface without using a top needle electrode.

Digital electrophoresis of charged droplets.

A digital microfluidic system based on a direct electric charging and subsequent electrophoretic manipulation of droplets is made by simple fabrication at low cost. Digitally controlled two-dimensional droplet motions are realized by digital polarity control of an array of electrodes. By independent control of droplets and colorimetric detection, the coalescence and mixing of droplets is analyzed quantitatively. The gelation of sodium alginate and the crystallization of calcium carbonate by multiple droplet translations and coalescence and the actuation of glassy carbon beads are demonstrated to show the versatile manipulation capability of the proposed technology. Finally, we discuss the implications and potentials of the present technology.

Effects of drop size and viscosity on spreading dynamics in DC electrowetting.

This study investigates the effects of drop size and viscosity on spreading dynamics, including response time, maximum velocity, and spreading pattern transition, in response to various DC voltages, based on both experiment and theoretical modeling. It is experimentally found that both switching time (i.e., time to reach maximum wetted radius) and settling time (i.e., time to reach equilibrium radius) are proportional to 1.5th power of the effective base radius. It is also found that the maximum velocity is slightly dependent on drop size but linearly proportional to the electrowetting number. The viscosity effect on drop spreading is investigated by observing spreading patterns with respect to applied voltages, and the critical viscosity at which a spreading pattern changes from under- to overdamped response is obtained. Theoretical models with contact angle hysteresis predict the spreading dynamics of drops with low and high viscosities fairly well. By fitting the theoretical models to experimental results, we obtain the friction coefficient, which is nearly proportional to 0.6th power of viscosity and is rarely influenced by applied voltage and drop size. Finally, we find that drop viscosity has a weak effect on maximum velocity but not a clear one on contact line friction.

Geometric characterization of optimal electrode designs for improved droplet charging and actuation.

Droplet charging characteristics depending on the geometry of charging electrodes have been investigated experimentally and numerically. In the experiments, two contrasting electrode systems are examined: pin-pin versus planar-planar types. To confirm the different charging behaviours on each electrode, an asymmetric system of a pin-planar type has also been examined. From the experimental and numerical results, it has been found that the droplet charge can be significantly increased (more than four times) with pin type electrodes compared with planar ones due to the increase in surface charge density by the intensification of the electric field around the charging electrode. Moreover, as the system scale becomes smaller, the superior charging effect becomes greater. Through comprehensive numerical studies on the effects of the cross-sectional area and length of a charging electrode, we have found the optimal geometric design of an electrode for droplet charging and actuation. The implications for basic understanding of the charging phenomenon and electrode design of microfluidic systems are discussed.

Discrete electrostatic charge transfer by the electrophoresis of a charged droplet in a dielectric liquid.

We have experimentally investigated the electrostatic charging of a water droplet on an electrified electrode surface to explain the detailed inductive charging processes and use them for the detection of droplet position in a lab-on-a-chip system. The periodic bouncing motion of a droplet between two planar electrodes has been examined by using a high-resolution electrometer and an image analysis method. We have found that this charging process consists of three steps. The first step is inductive charge accumulation on the opposite electrode by the charge of a droplet. This induction process occurs while the droplet approaches the electrode, and it produces an induction current signal at the electrometer. The second step is the discharging of the droplet by the accumulated induced charge at the moment of contact. For this second step, there is no charge-transfer detection at the electrometer. The third step is the charging of the neutralized droplet to a certain charged state while the droplet is in contact with the electrode. The charge transfer of the third step is detected as the pulse-type signal of an electrometer. The second and third steps occur simultaneously and rapidly. We have found that the induction current by the movement of a charged droplet can be accurately used to measure the charge of the droplet and can also be used to monitor the position of a droplet under actuation. The implications of the current findings for understanding and measuring the charging process are discussed.

Cascade reaction networks within audible sound induced transient domains in a solution.

Spatiotemporal control of chemical cascade reactions within compartmentalized domains is one of the difficult challenges to achieve. To implement such control, scientists have been working on the development of various artificial compartmentalized systems such as liposomes, vesicles, polymersomes, etc. Although a considerable amount of progress has been made in this direction, one still needs to develop alternative strategies for controlling cascade reaction networks within spatiotemporally controlled domains in a solution, which remains a non-trivial issue. Herein, we present the utilization of audible sound induced liquid vibrations for the generation of transient domains in an aqueous medium, which can be used for the control of cascade chemical reactions in a spatiotemporal fashion. This approach gives us access to highly reproducible spatiotemporal chemical gradients and patterns, in situ growth and aggregation of gold nanoparticles at predetermined locations or domains formed in a solution. Our strategy also gives us access to nanoparticle patterned hydrogels and their applications for region specific cell growth.

Electrophoresis of a charged droplet in a dielectric liquid for droplet actuation.

Electrophoretic motion of a charged droplet in a dielectric fluid under an electric field has been investigated experimentally for use as a microdroplet actuation method. The effects of the droplet size, electric field strength, and electrolyte concentration and ion species on the charging of an aqueous droplet have been examined. The amount of electrical charging has been measured by two different methods: indirect measurement using the image analysis of droplet motion and direct measurement using the electrometer. Quantitative comparison of the droplet charge measured experimentally and the theoretical value of a perfectly conductive sphere shows that an aqueous droplet is less charged than the corresponding perfectly conductive sphere. The limiting effect on electrical charging is more significant for an electrolyte droplet, and the effect is positively correlated to the electrolyte concentration rather than the ion species. This implies that the low electrical conductivity of water is not a major cause of the limiting effect. The scaling law of the charging amount for a deionized water droplet nearly follows that of the perfect conductor, whereas for an electrolyte droplet, the scaling law exponent is slightly higher. Some advantages and potentials of the current droplet actuation method are also discussed in comparison with the conventional ones.

Pumpless dispensing of a droplet by breaking up a liquid bridge formed by electric induction.

Dispensing uniform pico-to-nanoliter droplets has become one of essential components in various application fields from high-throughput bio-analysis to printing. In this study, a new method is suggested and demonstrated for dispensing a droplet on the top plate with an inverted geometry by using electric field. The process of dispensing droplets consists of two stages: (i) formation of liquid bridge by moving up the charged fluid mass using the electrostatic force between the charges on the fluid mass and the induced charges on the substrate and (ii) its break-up by the motion of the top plate. Different from conventional electrohydrodynamic methods, electric induction enables the droplets to be dispensed on various surfaces including non-conducting substrate. The use of capillarity with an inverted geometry removes the need of external pumps or elaborates control for constant flow feed. The droplet diameter has been characterized as a function of the nozzle-to-plate distance and the plate moving velocity. The robustness of the present method is shown in terms of nozzle length and applied voltage. Finally, its practical applicability is confirmed by rendering a 19 by 24 array of highly uniform droplets with only 1.8% size variation without use of any active feedback control.

Compact reaction-module on a pad for scalable flow-production of organophosphates as drug scaffolds.

Continuous pharmaceutical manufacturing receives intense attention as an alternative way to meet flexible market needs with the assurance of higher safety and quality control. Here, we report a compact reaction-module on a pad (CRP, 170 × 170 × 1.2 mm) for scale-up production of drug precursors in a continuous-flow. The CRP system was devised by stacking 9 films of the patterned polyimide to integrate micro-flow circuits, combining the functions of the even distribution of feeds, being completely mixed in less than a few milliseconds. A methodology of using a highly reactive species for the single-step synthesis of α-phosphonyloxy ketone, a drug scaffold, required the synthesis time of a few seconds in microfluidics. The fast reaction in the single CRP was capable of producing 19.2 g h-1 drug precursor, which indicates a solid step toward kilogram-scale pharmaceutical manufacturing in small footage.

Specific ion effects on electrocapillarity in aqueous electrolytes confined within nanochannels.

A nanoslit is a long, extremely narrow (nanometers apart) opening between two parallel plates. An overlapped electric double layer is formed when an electrolyte is present inside the slit and there exist distributions of the osmotic pressure and the Maxwell stress across the nanoslit, which lead to the electrocapillarity effect. This feature can be incorporated with the specific ion effects by considering the nonelectrostatic interactions between ions and confining walls, as they significantly influence the potential, electric field, and ion distributions across the nanoslit. In the present work, the electromechanical approach is integrated with the concept of specific ion effects to analyze the behavior of an electrolyte confined in a one-dimensional nanochannel. For a nanochannel, the average outward normal stress exerted on the cross section of a channel (P_{zz}[over ¯]) can be regarded as a measure of electrocapillarity and it is the driving force of the flow. This electrocapillarity measure is analyzed by using the solution of the modified Poisson-Boltzmann equation as a function of the bulk concentration of the electrolyte, the boundary potential, and most importantly, the ion-specific interfacial interactions. The significance of the present work can be manifested by the increasing usage of extremely narrow channels in nanoscaled systems, which will require proper consideration of specific ion effects in determining the behavior of the confined electrolyte.

Seismic phononic crystals by elastodynamic Navier equation.

The phenomenon, known as a complete band gap in photonic crystals consisting of periodically arranged manmade nanostructures, caused a huge sensation in photonics. Inspired by the physical methodology, we extend it to large-scale wave propagation for seismic waves. In particular, we exploit the elastodynamic Navier equation in the medium for seismic phononic crystals to induce complete band gaps of body (P) and shear (S) waves. We also show a technique that uses weak formulation to analyze band structures. Estimation of evanescent modes by complex-valued wave vectors yields propagation length, then we redesign bulky phononic crystals to be as thin as possible. We also investigate how material properties affect the relation between propagation length and effective particle velocity. This study will contribute to seismic-resistant techniques in seismology.

One-directional flow of ionic solutions along fine electrodes under an alternating current electric field.

Electric fields are widely used for controlling liquids in various research fields. To control a liquid, an alternating current (AC) electric field can offer unique advantages over a direct current (DC) electric field, such as fast and programmable flows and reduced side effects, namely the generation of gas bubbles. Here, we demonstrate one-directional flow along carbon nanotube nanowires under an AC electric field, with no additional equipment or frequency matching. This phenomenon has the following characteristics: First, the flow rates of the transported liquid were changed by altering the frequency showing Gaussian behaviour. Second, a particular frequency generated maximum liquid flow. Third, flow rates with an AC electric field (approximately nanolitre per minute) were much faster than those of a DC electric field (approximately picolitre per minute). Fourth, the flow rates could be controlled by changing the applied voltage, frequency, ion concentration of the solution and offset voltage. Our finding of microfluidic control using an AC electric field could provide a new method for controlling liquids in various research fields.

Electrical charging of a conducting water droplet in a dielectric fluid on the electrode surface.

It has been conceived that a charged droplet driven by Coulombic force can be used as a droplet-based microreactor. As a basic research for such applications, electrical charging of a conducting water droplet is studied experimentally. The effects of electric field, medium viscosity, and droplet size are investigated. It is found that the amount of electrical charging increases with the droplet size and the electric field. However, the medium viscosity does not have a significant effect in the range of the present study. A scaling law is derived from the experimental results. Unlike the case of a perfect conductor, the estimated amount of electrical charge (Q(est)) of a water droplet is proportional to the 1.59 power of the droplet radius (R) and the 1.33 power of the electric field strength (E). (For a spherical perfect conductor, Q is proportional to R(2) and E.) In order to understand these differences, numerical simulations are performed for the idealized droplets of perfect conductor. Comparison of the numerical and experimental results suggests that the differences are mainly due to incomplete charging of a water droplet resulted from the combined effect of electrochemical reaction at electrode and the relatively low conductivity of water.

Effect of ion migration in electro-generated chemiluminescence depending on the luminophore types and operating conditions.

Electro-generated chemiluminescence (ECL) has attracted increasing attention as a new platform for light-emitting devices; in particular, the use of mechanically stretchable ECL gels opens up the opportunity to achieve deformable displays. The movements of radical ions under an external electric field include short-range diffusion near the electrodes and long-distance migration between the electrodes. So far, only the diffusion of radical ions has been considered as the operating principle behind ECL. In this study, electrochemical and optical analysis was performed systematically to investigate the role of ion migration in ECL devices. This study reveals that long-distance migration of radical ions can be a key variable in ECL at low frequencies and that this effect depends on the type of ion species and the operating conditions (e.g. voltage and frequency). We also report that the emissions from the two electrodes are not identical, and the emission behaviors are different in the optimal operating conditions for the red, green, and blue ECL emissions.

Electrohydrodynamic (EHD) dispensing of nanoliter DNA droplets for microarrays.

In the present paper, we first demonstrated the possibility of electrohydrodynamic (EHD) dispensing method for preparing nanoliter probe DNA droplets on surfaces in DNA microarrays. To study the effect of an electric field on the dynamic behavior of pendent DNA droplet, visualization experiments with three kinds of electrode shapes are performed. In the early stage of droplet dispensing, it is shown that applied electric field assists a gravitational force exerted on DNA droplet. The pendent droplet is elongated in the parallel direction of applied electric field. However, after making fluid bridge between electrodes, it is shown that the electric force accelerates the capillary breaking of droplet by assisting a surface tension force exerted on droplet surface. Specifically, nanoliter dispensing volume (2 nL) is obtained in the needle-type electrode configuration. In addition, for the case of hydrophobic electrode surface, it is shown that the dispensing volume and spot size are remarkably decreased. Under the high relative humidity condition, it is observed that spot size is rapidly increased because of reduction in evaporation rate on droplet surface during the dispensing procedure. On the other hand, it is obtained that the spot size is not changed significantly in the wide range of DNA concentration from 1 to 10,000 nM. To monitor the influence of high electric voltage on DNA stability, we prepared a silicon-based chip with five capture probes for pathogens related with respiratory infectious diseases by EHD dispensing method. From the examination, it is clearly confirmed that pathogens are detected and the effective signal levels of pathogenic bacteria after hybridization are retained. Consequently, it is found that EHD dispensing method can be used to make cost-effective DNA microarrays with no thermal and electrical influences on DNA properties.

Drop formation via breakup of a liquid bridge in an AC electric field.

Experimental results are presented for the study of drop formation mechanism in a newly proposed electrohydrodynamic (EHD) method of drop generation in an AC electric field. In the method, a small drop is generated in two stages. A pendant drop is elongated with large oscillation by an electric force in the first stage. Then, it undergoes formation and breakup of a liquid bridge between the upper nozzle and the insulator-coated lower flat plate in the second stage. It is found that there exists a resonant frequency for maximum oscillation, which leads to an efficient drop formation in the latter stage. It is also found that breakup of liquid bridge is accelerated by the electrowetting tension acting on the drop perimeter contacting the insulator-coated flat plate. Thus the whole procedure of drop formation depends heavily on the frequency of AC field and the properties of the insulator such as hydrophilicity, thickness, and the dielectric constant. It is demonstrated that a wide range of drop size, from picoliter to nanoliter, can be obtained by controlling such key parameters without changing the nozzle diameter.