3D Cell Culture Method in Channel-Free Water-in-Oil Droplets.

A new channel-free water-in-oil (WO) droplet 3D cell culture method is proposed to address the challenges while maintaining the advantages of the conventional 3D cell culture methods. The proposed WO method can fundamentally solve the constraint of spheroids size, a common challenge in conventional 3D culture, by using droplet size controllability. The 3D cell culture performance of the WO method is verified by comparing it with the conventional 3D cell culture methods.

Simultaneous Separating, Splitting, Collecting, and Dispensing by Droplet Pinch-Off for Droplet Cell Culture.

Simultaneous separating, splitting, collecting, and dispensing a cell suspension droplet has been demonstrated by aspiration and subsequent droplet pinch-off for use in microfluidic droplet cell culture systems. This method is applied to cell manipulations including aliquots and concentrations of microalgal and mammalian cell suspensions. Especially, medium exchange of spheroid droplets is successfully demonstrated by collecting more than 99% of all culture medium without damaging the spheroids, demonstrating its potential for a 3D cell culture system.

Safe and efficient RNA and DNA introduction into cells using digital electroporation system.

We systematically compared the delivery and expression efficiencies according to cell types (plant and animal cells) and genetic materials (RNA and DNA) to deliver RNA using a digital electroporation system. Despite the significantly lower RNA delivery in Chlamydomoans reinhartii than DNA delivery due to RNA secondary structure and cell wall, the expression/delivery ratio of RNA was significantly higher than that of DNA (up to 90%), confirming the generally known fact that RNA is more favorable for expression than DNA. On the other hand, in K562 cells, the difference in RNA and DNA delivery efficiency was negligible.

Effect of Deformation on Droplet Contact Charge Electrophoresis.

The effect of deformation on the droplet contact charge electrophoresis (CCEP) was investigated for consistent droplet movement control. Through systematic experiments and numerical simulations, it has been found that overcharging by deformation is up to about 130% of the sphere and is mainly driven by the concentration of the electric field near the tip of the droplet rather than an increase in the surface area. Dimensional analysis revealed a consistent droplet CCEP motion with the electric capillary number range of 0.

Wall Effects on Hydrodynamic Drag and the Corresponding Accuracy of Charge Measurement in Droplet Contact Charge Electrophoresis.

Droplet size dependent wall effects on hydrodynamic drag and the corresponding droplet contact charge estimations were experimentally and theoretically investigated. The consistent reduction in the dimensionless droplet contact charges proportional to droplet size was reported and explained by the parallel and approaching wall effects on the drag coefficient. Extrapolation of the size dependent droplet charge data showed that the droplet charge follows the perfect conductor theory when the droplet radius approaches zero.

Enhanced Controllability of Fries Rearrangements Using High-Resolution 3D-Printed Metal Microreactor with Circular Channel.

High-resolution 3D-printed stainless steel metal microreactors (3D-PMRs) with different cross-sectional geometry are fabricated to control ultrafast intramolecular rearrangement reactions in a comparative manner. The 3D-PMR with circular channel demonstrates the improved controllability in rapid Fries-type rearrangement reactions, because of the superior mixing efficiency to rectangular cross-section channels (250 µm × 125 µm) which is confirmed based on the computational flow dynamics simulation. Even in case of very rapid intramolecular rearrangement of sterically small acetyl group occurring in 333 µs of reaction time, the desired intermolecular reaction can outpace to the undesired intramolecular rearrangement using 3D-PMR to result in high conversion and yield.

Correction: Indirect fabrication of versatile 3D microfluidic device by a rotating plate combined 3D printing system.

[This corrects the article DOI: 10.1039/C8RA08465C.].

Optimization of the droplet electroporation method for wild type Chlamydomonas reinhardtii transformation.

We performed the transformation of a wild type Chlamydomonas reinhardtii by optimizing previously developed droplet EP method. For more effective and faster optimization, we used DNA dying fluorescent molecule (Yo-Pro-1) for finding optimal EP conditions instead of using protein expression based evaluation method. By examining wider range of electrical parameter space together with the analysis of total current flow of EP process, we found optimal EP conditions.

Indirect fabrication of versatile 3D microfluidic device by a rotating plate combined 3D printing system.

In the past decade, 3D-printing technology has been applied in the field of microfluidics to fabricate microfluidic devices for wide-ranging areas of study including chemistry, biology, medicine, and others. However, these methods face several limitations such as insufficient resolution and long fabrication time. In this study, versatile microfluidic devices with different functions were indirectly fabricated by a rapid sacrificial template printing process using a photocurable fluoropolymer with chemical durability.

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.

Electrostatic Origins of the Positive and Negative Charging Difference in the Contact Charge Electrophoresis of a Water Droplet.

The positive and negative charging difference in the contact charge electrophoresis of a water droplet suspended in oil was investigated to find out the origin of this charging difference. Through numerous experiments and numerical analysis, the charging difference has been found to be mainly originated from electrostatic sources. Two electrostatic sources were found in the present experimental setup, and by excluding those two sources the charging difference was successfully diminished.

Electrically Controllable Microparticle Synthesis and Digital Microfluidic Manipulation by Electric-Field-Induced Droplet Dispensing into Immiscible Fluids.

The dispensing of tiny droplets is a basic and crucial process in a myriad of applications, such as DNA/protein microarray, cell cultures, chemical synthesis of microparticles, and digital microfluidics. This work systematically demonstrates droplet dispensing into immiscible fluids through electric charge concentration (ECC) method. It exhibits three main modes (i.

Submillisecond organic synthesis: Outpacing Fries rearrangement through microfluidic rapid mixing.

In chemical synthesis, rapid intramolecular rearrangements often foil attempts at site-selective bimolecular functionalization. We developed a microfluidic technique that outpaces the very rapid anionic Fries rearrangement to chemoselectively functionalize iodophenyl carbamates at the ortho position. Central to the technique is a chip microreactor of our design, which can deliver a reaction time in the submillisecond range even at cryogenic temperatures.

Multilayered film microreactors fabricated by a one-step thermal bonding technique with high reproducibility and their applications.

We report the versatile uses of multilayered polyimide (PI) film microreactors with various functions including pressure tolerance, three-dimensional mixing and multistep membrane emulsification. Such PI film microreactors were fabricated by a simple one-step thermal bonding technique with high reproducibility. Upon bonding at 300 °C for 1 hour, the thin and flexible film microdevices could withstand pressure up to 8.

Extraction of Cations from an Ionic Liquid Droplet in a Dielectric Liquid under Electric Field.

Ionic liquids show great promise as excellent solvents or catalysts in energy and biological fields due to their unique chemical and physical properties. In this work, the characteristics of various ionic liquids are investigated with the electrophoresis of a charged droplet (ECD) method. Under normal situation, a charged droplet in a dielectric liquid shows back-and-forth bouncing motion between the positive and negative electrodes continuously.

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.

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.

A monolithic and flexible fluoropolymer film microreactor for organic synthesis applications.

A photocurable and viscous fluoropolymer with chemical stability is a highly desirable material for fabrication of microchemical devices. Lack of a reliable fabrication method, however, limits actual applications for organic reactions. Herein, we report fabrication of a monolithic and flexible fluoropolymer film microreactor and its use as a new microfluidic platform.

Three-dimensional flash flow microreactor for scale-up production of monodisperse PEG-PLGA nanoparticles.

We present a pressure-tolerant 3D parallel polyimide (PI) film microreactor operating at up to ~160 bars with direct 3D flow focusing geometry for mass production of PEG-PLGA nanoparticles in a ~10(1) gram-scale (g h(-1)).

Chitosan-microreactor: a versatile approach for heterogeneous organic synthesis in microfluidics.

Microreactors have been proven to be efficient tools for a variety of homogeneous organic transformations due to their mixing efficiency, which results in very fast reactions, better heat and mass transfer, and simple scale-up. However, in heterogeneous catalytic reactions each catalyst needs an individual substrate as support. Herein, a versatile approach to immobilize metal catalysts on chitosan as a common substrate is presented.

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.

Spontaneous electrical charging of droplets by conventional pipetting.

We report that a droplet dispensed from a micropipette almost always has a considerable electrical charge of a magnitude dependent on the constituents of the droplet, on atmospheric humidity and on the coating material of pipette tip. We show that this natural electrification of a droplet originates from the charge separation between a droplet and pipette tip surface by contact with water due to the ionization of surface chemical groups. Charge on a droplet can make it difficult to detach the droplet from the pipette tip, can decrease its surface tension, can affect the chemical characteristics of solutions due to interactions with charged molecules, and can influence the combination and localization of charged bio-molecules; in all cases, the charge may affect results of experiments in which any of these factors is important.

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.

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.