A simplified PDMS microfluidic device with a built-in suction actuator for rapid production of monodisperse water-in-oil droplets

We previously established an automatic droplet-creation technique that only required air evacuation of a PDMS microfluidic device prior to use. Although the rate of droplet production with this technique was originally slow (∼10 droplets per second), this was greatly improved (∼470 droplets per second) in our recent study by remodeling the original device configuration. This improvement was realized by the addition of a degassed PDMS layer with a large surface area-to-volume ratio that served as a powerful vacuum generator. However, the incorporation of the additional PDMS layer (which was separate from the microfluidic PDMS layer itself) into the device required reversible bonding of five different layers. In the current study, we aimed to simplify the device architecture by reducing the number of constituent layers for enhancing usability of this microfluidic droplet generator while retaining its rapid production rate. The new device consisted of three layers. This comprised a degassed PDMS slab with microfluidic channels on one surface and tens of thousands of vacuum-generating micropillars on the other surface, which was simply sandwiched by PMMA layers. Despite its simplified configuration, this new device created monodisperse droplets at an even faster rate (>1000 droplets per second).     

Simultaneous generation of multiple aqueous droplets in a microfluidic device

This paper describes a microfluidic platform for the on-demand generation of multiple aqueous droplets, with varying chemical contents or chemical concentrations, for use in droplet based experiments. This generation technique was developed as a complement to existing techniques of continuous-flow (streaming) and discrete-droplet generation by enabling the formation of multiple discrete droplets simultaneously. Here sets of droplets with varying chemical contents can be generated without running the risk of cross-contamination due to the isolated nature of each supply inlet. The use of pressure pulses to generate droplets in parallel is described, and the effect of droplet size is examined in the context of flow rates and surfactant concentrations. To illustrate this technique, an array of different dye-containing droplets was generated, as well as a set of droplets that displayed a concentration gradient of a fluorescent dye.     

Recent advances in microfluidic production of Janus droplets and particles

Microfluidic production of multicompartmental emulsion droplets and particles has received considerable attention of late. In particular, droplets having two physically and chemically distinct segments (so-called Janus droplets) and the anisotropic particles synthesized from these droplets, are becoming increasingly popular because of their novel and promising properties, which make them suitable for use in numerous applications, including for controlled drug release, display devices, and self-assembly. So far, a range of interesting anisotropies have been accorded to Janus droplets and particles via microfluidics; these span from chromatic, magnetic, and hydrophobic–hydrophilic characteristics to selective degradation properties. Here, we summarize and discuss the recent trends related to Janus droplets and particles produced through microfluidic processing. We also review the parallelization technologies being developed for scaling up microfluidic emulsification in the industry.     

A microfluidic electroporation device for cell lysis

We demonstrate a micro-electroporation device for cell lysis prior to subcellular analysis. Simple circuit models show that electrical lysis method is advantageous because it is selective towards plasma membrane while leaving organelle membrane undamaged. In addition, miniaturization of this concept leads to negligible heat generation and bubble formation. The designed microdevices were fabricated using a combination of photolithography, metal-film deposition, and electroplating. We demonstrate the electro-lysis of human carcinoma cells in these devices to release the subcellular materials.     

A software-programmable microfluidic device for automated biology

Specific-purpose microfluidic devices have had considerable impact on the biological and chemical sciences, yet their use has largely remained limited to specialized laboratories. Here we present a general-purpose software-programmable microfluidic device which is capable of performing a multitude of low- and high-level functions without requiring any hardware modifications. To demonstrate the applicability and modularity of the device we implemented a variety of applications such as a microfluidic display, fluid metering and active mixing, surface immunoassays, and cell culture. We believe that analogously to personal computers, programmable, general-purpose devices will increase the accessibility and advance the pervasiveness of microfluidic technology.     

A polymer-based microfluidic device for immunosensing biochips

This paper describes the design, fabrication, and test of a PDMS/PMMA-laminated microfluidic device for an immunosensing biochip. A poly(dimethyl siloxane)(PDMS) top substrate molded by polymer casting and a poly(methyl methacrylate)(PMMA) bottom substrate fabricated by hot embossing are bonded with pressure and hermetically sealed. Two inlet ports and an air vent are opened through the PDMS top substrate, while gold electrodes for electrochemical biosensing are patterned onto the PMMA bottom substrate. The analyte sample is loaded from the sample inlet port to the detection chamber by capillary force, without any external intervening forces. For this and to control the time duration of sample fluid in each compartment of the device, including the inlet port, diffusion barrier, reaction chamber, flow-delay neck, and detection chamber, the fluid conduit has been designed with various geometries of channel width, depth, and shape. Especially, the fluid path has been designed so that the sample flow naturally stops after filling the detection chamber to allow sufficient time for biochemical reaction and subsequent washing steps. As model immunosensing tests for the microfluidic device, functionalizations of ferritin and biotin to the sensing surfaces on gold electrodes and their biospecific interactions with antiferritin antiserum and streptavidin have been investigated. An electrochemical detection method for immunosensing by biocatalyzed precipitation has been developed and applied for signal registration. With the biochip, the whole immunosensing processes could be completed within 30 min.     

A microfluidic device for performing pressure-driven separations

Microchannels in microfluidic devices are frequently chemically modified to introduce specific functional elements or operational modalities. In this work, we describe a miniaturized hydraulic pump created by coating selective channels in a glass microfluidic manifold with a polyelectrolyte multilayer (PEM) that alters the surface charge of the substrate. Pressure-driven flow is generated due to a mismatch in the electroosmotic flow (EOF) rates induced upon the application of an electric field to a tee channel junction that has one arm coated with a positively charged PEM and the other arm left uncoated in its native state. In this design, the channels that generate the hydraulic pressure are interconnected via the third arm of the tee to a field-free analysis channel for performing pressure-driven separations. We have also shown that modifications in the cross-sectional area of the channels in the pumping unit can enhance the hydrodynamic flow through the separation section of the manifold. The integrated device has been demonstrated by separating Coumarin dyes in the field-free analysis channel using open-channel liquid chromatography under pressure-driven flow conditions.     

A microfluidic device for accurate detection of hs-cTnI

This device is aimed at ensuring that the sample is uniformly and equivalently reacted with the antibody on the NC membrane in each test when the microfluidic liquid system is introduced to the chip. In this study, the developed microfluidic chip can avoid the presence of the sample and conjugate pads in the chip, while the precision of the chromatography system can be greatly improved using the same particles, NC membrane and antibody alongside the traditional strip. The results, taking the detection of cTnI as an example, revealed that the coefficient of variation (CV) is controlled within 4%, while the maximum record of the contrast chromatographic reagent strip can reach 15%. Additionally, the detection sensitivity can maintain the same order of magnitudes with that of the traditional chromatographic strip. With the results, the determination correlation of the developed microfluidic chip has been greatly improved. In addition, the CV of the chip in this study is greatly improved in comparison with that of the traditional strip. The biggest improvement lies in the mixing between the sample and the microspheres, indicating that this is a new approach to improve the CV of the traditional strip.     

Pneumatic handling of droplets on-demand on a microfluidic device for seamless processing of reaction and electrophoretic separation

Sequential operations of pre-separation reaction process by picoliter droplets and following electrophoretic separation process were realized in a single microfluidic device with pneumatic handling of liquid. The developed device consists of a fluidic chip made of PDMS, an electrode substrate, and a temperature control substrate on which thin film heater/sensor structures are fabricated. Liquid handling, including introduction of liquid samples, droplet generation, and merging of droplets, was implemented by pneumatic manipulation through microcapillary vent structures, allowing air to pass and stop liquid flow. Since the pneumatic manipulations are conducted in a fully automated manner by using a programmable air pressure control system, the user simply has to load liquid samples on each liquid port of the device. Droplets of 420 pL were generated with an accuracy of ± 2 pL by applying droplet generation pressure in the range of 40-100 kPa. As a demonstration, a binding reaction of a 15 mer ssDNA with a peptide nucleic acid oligomer used as an oligoprobe followed by denaturing electrophoresis to discriminate a single-base substitution was performed within 1.5 min. By exploiting the droplet-on-demand capability of the device, the influence of various factors, such as reaction time, mixing ratio and droplet configurations on the ssDNA-peptide nucleic acid binding reaction in the droplet-based process, was studied toward realization of a rapid detection method to discriminate rapid single-base substitution.     

A linear dilution microfluidic device for cytotoxicity assays

A two-layer polymer microfluidic device is presented which creates nine linear dilutions from two input fluid streams mixed in varying volumetric proportions. The linearity of the nine dilutions is conserved when the flow rate is held constant at 1.0 microl min(-1) (R(2) = 0.9995) and when it is varied from 0.5-16 microl min(-1) (R(2) = 0.9998). An analytical expression is presented for designing microfluidic devices with arbitrary numbers of linear dilutions. To demonstrate the efficacy of this device, primary human epidermal keratinocytes (HEK) were stained with nine dilutions of calcein, resulting in a linear spread of fluorescent intensities (R(2) = 0.94). The operating principles of the device can be scaled up to incorporate any number of linear dilutions. This scalability, coupled with an intrinsic ability to create linear dilutions under a variety of operating conditions, makes the device applicable to high throughput screening applications such as combinatorial chemistry or cytotoxicity assays.     

A magnetically active microfluidic device for chemiluminescence bioassays

Highly active horseradish peroxidase functionalized magnetic nanoparticles were prepared and packed into a microfluidic channel, producing an in-line bioreactor that enabled a sensitive chemiluminescence assay of H(2)O(2). The proposed magnetically active microfluidic device proved useful for chemiluminescence assays of biomedically interesting compounds.     

A passive microfluidic device for continuous microparticle enrichment

A passive microfluidic device is reported for continuous microparticle enrichment. The microparticle is enriched based on the inertial effect in a microchannel with contracting‐expanding structures on one side where microparticles/cells are subjected to the inertial lift force and the momentum‐change‐induced inertial force induced by highly curved streamlines. Under the combined effect of the two forces, yeast cells and microparticles of different sizes were continuously focused in the present device over a range of Reynolds numbers from 16.7 to 125. ～68% of the particle‐free liquid was separated from the sample at Re = 66.7, and ～18 μL particle‐free liquid was fast obtained within 10 s. Results also showed that the geometry of the contracting‐expanding structure significantly influenced the lateral migration of the particle. Structures with a large angle induced strong inertial effect and weak disturbance effect of vortex on the particle, both of which enhanced the microparticle enrichment in microchannel. With simple structure, small footprint (18 × 0.35 mm), easy operation and cell‐friendly property, the present device has great potential in biomedical applications, such as the enrichment of cells and the fast extraction of plasma from blood for disease diagnose and therapy.     

Water-oil core-shell droplets for electrowetting-based digital microfluidic devices

Digital microfluidics based on electrowetting-on-dielectric (EWOD) has recently emerged as one of the most promising technologies to realize integrated and highly flexible lab-on-a-chip systems. In such EWOD-based digital microfluidic devices, the aqueous droplets have traditionally been manipulated either directly in air or in an immiscible fluid such as silicone oil. However, both transporting mediums have important limitations and neither offers the flexibility required to fulfil the needs of several applications. In this paper, we report on an alternative mode of operation for EWOD-based devices in which droplets enclosed in a thin layer of oil are manipulated in air. We demonstrate the possibility to perform on-chip the fundamental fluidic operations by using such water-oil core-shell droplets and compare systematically the results with the traditional approach where the aqueous droplets are manipulated directly in air or oil. We show that the core-shell configuration combines several advantages of both the air and oil mediums. In particular, this configuration not only reduces the operation voltage of EWOD-based devices but also leads to higher transport velocities when compared with the manipulation of droplets directly in air or oil.     

A plug and play microfluidic device

Chip-to-world interface is a major issue in the field of microfluidics and its applications. We developed a plug and play microfluidic device composed of a fluid driving unit and a polymer chip containing microfluidic channels and reservoirs. The one and only connection of the device to the external world is a set of electric control lines for the driving unit. Just putting the reagents and samples onto the reservoirs, the chip can be operated for chemical or biochemical reaction and analysis. We demonstrate here that silicon-based micropumps embedded in the present device allow us to achieve flexible fluidic manipulations with minimum time delay and dead volume.     

Reactions in droplets in microfluidic channels

Fundamental and applied research in chemistry and biology benefits from opportunities provided by droplet-based microfluidic systems. These systems enable the miniaturization of reactions by compartmentalizing reactions in droplets of femoliter to microliter volumes. Compartmentalization in droplets provides rapid mixing of reagents, control of the timing of reactions on timescales from milliseconds to months, control of interfacial properties, and the ability to synthesize and transport solid reagents and products. Droplet-based microfluidics can help to enhance and accelerate chemical and biochemical screening, protein crystallization, enzymatic kinetics, and assays. Moreover, the control provided by droplets in microfluidic devices can lead to new scientific methods and insights.     

Single-molecule emulsion PCR in microfluidic droplets

The application of microfluidic droplet PCR for single-molecule amplification and analysis has recently been extensively studied. Microfluidic droplet technology has the advantages of compartmentalizing reactions into discrete volumes, performing highly parallel reactions in monodisperse droplets, reducing cross-contamination between droplets, eliminating PCR bias and nonspecific amplification, as well as enabling fast amplification with rapid thermocycling. Here, we have reviewed the important technical breakthroughs of microfluidic droplet PCR in the past five years and their applications to single-molecule amplification and analysis, such as high-throughput screening, next generation DNA sequencing, and quantitative detection of rare mutations. Although the utilization of microfluidic droplet single-molecule PCR is still in the early stages, its great potential has already been demonstrated and will provide novel solutions to today's biomedical engineering challenges in single-molecule amplification and analysis.     

A parallel diffusion-based microfluidic device for bacterial chemotaxis analysis

We developed a multiple-channel microfluidic device for bacterial chemotaxis detection. Some characteristics such as easy operation, parallel sample adding design and fast result readout make this device convenient for most biology labs. The characteristic feature of the design is the agarose gel channels, which serve as a semi-permeable membrane. They can stop the fluid flow and prevent bacteria getting across, but permit the diffusion of small molecules. In the device fabrication process a novel thermal-based method was used to control the shape of agarose gel in the microfluidic channel. The chemical gradient is established by diffusion which can be precisely controlled and measured. Combined with an 8-channel pipette, different attractants, repellent chemicals or different bacteria were analyzed by a two step operation with a readout time of one hour. This device may be useful in the high throughput detection of chemotaxis related molecules and genes.     

A microfluidic electrochemiluminescent device for detecting cancer biomarker proteins

We describe an electrochemiluminescence (ECL) immunoarray incorporated into a prototype microfluidic device for highly sensitive protein detection and apply this system to accurate, sensitive measurements of prostate-specific antigen (PSA) and interleukin-6 (IL-6) in serum. The microfluidic system employed three molded polydimethylsiloxane (PDMS) channels on a conductive pyrolytic graphite chip (2.5?×?2.5 cm) inserted into a machined chamber and interfaced with a pump, switching valve, and sample injector. Each of the three PDMS channels encompasses three 3 μL analytical wells. Capture-antibody-decorated single-wall carbon nanotube forests are fabricated in the bottom of the wells. The antigen is captured by these antibodies on the well bottoms. Then, a RuBPY-silica-secondary antibody (Ab₂) label is injected to bind to antigen on the array, followed by injection of sacrificial reductant tripropylamine (TPrA) to produce ECL. For detection, the chip is placed into an open-top ECL measuring cell, and the channels are in contact with electrolyte in the chamber. Potential applied at 0.95 V versus Ag/AgCl oxidizes TPrA to produce ECL by redox cycling the RuBPY species in the particles, and ECL light is measured by a charge-coupled device camera. This approach achieved ultralow detection limits of 100 fg?mL^?1 for PSA (9 zeptomole) and 10 fg?mL^?1 (1 zeptomole) for IL-6 in calf serum, a 10–25-fold improvement of a similar non-microfluidic array. PSA and IL-6 in synthetic cancer patient serum samples were detected in 1.1 h and results correlated well with single-protein enzyme-linked immunosorbent assays.     

A microfluidic device for high throughput bacterial biofilm studies

Bacteria are almost always found in ecological niches as matrix-encased, surface-associated, multi-species communities known as biofilms. It is well established that soluble chemical signals produced by the bacteria influence the organization and structure of the biofilm; therefore, there is significant interest in understanding how different chemical signals are coordinately utilized for community development. Conventional methods for investigating biofilm formation such as macro-scale flow cells are low-throughput, require large volumes, and do not allow spatial and temporal control of biofilm community formation. Here, we describe the development of a PDMS-based two-layer microfluidic flow cell (μFC) device for investigating bacterial biofilm formation and organization in response to different concentrations of soluble signals. The μFC device contains eight separate microchambers for cultivating biofilms exposed to eight different concentrations of signals through a single diffusive mixing-based concentration gradient generator. The presence of pneumatic valves and a separate cell seeding port that is independent from gradient-mixing channels offers complete isolation of the biofilm microchamber from the gradient mixer, and also performs well under continuous, batch or semi-batch conditions. We demonstrate the utility of the μFC by studying the effect of different concentrations of indole-like biofilm signals (7-hydroxyindole and isatin), either individually or in combination, on biofilm development of pathogenic E. coli. This model can be used for developing a fundamental understanding of events leading to bacterial attachment to surfaces that are important in infections and chemicals that influence the biofilm formation or inhibition.