Concentration-adjustable micromixer using droplet injection into a microchannel

A novel micromixing technique that exploit a thrust of droplets into the mixing interface is developed. The technique enhances the mixing by injecting immiscible droplets in a mixing channel and the methodology enables a control of the mixing level simply by changing the droplet injection frequency. We experimentally characterize the mixing performance with various droplet injection frequencies, channel geometries, and diffusion coefficients. Consequently, it is revealed that the mixing level increases with the injection frequency, the droplet-diameter-to-channel-width ratio, and the diffusion coefficient. Moreover, the mixing level is found to be a linear function of the droplet volume fraction in the mixing section. The results suggest that the developed technique can produce a large amount of sample solution whose concentration is arbitrary and precisely controllable with a simple and stable operation.Comment: 12 + 3 pages, 6 + 4 figure

Droplet Microfluidics

Droplet microfluidics has dramatically developed in the past decade and has been established as a microfluidic technology that can translate into commercial products. Its rapid development and adoption have relied not only on an efficient stabilizing system (oil and surfactant), but also on a library of modules that can manipulate droplets at a high-throughput. Droplet microfluidics is a vibrant field that keeps evolving, with advances that span technology development and applications. Recent examples include innovative methods to generate droplets, to perform single-cell encapsulation, magnetic extraction, or sorting at an even higher throughput. The trend consists of improving parameters such as robustness, throughput, or ease of use. These developments rely on a firm understanding of the physics and chemistry involved in hydrodynamic flow at a small scale. Finally, droplet microfluidics has played a pivotal role in biological applications, such as single-cell genomics or high-throughput microbial screening, and chemical applications. This Special Issue will showcase all aspects of the exciting field of droplet microfluidics, including, but not limited to, technology development, applications, and open-source systems

REVERSE INSULATOR DIELECTROPHORESIS: UTILIZING DROPLET MICROENVIRONMENTS FOR DISCERNING MOLECULAR EXPRESSIONS ON CELL SURFACES

Lab-on-a-chip (LOC) technologies enable the development of portable analysis devices that use small sample and reagent volumes, allow for multiple unit operations, and couple with detectors to achieve high resolution and sensitivity, while having small footprints, low cost, short analysis times, and portability. Droplet microfluidics is a subset of LOCs with the unique benefit of enabling parallel analysis since each droplet can be utilized as an isolated microenvironment. This work explored adaptation of droplet microfluidics into a unique, previously unexplored application where the water/oil interface was harnessed to bend electric field lines within individual droplets for insulator dielectrophoretic (iDEP) characterizations. iDEP polarizes particles/cells within non-uniform electric fields shaped by insulating geometries. We termed this unique combination of droplet microfluidics and iDEP reverse insulator dielectrophoresis (riDEP). This riDEP approach has the potential to protect cell samples from unwanted sample-electrode interactions and decrease the number of required experiments for dielectrophoretic characterization by ~80% by harnessing the parallelization power of droplet microfluidics. Future research opportunities are discussed that could improve this reduction further to 93%. A microfluidic device was designed where aqueous-in-oil droplets were generated in a microchannel T-junction and packed into a microchamber. Reproducible droplets were achieved at the T-junction and were stable over long time periods in the microchamber using Krytox FSH 157 surfactant in the continuous oil FC-40 phase and isotonic salts and dextrose solutions as the dispersed aqueous phase. Surfactant, salts, and dextrose interact at the droplet interface influencing interfacial tension and droplet stability. Results provide foundational knowledge for engineering stable bio- and electro-compatible droplet microfluidic platforms. Electrodes were added to the microdevice to apply an electric field across the droplet packed chamber and explore riDEP responses. Operating windows for droplet stability were shown to depend on surfactant concentration in the oil phase and aqueous phase conductivity, where different voltage/frequency combinations resulted in either stable droplets or electrocoalescence. Experimental results provided a stability map for strategical applied electric field selection to avoid adverse droplet morphological changes while inducing riDEP. Within the microdevice, both polystyrene beads and red blood cells demonstrated weak dielectrophoretic responses, as evidenced by pearl-chain formation, confirming the preliminary feasibility of riDEP as a potential characterization technique. Two additional side projects included an alternative approach to isolate electrode surface reactions from the cell suspension via a hafnium oxide film over the electrodes. In addition, a commercially prevalent water-based polymer emulsion was found to adequately duplicate microchannel and microchamber features such that it could be used for microdevice replication

Development of functional droplet based microfluidic systems for synthetic biology and biomedical high-throughput applications

Die Tröpfchen-basierte Mikrofluidik kombiniert wissenschaftliche Prinzipen mit technologischen Ansätzen und ermöglicht ihrem Nutzer die präzise Verarbeitung und Manipulation von Wasser-in-Öl Tröpfchen. Dabei repräsentiert jedes Tröpfchen einen in sich geschlossenen Mikroreaktor, der zur Beobachtung interner chemischer und biologischer Reaktionen geeignet ist. Des Weiteren erfordert die Technologie nur minimalen Eingriff des Nutzers, ist sparsam im Probenverbrauch und ermöglicht hohe Analysegeschwindigkeiten bei erhöhter Präzision. Diese Vorteile verdeutlichen das enorme Potential dieser Technologie für die Miniaturisierung und Automatisierung biomedizinischer Tests. Trotz der in den letzten Jahren erzielten Fortschritte befindet sich die Tröpfchen-basierte Mikrofluidik immer noch im Entwicklungsstadium. Ziel meiner interdisziplinären Doktorarbeit ist es, die Tröpfchen-basierte Mikrofluidik für automatisierte Anwendungen in der biophysikalischen und biochemischen Grundlagenforschung weiterzuentwickeln. Zu diesem Zweck habe ich während meiner Promotion mehrere Chip-basierte Tröpfchenmanipulationseinheiten entwickelt und optimiert. Insbesondere wandte ich grundlegende physikalische und chemische Prinzipien an, um ihre Leistung zu verbessern. Unter anderem führten meine Entwicklungen zu einer Erhöhung der Tröpfchen-Produktionsrate, indem ich die Geometrie der Tröpfchenmanipulationseinheit modifizierte. Darüber hinaus habe ich die mikrofluidische Tröpfcheninjektionseinheit optimiert, die für die nachträgliche Manipulation des Tröpfcheninhhalts eingesetzt wird. Ich entwickelte ein neuartiges Design zur Destabilisierung der schützenden Tensidschicht ohne Notwendigkeit eines elektrischen Feldes. Die Injektion wird infolge einer schnellen Verformung des Tröpfchens und der damit verbundenen Bildung von Poren in der Tensidschicht ermöglicht. Hervorzuheben ist die Entwicklung einer Einheit zur kontrollierten Freisetzung des Tröpfcheninhalts. Durch das Anlegen eines elektrischen Feldes war es mir möglich, eingekapselte Suspensionszellen in eine kontinuierliche wässrige Phase freizusetzen und somit den Inhalt von der umgebenden Ölschicht zu trennen. Eine Kombination der entwickelten Einheit mit programmierbarer DNA-Funktionalisierung der inneren Tröpfchenfläche ermöglichte die kontrollierbare Filtration des Tröpfcheninhaltes durch kontrollierte Freisetzung der eingekapselten Materialien. Ein weiterer Fokus meiner Arbeit lag in der Entwicklung optischer Verfahren zur Echtzeitüberwachung der Wasser-in-Öl Tröpfchen. Zusammen mit Kollegen habe ich zwei entsprechende Techniken entwickelt. Eine dieser Techniken nutzt eine veränderte Auslesemethode der Fluoreszenzkorrelationsspektroskopie (FCS). Durch Neuinterpretation der Autokorrelationskurve können Aussagen über die Tröpfchenflussrate, deren Homogenität und sogar über den Tröpfcheninhalt getroffen werden. Im zweiten Ansatz wurde eine empfindliche optische Vorrichtung zur markierungsfreien Beobachtung, Charakterisierung und aktiven Manipulation vorbeifließender Tröpfchen entwickelt. Die fortschrittlichen Eigenschaften des entwickelten optischen Geräts wurden durch Messung verschiedener Tröpfchen-Produktionsparameter sowie durch den markierungsfreien Nachweis von eingekapselten Zellen bewiesen. Zusätzlich kann anhand gemessener Parameter eine aktive Manipulation der Tröpfchen durch die Vorrichtung ausgelöst werden. Dies wurde anhand einer markierungsfreien Tröpfchensortierung verdeutlicht. Zusammenfassend konnte ich die Leistung einzelner mikrofluidischer Einheiten verbessern und Anwendungsbereiche aufzeigen. Darüber hinaus verfügt das entwickelte optische Gerät über das Potential zur aktiven Überwachung und Steuerung zusammengeschalteter funktioneller Einheiten, wodurch eine gesamte Prozesskette auf einem einzigen mikrofluidischen Chip durchgeführt werden kann

An integrated droplet based microfluidic platform for high throughput, multi-parameter screening of photosensitiser activity

With rapid advances in the field of cellomics, genomics, and proteomics, the demands for development of enabling technologies for performing high throughput biological experimentation are ever increasing. Droplet based microfluidic systems have recently been developed to perform high throughput experimentations. With the ability to generate droplets over 1 kHz frequency and perform combinatorial experiments via various passive and active manipulating techniques, microdroplet technology provides an ideal platform for combinatorial biological experiments whilst consuming minimal amount of reagent. As it is possible to generate droplets, manipulate them, and characterise droplets using highly sensitive on-line detection systems, it is now crucial to bring various functionalities together to create a micro total analysis system capable of performing complex biological experiments within microfluidic devices. As such, an integrated droplet based microfluidic platform was developed to assess the efficacy of photodynamic therapy against microbial organisms. Photodynamic therapy is an alternative efficacious treatment method for the treatment of localized microbial infections with several favourable features such as broad spectrum of action, efficient inactivation of multidrug-resistant bacteria, and low mutagenic potential. In order to perform the photosensitiser cytotoxicity screening, various microfluidic modules such as droplet generation, chamber based microdroplet storage and light irradiation, droplet reinjection, electrocoalescence and on-chip viability scoring of cells within droplets using a combination of carboxyfluorescein diacetate and propidium iodide were developed and integrated within the microfluidic platform. The microfluidic system was then used to screen the cytotoxicity of TBO against E.coli cells and the results were validated against conventional colony forming unit assays. Finally, the integrated system was used to assess the effects of several parameters on E.coli viability such as dark toxicity, photosensitiser concentration, light dose and poor oxygenation condition.Open Acces

Microvortices In Droplets: Generation & Applications

The emerging field of droplet microfluidics deals with the manipulation of nL-fL droplets encapsulated within an immiscible carrier phase. The droplets are used as reaction containers for biochemical assays, enabling drastic reduction in assay volumes needed for modern life sciences research. To achieve this, basic laboratory processes such as mixing, detection, and metering must be emulated in the droplet format. Three important unit operations relevant to high throughput screening include 1) the concentration of particles and species within droplets, which is necessary for heterogeneous assays; 2) sensing the biochemical contents of a droplet; and 3) the sorting of droplets based on physical or chemical properties, which is important for single cell and proteomic assays. Currently, particle concentration in droplets requires active components, such as on-chip electrodes or magnets, along with charged or magnetic particles. Similarly, sensing and sorting droplets by chemical composition is based on flow cytometry, which also requires on-chip electrodes, feedback control, and chemical labeling. It is desirable to avoid active field techniques due to complexity, size, and cost constraints, and replace them with more simple and passive techniques. In this thesis, we utilize microvortices, the rotational motion of fluid, to enhance the capabilities of droplet microfluidics in the above three areas. The microvortices are generated using two methods: (i) hydrodynamic recirculation drag and (ii) tensiophoresis. In the first method, species concentration is accomplished by exploiting the shear-induced vortices that occur naturally inside a droplet/plug as it moves through a microchannel. Prior studies utilized these flows for enhancing mixing or interphase mass transfer. This work exploits microvortices together with two other independent phenomena--sedimentation of particles and interfacial adsorption of proteins--to concentrate both types of species at the rear of the droplet, where they can be extracted from the drop. In the latter case, the protein localization at the rear of drop reduces the interfacial tension locally resulting in an asymmetry in the drop shape. Under laminar flow, the shape deformation is deterministic and can serve as a sensitive, label-free indicator of protein concentration in proteomic screening. In the second method, label-free sorting of droplets is accomplished by a novel droplet actuation technique termed Tensiophoresis. A microchemical gradient across the droplet is transduced into a microvortex flow which propels the droplets up the chemical gradient. Using laminar flow to precisely control the gradient, droplets can be sorted by size with 3.3% resolution over a wide turning range. Droplets can be also sorted based on chemical composition because tensiophoresis is inhibited by surface active agents adsorbed on the droplet surface. Studies conducted using Bovine Serum Albumin (BSA) show that the droplet migration velocity scales inversely with protein concentration in the droplet, and migration velocity can be correlated to protein concentration with a 1 femtomole limit of detection. As modern life sciences research becomes increasingly reliant on high throughput workflows, microdroplet technology can meet the growing demand to perform screening at ultra-high throughputs with reduced sample volume. This thesis contributes three important unit operations which expand the capabilities of droplet-based workflows in proteomics, cell biology, and other areas of biomedical research

Microfluidic Production of Polymeric Functional Microparticles

This dissertation focuses on applying droplet-based microfluidics to fabricate new classes of polymeric microparticles with customized properties for various applications. The integration of microfluidic techniques with microparticle engineering allows for unprecedented control over particle size, shape, and functional properties. Specifically, three types of microparticles are discussed here: (1) Magnetic and fluorescent chitosan hydrogel microparticles and their in-situ assembly into higher-order microstructures; (2) Polydimethylsiloxane (PDMS) microbeads with phosphorescent properties for oxygen sensing; (3) Macroporous microparticles as biological immunosensors. First, we describe a microfluidic approach to generate monodisperse chitosan hydrogel microparticles that can be further connected in-situ into higher-order microstructures. Microparticles of the biopolymer chitosan are created continuously by contacting an aqueous solution of chitosan at a microfluidic T-junction with a stream of hexadecane containing a nonionic detergent, followed by downstream crosslinking of the generated droplets by a ternary flow of glutaraldehyde. Functional properties of the microparticles can be easily varied by introducing payloads such as magnetic nanoparticles and/or fluorescent dyes into the chitosan solution. We then use these prepared microparticles as "building blocks" and assemble them into high ordered microstructures, i.e. microchains with controlled geometry and flexibility. Next, we describe a new approach to produce monodisperse microbeads of PDMS using microfluidics. Using a flow-focusing configuration, a PDMS precursor solution is dispersed into microdroplets within an aqueous continuous phase. These droplets are collected and thermally cured off-chip into soft, solid microbeads. In addition, our technique allows for direct integration of payloads, such as an oxygen-sensitive porphyrin dye, into the PDMS microbeads. We then show that the resulting dye-bearing beads can function as non-invasive and real-time oxygen micro-sensors. Finally, we report a co-flow microfluidic method to prepare uniform polymer microparticles with macroporous texture, and investigate their application as discrete immunological biosensors for the detection of biological species. The matrix of such microparticles is based on macroporous polymethacrylate polymers configured with tailored pores ranging from hundreds of nanometers to a few microns. Subsequently, we immobilize bioactive antibodies on the particle surface, and demonstrate the immunological performance of these functionalized porous microbeads over a range of antigen concentrations

Non-conventional solutions to physical and engineering problems facing microfluidics

Microfluidics is a vital tool for scientific research utilising micrometre scale features to provide unparalleled control of micro-, nano- and pico-litres of fluid. Planar lithographic design and fabrication techniques have become more versatile and refined over time. However, stagnation of novel designs, fabrication methodologies and experimental conditions is increasing due to the current limitations. 3D printing is approaching the resolution required in microfluidics, whilst also providing greater freedom of design, materials, and fabrication techniques. This thesis seeks to overcome the traditional limitations using 3D printing to innovate design and production, enabling rapid prototyping methodologies and truly 3D structures, which are typically expensive and labour intensive. The first system discussed within this work generates cells-laden gelatin microdroplets and featured heating and cooling water channels, with a performance comparative to commercial devices. Secondly, a flow cell for the screening of extracellular lectins via glycomimetic liposomes was produced. Additionally, stereolithographic printing was used to produce a bioinspired monolithic droplet generator which featured intertwined channels. Finally, a Van de Graaff generator-based electrophoresis system was developed in order to generate record breaking separation resolutions whilst extended capillary lifespan compared to other experimental systems.Die Mikrofluidik ist ein wichtiges Werkzeug für die wissenschaftliche Forschung, das Merkmale im Mikrometerbereich nutzt, um eine beispiellose Kontrolle von Mikro-, Nano- und Pikolitern von Flüssigkeiten zu ermöglichen. Planare lithografische Konstruktions- und Herstellungstechniken sind im Laufe der Zeit vielseitiger und verfeinert worden. Aufgrund der derzeitigen Einschränkungen nimmt jedoch die Stagnation neuartiger Designs, Herstellungsmethoden und experimenteller Bedingungen zu. Der 3D-Druck nähert sich der in der Mikrofluidik erforderlichen Auflösung und bietet gleichzeitig eine größere Freiheit bei Design, Materialien und Herstellungstechniken. Diese Dissertation versucht, die traditionellen Einschränkungen bei der Verwendung von 3D-Druck zu überwinden, um Design und Produktion zu erneuern und schnelle Prototyping-Methoden und echte 3D-Strukturen zu ermöglichen, die normalerweise teuer und arbeitsintensiv sind. Das erste in dieser Arbeit diskutierte System erzeugt mit Zellen beladene Gelatine-Mikrotröpfchen und verfügt über Heiz- und Kühlwasserkanäle mit einer Leistung, die mit kommerziellen Geräten vergleichbar ist. Zweitens wurde eine Durchflusszelle für das Screening von extrazellulären Lektinen über glykomimetische Liposomen hergestellt. Darüber hinaus wurde stereolithografischer Druck verwendet, um einen bioinspirierten monolithischen Tröpfchengenerator herzustellen, der ineinander verschlungene Kanäle aufweist. Schließlich wurde ein auf einem Van-de-Graaff-Generator basierendes Elektrophoresesystem entwickelt, um rekordverdächtige Trennauflösungen zu erzeugen und gleichzeitig die Kapillarlebensdauer im Vergleich zu anderen experimentellen Systemen zu verlängern