Engineering tissue barrier models on hydrogel microfluidic platforms

Tissue barriers play a crucial role in human physiology by establishing tissue compartmentalization and regulating organ homeostasis. At the interface between the extracellular matrix (ECM) and flowing fluids, epithelial and endothelial barriers are responsible for solute and gas exchange. In the past decade, microfluidic technologies and organ-on-chip devices became popular as in vitro models able to recapitulate these biological barriers. However, in conventional microfluidic devices, cell barriers are primarily grown on hard polymeric membranes within polydimethylsiloxane (PDMS) channels that do not mimic the cell¿ECM interactions nor allow the incorporation of other cellular compartments such as stromal tissue or vascular structures. To develop models that accurately account for the different cellular and acellular compartments of tissue barriers, researchers have integrated hydrogels into microfluidic setups for tissue barrier-on-chips, either as cell substrates inside the chip, or as self-contained devices. These biomaterials provide the soft mechanical properties of tissue barriers and allow the embedding of stromal cells. Combining hydrogels with microfluidics technology provides unique opportunities to better recreate in vitro the tissue barrier models including the cellular components and the functionality of the in vivo tissues. Such platforms have the potential of greatly improving the predictive capacities of the in vitro systems in applications such as drug development, or disease modeling. Nevertheless, their development is not without challenges in their microfabrication. In this review, we will discuss the recent advances driving the fabrication of hydrogel microfluidic platforms and their applications in multiple tissue barrier models

Microfluidic on-chip biomimicry for 3D cell culture: a fit-for-purpose investigation from the end user standpoint

A plethora of 3D and microfluidics-based culture models have been demonstrated in the recent years with the ultimate aim to facilitate predictive in vitro models for pharmaceutical development. This article summarizes to date the progress in the microfluidics-based tissue culture models, including organ-on-a-chip and vasculature-on-a-chip. Specific focus is placed on addressing the question of what kinds of 3D culture and system complexities are deemed desirable by the biological and biomedical community. This question is addressed through analysis of a research survey to evaluate the potential use of microfluidic cell culture models among the end users. Our results showed a willingness to adopt 3D culture technology among biomedical researchers, although a significant gap still exists between the desired systems and existing 3D culture options. With these results, key challenges and future directions are highlighted.This work has been supported by The Engineering and Physical Sciences Research Council (EPSRC) UK under the grant number EP/M018989/1, and a Royal Society Research Grant. Y Liu received a studentship from the Schlumberger Foundation, and E Gill received a scholarship from the WD Armstrong Trust

FABRICATION OF MAGNETIC TWO-DIMENSIONAL AND THREE-DIMENSIONAL MICROSTRUCTURES FOR MICROFLUIDICS AND MICROROBOTICS APPLICATIONS

Micro-electro-mechanical systems (MEMS) technology has had an increasing impact on industry and our society. A wide range of MEMS devices are used in every aspects of our life, from microaccelerators and microgyroscopes to microscale drug-delivery systems. The increasing complexity of microsystems demands diverse microfabrication methods and actuation strategies to realize. Currently, it is challenging for existing microfabrication methods—particularly 3D microfabrication methods—to integrate multiple materials into the same component. This is a particular challenge for some applications, such as microrobotics and microfluidics, where integration of magnetically-responsive materials would be beneficial, because it enables contact-free actuation. In addition, most existing microfabrication methods can only fabricate flat, layered geometries; the few that can fabricate real 3D microstructures are not cost efficient and cannot realize mass production. This dissertation explores two solutions to these microfabrication problems: first, a method for integrating magnetically responsive regions into microstructures using photolithography, and second, a method for creating three-dimensional freestanding microstructures using a modified micromolding technique. The first method is a facile method of producing inexpensive freestanding photopatternable polymer micromagnets composed NdFeB microparticles dispersed in SU-8 photoresist. The microfabrication process is capable of fabricating polymer micromagnets with 3 µm feature resolution and greater than 10:1 aspect ratio. This method was used to demonstrate the creation of freestanding microrobots with an encapsulated magnetic core. A magnetic control system was developed and the magnetic microrobots were moved along a desired path at an average speed of 1.7 mm/s in a fluid environment under the presence of external magnetic field. A microfabrication process using aligned mask micromolding and soft lithography was also developed for creating freestanding microstructures with true 3D geometry. Characterization of this method and resolution limits were demonstrated. The combination of these two microfabrication methods has great potential for integrating several material types into one microstructure for a variety of applications

Organ-on-a-Disc: A Scalable Platform Technology for the Generation and Cultivation of Microphysiological Tissues

Organ-on-Chip (OoC) systems culture human tissues in a controllable environment under microfluidic perfusion and enable a precise recapitulation of human physiology. Although recent studies demonstrate the potential of OoCs as alternative to traditional cell assays and animal models in drug development as well as personalized medicine, unmet challenges in device fabrication, parallelization and operation hinder their widespread application. In order to overcome these obstacles, this thesis focuses on the development of the Organ-on-a-Disc technology for the scalable generation and cultivation of microphysiological tissues. Organ-Discs are fabricated using precise, rapid and scalable microfabrication techniques. They enable the pump- and tubing-free perfusion as well as the parallelized generation and culture of tailorable and functional microtissues using rotation-based operations. The Organ-Disc setup is suitable for versatile tissue readouts, treatments and even whole blood perfusion with minimal handling and equipment requirements. Overall, the Organ-Disc creates a scalable and userfriendly platform technology for microphysiological tissue models and paves the way for their transition towards high-throughput systems.:Abbreviations Symbols 1 Introduction 2 Background 2.1 Fluid Dynamics 2.1.1 Flow Equations 2.1.2 Hydraulic Resistance 2.1.3 Wall Shear Stress 2.1.4 Centrifugal Microfluidics 2.2 Microfluidic Chip Fabrication 2.2.1 Chip Materials 2.2.2 Microstructuring 2.2.3 Bonding 3 State of the Art 3.1 Cell Culture Systems 3.2 3D Tissue Generation in Microfluidic Systems 3.3 Organ-on-Chip 3.4 Scale-up of Organ-on-Chip Systems 3.4.1 Scalable Fabrication Technologies 3.4.2 Parallelization Approaches 3.4.3 Integrated Fluid Actuation 3.5 Centrifugal Microfluidics 4 Objectives 5 Materials and Methods 5.1 Organ-Disc Fabrication 5.1.1 Materials 5.1.2 2D Structuring 5.1.3 Hot Embossing Stamp Fabrication TPE Hot Embossing 5.1.4 Bonding Solvent Vapor Bonding Thermal Fusion Bonding TPE Bonding 5.1.5 Characterization Methods Structure Sizes Bonding Strength Optical Properties 5.2 Organ-Disc Spinner 5.2.1 Centrifugal Loading Setup 5.2.2 Centrifugal Perfusion Setup 5.2.3 Peristaltic Pumping Setup 5.3 Organ-Disc Perfusion 5.3.1 Centrifugal Perfusion 5.3.2 Peristaltic Perfusion 5.4 Preparatory Cell Culture 5.5 Organ-Disc Cell Loading 5.5.1 Centrifugal Cell Loading 5.5.2 Endothelial-lining 5.6 Organ-Disc Cell Culture 5.6.1 Staining and Imaging Live Cell Labeling Live/Dead Staining CD106 Staining CD41 Staining Fixation, Permeabilization and Blocking Actin/Nuclei Staining CD31/Nuclei Staining 5.6.2 Media Analysis 5.6.3 Endothelial Cell Activation 5.6.4 Whole Blood Perfusion 5.7 Data Presentation and Statistics 6 Concept and Design 6.1 Organ-Disc Technology 6.2 Organ-Disc Design 6.3 Centrifugal Cell Loading 6.4 Endothelial Cell Lining 6.5 Centrifugal Perfusion 6.6 Peristaltic Perfusion 7 Building Blocks 7.1 Microfabrication Technology 7.1.1 Structuring 2D Structuring Hot Embossing 7.1.2 Bonding Solvent Vapor Bonding Thermal Fusion Bonding TPE Bonding 7.2 Organ-Disc Spinner 8 Perfusion 8.1 Centrifugal Pumping 8.2 Peristaltic Pumping 9 Tissue Generation and Culture 9.1 3D Tissue Generation 9.2 Stratified Tissue Construction 9.3 Generation of Endothelial-lined Channels 9.4 Perfusion of Endothelial-lined Channels 9.4.1 Media Monitoring Evaporation Cell Metabolism 9.4.2 Inflammatory Cell Stimulation 9.4.3 Whole Blood Perfusion 10 Discussion 10.1 Organ-Disc Technology 10.2 Scalable, Precise and Robust Organ-Disc Fabrication 10.2.1 Fabrication of Thermoplastic Organ-Discs 10.2.2 Fabrication of TPE Modules 10.2.3 Integration of TPE Modules to Organ-Discs 10.3 Tunable, Pump- and Tubing-free Perfusion 10.4 On-Disc Tissue Culture 10.4.1 3D Tissues 10.4.2 Blood Vessel-like Structures 10.4.3 Tissue Characterization and Treatment 10.5 On-Disc Blood Perfusion 11 Summary and Conclusion 12 References 13 AppendixIn Organ-on-Chip (OoC)-Systemen werden menschliche Gewebe mittels mikrofluidischer Versorgung in einer kontrollierten Umgebung kultiviert und so die Physiologie des Menschen nachgebildet. Obwohl aktuelle Studien zeigen, dass dieser Ansatz Alternativen zu herkömmlichen Zellbasierten Tests und Tiermodellen in der Arzneimittelentwicklung und der personalisierten Medizin bietet, stehen einer breiteren Anwendung Hürden im Bereich der Herstellung, Parallelisierung und Handhabung im Weg. Deshalb ist das Ziel dieser Arbeit die Entwicklung der Organ-on-a-Disc-Technologie, die eine skalierbare Erzeugung und Kultur von mikrophysiologischen Geweben ermöglicht. Für die Herstellung von der Organ-Disc kommen präzise, schnelle und skalierbare Mikrofabrikationsmethoden zum Einsatz. Die Organ-Disc schafft die Basis für die parallelisierte Erzeugung und Kultur von maßgeschneiderten und funktionellen Mikrogeweben, sowie deren Versorgung durch rotationsbasierte Prozesse und ohne zur Hilfenahme von Pumpen oder Schläuchen. Die Organ-Disc eignet sich für unterschiedliche Charakterisierungsmethoden sowie der Gewebestimulation und sogar der Vollblutperfusion mit minimalem Aufwand und Equipment. Insgesamt stellt die Organ-Disc eine skalierbare und benutzerfreundliche Plattformtechnologie für mikrophysiologische Modelle dar und bereitet den Weg für Hochdurchsatzanwendungen.:Abbreviations Symbols 1 Introduction 2 Background 2.1 Fluid Dynamics 2.1.1 Flow Equations 2.1.2 Hydraulic Resistance 2.1.3 Wall Shear Stress 2.1.4 Centrifugal Microfluidics 2.2 Microfluidic Chip Fabrication 2.2.1 Chip Materials 2.2.2 Microstructuring 2.2.3 Bonding 3 State of the Art 3.1 Cell Culture Systems 3.2 3D Tissue Generation in Microfluidic Systems 3.3 Organ-on-Chip 3.4 Scale-up of Organ-on-Chip Systems 3.4.1 Scalable Fabrication Technologies 3.4.2 Parallelization Approaches 3.4.3 Integrated Fluid Actuation 3.5 Centrifugal Microfluidics 4 Objectives 5 Materials and Methods 5.1 Organ-Disc Fabrication 5.1.1 Materials 5.1.2 2D Structuring 5.1.3 Hot Embossing Stamp Fabrication TPE Hot Embossing 5.1.4 Bonding Solvent Vapor Bonding Thermal Fusion Bonding TPE Bonding 5.1.5 Characterization Methods Structure Sizes Bonding Strength Optical Properties 5.2 Organ-Disc Spinner 5.2.1 Centrifugal Loading Setup 5.2.2 Centrifugal Perfusion Setup 5.2.3 Peristaltic Pumping Setup 5.3 Organ-Disc Perfusion 5.3.1 Centrifugal Perfusion 5.3.2 Peristaltic Perfusion 5.4 Preparatory Cell Culture 5.5 Organ-Disc Cell Loading 5.5.1 Centrifugal Cell Loading 5.5.2 Endothelial-lining 5.6 Organ-Disc Cell Culture 5.6.1 Staining and Imaging Live Cell Labeling Live/Dead Staining CD106 Staining CD41 Staining Fixation, Permeabilization and Blocking Actin/Nuclei Staining CD31/Nuclei Staining 5.6.2 Media Analysis 5.6.3 Endothelial Cell Activation 5.6.4 Whole Blood Perfusion 5.7 Data Presentation and Statistics 6 Concept and Design 6.1 Organ-Disc Technology 6.2 Organ-Disc Design 6.3 Centrifugal Cell Loading 6.4 Endothelial Cell Lining 6.5 Centrifugal Perfusion 6.6 Peristaltic Perfusion 7 Building Blocks 7.1 Microfabrication Technology 7.1.1 Structuring 2D Structuring Hot Embossing 7.1.2 Bonding Solvent Vapor Bonding Thermal Fusion Bonding TPE Bonding 7.2 Organ-Disc Spinner 8 Perfusion 8.1 Centrifugal Pumping 8.2 Peristaltic Pumping 9 Tissue Generation and Culture 9.1 3D Tissue Generation 9.2 Stratified Tissue Construction 9.3 Generation of Endothelial-lined Channels 9.4 Perfusion of Endothelial-lined Channels 9.4.1 Media Monitoring Evaporation Cell Metabolism 9.4.2 Inflammatory Cell Stimulation 9.4.3 Whole Blood Perfusion 10 Discussion 10.1 Organ-Disc Technology 10.2 Scalable, Precise and Robust Organ-Disc Fabrication 10.2.1 Fabrication of Thermoplastic Organ-Discs 10.2.2 Fabrication of TPE Modules 10.2.3 Integration of TPE Modules to Organ-Discs 10.3 Tunable, Pump- and Tubing-free Perfusion 10.4 On-Disc Tissue Culture 10.4.1 3D Tissues 10.4.2 Blood Vessel-like Structures 10.4.3 Tissue Characterization and Treatment 10.5 On-Disc Blood Perfusion 11 Summary and Conclusion 12 References 13 Appendi

Microfluidic device prototyping via laser processing of glass and polymer materials

In this thesis, three different processes for the fabrication of microchannels in three different base materials were experimentally and numerically modelled in detail in order to understand the effects of processing conditions on process fabrication capabilities. CO2 and Nd:YAG laser processing systems as well as a xurography technique were employed in this work for the development of microfluidic channels. The effects of CO2 laser processing on the process of directly writing microchannels on surface of four different types of glass: soda lime, fused silica, borosilicate and quartz were studied. Mathematical models were developed to relate the process input parameters to the dimensions of the microchannels. The effect of laser processing on the optical transmission capabilities of the glass was also assessed. A novel method, using Nd:YAG laser system, was employed for the fabrication of internal microchannels inside polymeric materials. Microchannels up to three millimetres long were successfully created inside a polycarbonate within a single laser processing step. Mathematical models were developed to express the relationship between laser processing input parameters and the width of these internal microchannels. The Nd:YAG processing parameters for laser welding of polycarbonate sheets were also determined. A new rapid low-cost prototyping method for the fabrication of multilayer microfluidic devices from cyclic olefin copolymer (COC) films was developed. CO2 laser cutting and xurography techniques were employed for the fabrication of the microfluidic features, followed by multilayer lamination via cyclohexane vapour exposure. Process parameters were optimised including solvent exposure time. Functional UV-transparent microfluidic mixing devices were demonstrated which included internally bound polymer monolithic columns within the microfluidic channels. There is a growing interest to use technologies which are in this thesis, the three different developed processes for the fabrication of microchannels in three different base materials provides the basis for achieving higher dimensional accuracies and novel designs within lab-on-a-chip microfluidic sensing devices

Glassy Materials Based Microdevices

Microtechnology has changed our world since the last century, when silicon microelectronics revolutionized sensor, control and communication areas, with applications extending from domotics to automotive, and from security to biomedicine. The present century, however, is also seeing an accelerating pace of innovation in glassy materials; as an example, glass-ceramics, which successfully combine the properties of an amorphous matrix with those of micro- or nano-crystals, offer a very high flexibility of design to chemists, physicists and engineers, who can conceive and implement advanced microdevices. In a very similar way, the synthesis of glassy polymers in a very wide range of chemical structures offers unprecedented potential of applications. The contemporary availability of microfabrication technologies, such as direct laser writing or 3D printing, which add to the most common processes (deposition, lithography and etching), facilitates the development of novel or advanced microdevices based on glassy materials. Biochemical and biomedical sensors, especially with the lab-on-a-chip target, are one of the most evident proofs of the success of this material platform. Other applications have also emerged in environment, food, and chemical industries. The present Special Issue of Micromachines aims at reviewing the current state-of-the-art and presenting perspectives of further development. Contributions related to the technologies, glassy materials, design and fabrication processes, characterization, and, eventually, applications are welcome

Microfluidics for Biosensing

There are 12 papers published with 8 research articles, 3 review articles and 1 perspective. The topics cover: Biomedical microfluidics Lab-on-a-chip Miniaturized systems for chemistry and life science (MicroTAS) Biosensor development and characteristics Imaging and other detection technologies Imaging and signal processing Point-of-care testing microdevices Food and water quality testing and control We hope this collection could promote the development of microfluidics and point-of-care testing (POCT) devices for biosensing

A portable artificial kidney system using microfluidics and multi-step filtration

Natural kidney filtration is a compact, multi-step filtration process which passes wastes and exceeded fluids via microscale vessels in glomerulus and tubules. The principal renal replacement therapy (RRT), commonly called dialysis, is a single-step filtration process based on diffusion to replace kidney failure. Conventional dialysis is limited in its effectiveness (not a continuous treatment), its impact on quality of life (typically requiring patients to spend several days per week in a clinic), and its cost (large systems, requiring frequent membrane replacement). This thesis is an investigation into the feasibility of using microfluidics and membrane technology to create portable alternatives to dialysis systems. It starts with a comprehensive review of the state-of-the-art in portable artificial kidneys, microfluidics, membrane science, and other related fields. An innovative, multi-step process was designed to mimic kidney filtration using two membranes; one to filter out large particles and one to remove urea and recycle water, thus mitigating the need for a dialysate system. The underlying physics (the mixing and shear stress) of the mechanisms which could enhance filtration performance at microscale was then studied. It was found that by adding microspacers into narrow-channel flows, it is possible to significantly enhance filtration. Optimized 3D-printed spacer designs (e.g., a ‘gyroid’ spacer) showed flux enhancement of up to 93% (compared to a plain channel) when using a plasma mimicking solution. The use of different blood and plasma mimicking solutions also suggested a prior step to separate large biological components (e.g., cells, proteins) is helpful to reduce cell contact and fouling in membrane filtration. The potential use of microfluidic diode valves and micropumps for pressure and flowrate regulation in the proposed small-format system was discussed. Membrane processes which mimic the filtration function of the tubules and have the potential for integration into portable systems (e.g., reverse osmosis and membrane distillation) are demonstrated to be useful potential alternatives to dialysis in toxin removal and in returning clean water to the blood stream