Siphon-Controlled Automation on a Lab-on-a-Disc Using Event-Triggered Dissolvable Film Valves

Within microfluidic technologies, the centrifugal microfluidic "Lab-on-a-Disc" (LoaD) platform offers great potential for use at the PoC and in low-resource settings due to its robustness and the ability to port and miniaturize \u27wet bench\u27 laboratory protocols. We present the combination of \u27event-triggered dissolvable film valves\u27 with a centrifugo-pneumatic siphon structure to enable control and timing, through changes in disc spin-speed, of the release and incubations of eight samples/reagents/wash buffers. Based on these microfluidic techniques, we integrated and automated a chemiluminescent immunoassay for detection of the CVD risk factor marker C-reactive protein displaying a limit of detection (LOD) of 44.87 ng mL$^{-1}$ and limit of quantitation (LoQ) of 135.87 ng mL$^{-1}$

Physics and Applications of a PDMS Based Centrifugal Microfluidic System

The objective of this research work is to develop a centrifugal microfluidic system for general purposes based on microfabrication technologies including SU-8 photolithography, polydimethylsiloxane (PDMS) casting. The main contribution of this research is to integrate a flyball governor system into the polymer based centrifugal microfluidic platform. A series of function units are developed based on this unique mechanism. In the first part, three pinch valve systems were designed and tested. The first one is based on the magnetic force and the second one is on the basis of spring force and the last one is a membrane valve. All valving system demonstrate good control of the fluid movement. The latter two valves are capable of sequential control. It proves that the flyball governor system is very compatible with centrifugal fluidic technologies. The major advantage of this new actuation technology is that its burst frequency can be conveniently manipulated by adjusting the parameters of the mechanical system without changes in the fluidic pattern. Next, two types of inward pumping systems were designed and tested. The result shows that both the inward pumps were capable of the pumping over a radial distance of 21mm in a short time. It thus improves the usage of space on the disc and paves the way to interconnect several functional units. Then as a proof of concept, a sequential valving system capable of metering and centrifugal sediment was developed for plasma extraction from whole blood. The resulting residual cell concentration was less than 0.5%. In the last part, a micromixer was developed based on the similar principle. The results show that the flyball governor system can effectively agitate the chaotic mixing of the sample liquids by periodically deflecting the PDMS membrane of the mixing chamber. The mixing effect can thus be enhanced

CD-based microfluidics for primary care in extreme point-of-care settings

We review the utility of centrifugal microfluidic technologies applied to point-of-care diagnosis in extremely under-resourced environments. The various challenges faced in these settings are showcased, using areas in India and Africa as examples. Measures for the ability of integrated devices to effectively address point-of-care challenges are highlighted, and centrifugal, often termed CD-based microfluidic technologies, technologies are presented as a promising platform to address these challenges. We describe the advantages of centrifugal liquid handling, as well as the ability of a standard CD player to perform a number of common laboratory tests, fulfilling the role of an integrated lab-on-a-CD. Innovative centrifugal approaches for point-of-care in extremely resource-poor settings are highlighted, including sensing and detection strategies, smart power sources and biomimetic inspiration for environmental control. The evolution of centrifugal microfluidics, along with examples of commercial and advanced prototype centrifugal microfluidic systems, is presented, illustrating the success of deployment at the point-of-care. A close fit of emerging centrifugal systems to address a critical panel of tests for under-resourced clinic settings, formulated by medical experts, is demonstrated. This emphasizes the potential of centrifugal microfluidic technologies to be applied effectively to extremely challenging point-of-care scenarios and in playing a role in improving primary care in resource-limited settings across the developing world

A centrifugal microfluidic platform for capturing, assaying and manipulation of beads and biological cells

Microfluidics is deemed a field with great opportunities, especially for applications in medical diagnostics. The vision is to miniaturize processes typically performed in a central clinical lab into small, simple to use devices - so called lab-on-a-chip (LOC) systems. A wide variety of concepts for liquid actuation have been developed, including pressure driven flow, electro-osmotic actuation or capillary driven methods. This work is based on the centrifugal platform (lab-on-a-disc). Fluid actuation is performed by the forces induced due to the rotation of the disc, thus eliminating the need for external pumps since only a spindle motor is necessary to rotate the disc and propel the liquids inside of the micro structures. Lab-on-a-disc systems are especially promising for point-of-care applications involving particles or cells due to the centrifugal force present in a rotating system. Capturing, assaying and identification of biological cells and microparticles are important operations for lab-on-a-disc platforms, and the focus of this work is to provide novel building blocks towards an integrated system for cell and particle based assays. As a main outcome of my work, a novel particle capturing and manipulation scheme on a centrifugal microfluidic platform has been developed. To capture particles (biological cells or micro-beads) I designed an array of V-shaped micro cups and characterized it. Particles sediment under stagnant flow conditions into the array where they are then mechanically trapped in spatially well-defined locations. Due to the absence of flow during the capturing process, i.e. particle sedimentation is driven by the artificial gravity field on the centrifugal platform, the capture efficiency of this approach is close to 100% which is notably higher than values reported for typical pressure driven systems. After capturing the particles, the surrounding medium can easily be exchanged to expose them to various conditions such as staining solutions or washing buffers, and thus perform assays on the captured particles. By scale matching the size of the capturing elements to the size of the particles, sharply peaked single occupancy can be achieved. Since all particles are arrayed in the same focal plane in spatially well defined locations, operations such as counting or fluorescent detection can be performed easily. The application of this platform to perform multiplexed bead-based immunoassays as well as the discrimination of various cell types based on intra cellular and membrane based markers using fluorescently tagged antibodies is demonstrated. Additionally, methods to manipulate captured particles either in batch mode or on an individual particle level have been developed and characterized. Batch release of captured particles is performed by a novel magnetic actuator which is solely controlled by the rotation frequency of the disc. Furthermore, the application of this actuator to rapidly mix liquids is shown. Manipulation of individual particles is performed using an optical tweezers setup which has been developed as part of this work. Additionally, this optical module also provides fluorescence detection capabilities. This is the first time that optical tweezers have been combined with a centrifugal microfluidic system. This work presents the core technology for an integrated centrifugal platform to perform cell and particle based assays for fundamental research as well as for point-of- care applications. The key outputs of my specific work are: 1. Design, fabrication and characterization of a novel particle capturing scheme on a centrifugal microfluidic platform (V-cups) with very high capture efficiency (close to 100%) and sharply peaked single occupancy (up to 99.7% single occupancy). 2. A novel rotation frequency controlled magnetic actuator for releasing captured particles as well as for rapidly mixing liquids has been developed, manufactured and characterized. 3. The V-cup platform has successfully been employed to capture cells and perform multi-step antibody staining assays for cell discrimination. 4. An optical tweezers setup has been built and integrated into a centrifugal teststand, and successful manipulation of individual particles trapped in the V-cup array is demonstrated

Microfluidic Technology and Application in Urinal Analysis

Microfluidic technology offers numerous advantages in minimizing and integrating the traditional assays. However, the lack of efficient control components of the microfluidic systems has been hindering the widely commercialization of the technology. The research work in this dissertation focused on the development of effective control components for microfluidic applications. A linear peristaltic pump was firstly designed, fabricated, and tested for conventional microfluidics by synchronously compressing the microfluidic channel with a miniature cam-follower system in Chapter 2. The miniature cam-follower system and microfluidic chip was prototyped using three-dimensional (3D) printing technology and soft lithography technology. Results from experimental test showed that the pump is self-priming and tolerant of bubbles. The pumping flowrate and back pressure could be controlled by changing the driving speed of the motor. Then a novel pinch-type valving system that can be used to realize both normally closed and normally open valves for centrifugal microfluidics was demonstrated in Chapter 3. A sliding wedge was actuated by centrifugal force to drive the valves. Experimental test and theoretical predication showed that the burst frequency of the valves could be tuned by changing the physical parameters of the valving system. In Chapter 4, the pinch type valving system was then further improved for better integration of multiple valves in limited space to realize sequential control of microfluidics. A valve chip with grooves on the surface was used to drive multiple valves. A flow switch which is capable of working at low rotation frequency and constant rotation direction is realized. Finally, the microfluidic platform was utilized for automatic urinalysis for the application at point of care (POC) to eliminate the difficulties in control of sample distribution and read-out time in manually conducted colorimetric urinalysis. 3D printed prototype of the microfluidic chip was used to test the proposed system. Commercial urinalysis strips was integrated with the microfluidic system for detecting glucose, specific gravity, PH, and protein from simulated urine sample. The color change of the pads was recorded using smartphone camera and analyzed to quantify the interested parameters

Advances in Microfluidics Technology for Diagnostics and Detection

Microfluidics and lab-on-a-chip have, in recent years, come to the forefront in diagnostics and detection. At point-of-care, in the emergency room, and at the hospital bed or GP clinic, lab-on-a-chip offers the potential to rapidly detect time-critical and life-threatening diseases such as sepsis and bacterial meningitis. Furthermore, portable and user-friendly diagnostic platforms can enable disease diagnostics and detection in resource-poor settings where centralised laboratory facilities may not be available. At point-of-use, microfluidics and lab-on-chip can be applied in the field to rapidly identify plant pathogens, thus reducing the need for damaging broad spectrum pesticides while also reducing food losses. Microfluidics can also be applied to the continuous monitoring of water quality and can support policy-makers and protection agencies in protecting the environment. Perhaps most excitingly, microfluidics also offers the potential to enable entirely new diagnostic tests that cannot be implemented using conventional laboratory tools. Examples of microfluidics at the frontier of new medical diagnostic tests include early detection of cancers through circulating tumour cells (CTCs) and highly sensitive genetic tests using droplet-based digital PCR.This Special Issue on “Advances in Microfluidics Technology for Diagnostics and Detection” aims to gather outstanding research and to carry out comprehensive coverage of all aspects related to microfluidics in diagnostics and detection

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

Development of novel advanced flow control systems on centrifugal microfluidic platforms for nucleic acid testing

In this work the development of novel flow control methods in centrifugal microfluidic systems for the nucleic acid testing are demonstrated. Nucleic acids make excellent biomarkers for the identification of numerous diseases, but their detection is a lengthy and labour intensive process. Centrifugal microfluidics has emerged as a highly useful tool in the area of biomedical diagnostics; however there are still limitations when it comes to sample preparation on these Lab-on-a-Disc systems. This is especially important in nucleic acid testing, where the main bottleneck in performing these processes on microfluidic devices is in sample preparation. Nucleic acid testing can be broken into three stages; extraction, purification and detection. To this end, this work outlines the development of two novel centrifugal routing systems for nucleic acid purification, through the integration of functional materials. The first is a solvent-selective router which integrated two solvent specific membrane valves. The capability of the system to purify total RNA with significant integrity and concentration was shown. The second system integrated multi-layer Graphene Oxide (GO) membranes into our Lab-on-a-Disc devices. Using this, two unique properties of the GO were investigated; its solvent selectivity and air impermeability. Finally, a novel, centrifugo-pneumatic scheme for solvent-selective routing of organic and aqueous flows was demonstrated. Also shown is the development of two separate extraction platforms. The first was a centrifugo-pneumatic ‘μHomogenizer’, which implements a 3-phase fluid extraction protocol of RNA. This system integrates chemical lysis and separation of the RNA containing aqueous phase and shows significant improvement over its time-consuming and labour intensive benchtop alternate. The second was the development of a mechanical lysis method that utilises a rotor stator grinding mill driven by the spindle motor. This system can be used for general lysis of a wide range of bacteria but would be of significant benefit for armoured cells

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

Frequency-controlled wireless passive microfluidic devices

Microfluidics is a promising technology that is increasingly attracting the attention of researchers due to its high efficiency and low-cost features. Micropumps, micromixers, and microvalves have been widely applied in various biomedical applications due to their compact size and precise dosage controllability. Nevertheless, despite the vast amount of research reported in this research area, the ability to implement these devices in portable and implantable applications is still limited. To date, such devices are constricted to the use of wires, or on-board power supplies, such as batteries. This thesis presents novel techniques that allow wireless control of passive microfluidic devices using an external radiofrequency magnetic field utilizing thermopneumatic principle. Three microfluidic devices are designed and developed to perform within the range of implantable drug-delivery devices. To demonstrate the wireless control of microfluidic devices, a wireless implantable thermopneumatic micropump is presented. Thermopneumatic pumping with a maximum flow rate of 2.86 μL/min is realized using a planar wirelessly-controlled passive inductor-capacitor heater. Then, this principle was extended in order to demonstrate the selective wireless control of multiple passive heaters. A passive wirelessly-controlled thermopneumatic zigzag micromixer is developed as a mean of a multiple drug delivery device. A maximum mixing efficiency of 96.1% is achieved by selectively activating two passive wireless planar inductor-capacitor heaters that have different resonant frequency values. To eliminate the heat associated with aforementioned wireless devices, a wireless piezoelectric normally-closed microvalve for drug delivery applications is developed. A piezoelectric diaphragm is operated wirelessly using the wireless power that is transferred from an external magnetic field. Valving is achieved with a percentage error as low as 3.11% in a 3 days long-term functionality test. The developed devices present a promising implementation of the reported wireless actuation principles in various portable and implantable biomedical applications, such as drug delivery, analytical assays, and cell lysis devices