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$^{-1}$

Design and fabrication of a low-cost wireless camera imaging system for centrifugal microfluidics

Centrifugal microfluidic devices offer a robust method for low-volume fluid handling by combining low-cost instrumentation with highly integrated automation. Crucial to the efficacy of Lab-on-a-Disc (LoaD) device operation is the selection of robust valving technology, the design of on-disc fluidic structures, and accurate control of disc spin-speeds (centrifugal force) during operation. The design and refinement of fluidic and valving structures is often guided by inspecting disc operation using high-speed camera systems. This approach involves synchronising image acquisition with disc rotation to visualise liquid flow through a series of images often presented in a video format. Depending on the decisions taken, such systems can cost from €4,000 upwards. This paper outlines the development of a low-cost centrifugal test-stand with an integrated imaging system using a generic wireless camera to record videos directly to a smartphone device. This imaging system can be fabricated using only 3D printers and a low-cost CNC milling machine from widely available materials for approximately €350. High-fidelity imaging of the entire disc for flow visualisation and the recording of real-time colour intensity measurements are facilitated by this standalone device. A vibration analysis study has been performed to determine the rotational velocity range at which the system can be safely operated. Furthermore, the efficacy of the imaging system has been demonstrated by performing real-time colour intensity measurements of dyed water dilution

3D printed microfluidic device for point-of-care anemia diagnosis

Master of ScienceDepartment of Biological & Agricultural EngineeringMei HeAnemia affects about 25% of the world’s population and causes roughly 8% of all disability cases. The development of an affordable point-of-care (POC) device for detecting anemia could be a significant for individuals in underdeveloped countries trying to manage their anemia. The objective of this study was to design and fabricate a 3D printed, low cost microfluidic mixing chip that could be used for the diagnosis of anemia. Microfluidic mixing chips use capillary flow to move fluids without the aid of external power. With new developments in 3D printing technology, microfluidic devices can be fabricated quickly and inexpensively. This study designed and demonstrated a passive microfluidic mixing chip that used capillary force to mix blood and a hemoglobin detecting assay. A 3D computational fluid dynamic simulation model of the chip design showed 96% efficiency when mixing two fluids. The mixing chip was fabricated using a desktop 3D printer in one hour for less than $0.50. Blood samples used for the clinical validation were provided by The University of Kansas Medical Center Biospecimen Repository. During clinical validation, RGB (red, green, blue) values of the hemoglobin detection assay color change within the chip showed consistent and repeatable results, indicating the chip design works efficiently as a passive mixing device. The anemia detection assay tended to overestimate hemoglobin levels at lower values while underestimating them in higher values, showing the assay needs to go through more troubleshooting

Development of a laser foaming process for high throughput three-dimensional tissue model devices

A three-dimensional (3D) porous structure on biodegradable or biocompatible polymers have attracted tremendous attention in numerous bio-related areas including 3D cell culturing and tissue engineering because of their microenvironment similar to ones in vivo. In this study, a novel fabrication process, named selective laser foaming, was developed to create localized 3D porous structure on a polymer chip. The effects of laser power and lasing time on the porous structure were studied both experimentally and through finite element modeling. A high throughput two-chamber tissue model platform was developed using the proposed selective laser foaming process. For comparison, cell culture studies were conducted with both selective laser foamed and unfoamed polylactic acid (PLA) samples using T98G cells. The results show that by laser foaming gas-impregnated polylactic acid it is possible to generate an array of inverse cone-shaped wells with porous walls. The size of the foamed region can be controlled with laser power and exposure time, while the pore size of the scaffold can be manipulated with the saturation pressure. The finite element modeling results showed good agreement with the experimental data; therefore, the model could be used to optimize and control the selective foaming process. T98G cells grew well in the foamed scaffolds, forming clusters that have not been observed in 2D cell cultures. Cells were more viable in the 3D scaffolds than in the 2D cell culture cases, suggesting that the 3D porous microarray could be used for parallel studies of drug toxicity, guided stem cell differentiation, and DNA binding profiles. As an example, a high-throughput two-chamber 3D tissue model platform driven by the centrifugal force was developed for drug screening. The selective laser foaming process was calibrated to fabricate 3D scaffold on a commercially available compact disc (CD) made of polycarbonate (PC). Laser foaming of gas saturated polycarbonate created inverse cone-shaped wells with micro-sized porous structure underneath the surface. The open pores were hundreds of micrometers in diameter and depth. The pore size of the underneath porous structure was several tens of micrometers. The size of the well was dependent on the laser power and laser exposure time. Two laser-foamed scaffolds were fabricated in tandem and two mechanically-machined chambers were placed adjacent to the scaffolds, respectively. All scaffolds and chambers were in line and all of them were connected with micro-channels. The surface was coated with polydopamine (PDA) in order to increase the hydrophilicity and biocompatibility. After sterilization, human glioblastoma multiforme (M059K) and hepatoblastoma (C3A sub28) were seeded in the two 3D scaffolds separately and cultured for up to four weeks. These cells grew well in the scaffolds and cell aggregates were observed, suggesting that the developed two-chamber tissue model array could be useful for high-throughput biochemical assays.Mechanical Engineerin

Size based platelet isolation on a centrifugal microfluidic device

Department of Biomedical EngineeringCardiovascular diseases are one of the leading causes of disability and mortality worldwide and directly associated with the enhanced reactivity of platelets. Platelet is the smallest (1 ~ 4 ??m) cells in the circulation, comprising the second largest volume fraction of blood, responsible for the maintenance of the circulatory systems in normal condition. However, platelets can significantly contribute to the formation of thrombus which blocks the blood flow if hyper-functional, or result in bleeding, if dysfunctional. Aside from its hemostatic role, platelets are also involved in other essential and versatile functions of immunity, wound healing, and inflammation. Its pathophysiological involvements in cancer, Alzheimer???s disease, cardiovascular disease, diabetes and viral infections were recently established and have gained much attention for its potential use in both diagnostics and therapy. Conventional platelet isolation uses density-based centrifugation, which lacks global standard, hence, high variations that influence clinical decision have been reported. In addition, platelet activation due to high shear stress during the centrifugation, and WBC contamination limited the use of platelet in bio-assays for protein quantification and RNA analysis. The conventional isolation approach also suffers from handling errors, long processing time, and labour intensiveness. Hence, microfluidics based technological interventions have been developed to overcome above limitations but was not able to achieve the optimal platelet isolation of uncompromised high purity, throughput, and recovery with minimal activation in short time from undiluted whole blood. Still, isolation of platelets for molecular diagnosis in small blood volume remains a challenging task. Therefore, we developed a fully automated lab-on-a-disc device to isolate platelets for downstream analysis. By integration of sequential filtration on disc with 3 ??m and 600 nm pore size membranes, highly pure platelet isolation was achieved. From our results, the disc based isolation significantly increased platelet count by 3~4 fold, while simultaneously lowering activation even in the absence of inhibitors. The flow cytometry and RT-PCR analysis of isolated platelets revealed that our disc platform results in ~ 99 % pure platelets free from WBC contamination having WBC specific gene undetected. In summary, the experimental result confirmed that disc is capable of separating platelets ideal for downstream analysis having high purity and recovery with minimal activation in time efficient manner. Prior to downstream analysis, platelet function test for screening of multiple platelet related disorders or conditions are commonly requested in clinical settings. As a proof of concept, we demonstrated the potential of full integration of light transmission aggregometry - the reference gold standard of addressing platelet function - on lab on a disc platform as a point of care testing device to improve time efficiency, overall cost, and higher precision without restriction to the staff and facilities.ope

Ultrafast Microfluidic Immunoassays Towards Real-time Intervention of Cytokine Storms

Biomarker-guided precision medicine holds great promise to provide personalized therapy with a good understanding of the molecular or cellular data of an individual patient. However, implementing this approach in critical care uniquely faces enormous challenges as it requires obtaining “real-time” data with high sensitivity, reliability, and multiplex capacity near the patient’s bedside in the quickly evolving illness. Current immunodiagnostic platforms generally compromise assay sensitivity and specificity for speed or face significantly increased complexity and cost for highly multiplexed detection with low sample volume. This thesis introduces two novel ultrafast immunoassay platforms: one is a machine learning-based digital molecular counting assay, and the other is a label-free nano-plasmonic sensor integrated with an electrokinetic mixer. Both of them incorporate microfluidic approaches to pave the way for near-real-time interventions of cytokine storms. In the first part of the thesis, we present an innovative concept and the theoretical study that enables ultrafast measurement of multiple protein biomarkers (<1 min assay incubation) with comparable sensitivity to the gold standard ELISA method. The approach, which we term “pre-equilibrium digital enzyme-linked immunosorbent assay” (PEdELISA) incorporates the single-molecular counting of proteins at the early, pre-equilibrium state to achieve the combination of high speed and sensitivity. We experimentally demonstrated the assay’s application in near-real-time monitoring of patients receiving chimeric antigen receptor (CAR) T-cell therapy and for longitudinal serum cytokine measurements in a mouse sepsis model. In the second part, we report the further development of a machine learning-based PEdELISA microarray data analysis approach with a significantly extended multiplex capacity using the spatial-spectral microfluidic encoding technique. This unique approach, together with a convolutional neural network-based image analysis algorithm, remarkably reduced errors faced by the highly multiplexed digital immunoassay at low analyte concentrations. As a result, we demonstrated the longitudinal data collection of 14 serum cytokines in human patients receiving CAR-T cell therapy at concentrations < 10pg/mL with a sample volume < 10 µL and 5-min assay incubation. In the third part, we demonstrate the clinical application of a machine learning-based digital protein microarray platform for rapid multiplex quantification of cytokines from critically ill COVID-19 patients admitted to the intensive care unit. The platform comprises two low-cost modules: (i) a semi-automated fluidic dispensing module that can be operated inside a biosafety cabinet to minimize the exposure of technician to the virus infection and (ii) a compact fluorescence optical scanner for the potential near-bedside readout. The automated system has achieved high interassay precision (~10% CV) with high sensitivity (<0.4pg/mL). Our data revealed large subject-to-subject variability in patient responses to anti-inflammatory treatment for COVID-19, reaffirming the need for a personalized strategy guided by rapid cytokine assays. Lastly, an AC electroosmosis-enhanced localized surface plasmon resonance (ACE-LSPR) biosensing device was presented for rapid analysis of cytokine IL-1β among sepsis patients. The ACE-LSPR device is constructed using both bottom-up and top-down sensor fabrication methods, allowing the seamless integration of antibody-conjugated gold nanorod (AuNR) biosensor arrays with microelectrodes on the same microfluidic platform. Applying an AC voltage to microelectrodes while scanning the scattering light intensity variation of the AuNR biosensors results in significantly enhanced biosensing performance. The technologies developed have enabled new capabilities with broad application to advance precision medicine of life-threatening acute illnesses in critical care, which potentially will allow the clinical team to make individualized treatment decisions based on a set of time-resolved biomarker signatures.PHDMechanical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/163129/1/yujing_1.pd

Charakterisierung von benetzbaren Systemen mittels Beschreibung des Kapillarverhaltens unter Rotationsbedingungen

Der Einsatz poröser Materialien wie bspw. für Verpackungen sind sowohl die mechanischen Eigenschaften als auch insbesondere die Interaktion mit Fluiden, besonders Wasser, von größter Bedeutung. Die Optimierung der mechanischen Papiereigenschaften für den vorgesehenen Einsatz erfolgt meist durch eine Vorbehandlung der Fasern vor der Herstellung. So wird z.B. durch die Erhöhung der Fibrillierung des ausgewählten Zellstoffs durch Mahlen die Zugfestigkeit des Papiers deutlich erhöht. Wird das Fasernetzwerk jedoch in einer wässrigen Umgebung eingesetzt, wie z. B. in mikrofluidischen Experimenten, wirkt sich die erhöhte Aufrauhung der Fasern auch erheblich auf das Flüssigkeitsverhalten aus, was sowohl auf die Porenstruktur als auch auf Reibungseffekten zurückgeführt werden kann. Trotz zahlreicher mathematischer Modelle, die in den letzten Jahren entwickelt wurden, ist die Zahl der verschiedenen intrinsischen und extrinsischen Parameter, die die Flüssigkeitsströmung in Papier beeinflussen können, zu groß, um eine angemessene Beschreibung zu liefern. Insbesondere die Reibungskräfte, die als Gegenkraft zur kapillargetriebenen Flüssigkeitsströmung wirken, sind in diesen komplexen Systemen schwer zu quantifizieren. Im Rahmen dieser Arbeit wurden die Reibungskraft durch die wohldefinierte Zentrifugalkraft als Gegenkraft ersetzt, welches ermöglichte, sowohl den mittleren Porenradius des Fasernetzes als auch einen quantitativen Wert, welcher mit dem Druckabfall korreliert, zu bestimmen. Auf diese Weise können die Einflüsse der Fasermorphologie, wie z.B. die Wahl des Zellstoffs, des Mahlgrads, des Fraktionierungsgrads, sowie die Auswirkungen der Fluidströmung durch verschiedene komprimierte Papiere und die Einflüsse der hydrophilen Kanalbreite bestimmt werden. Neben der Anwendung an verschiedenen Substraten wird ebenso die Konstruktion solche eines Rotationsgerätes beschrieben

LAB-ON-A-DISCS FOR QUANTIFICATION OF MICROALGAL LIPIDS AND NATURAL ANTIOXIDANTS OF BEVERAGE SAMPLES

Department of Chemical EngineeringSince the first introduction in 1960s, lab-on-a-disc platform has gained much attention due to its great advantages such as simple operation, rapid reaction, low cost, full integration and automation. Lab-on-a-disc has been applied for various research fields such as biomedical, environmental, energy and food agricultural field. In this thesis, fully integrated lab-on-a-disc platform was demonstrated for quantification of microalgal lipid and natural antioxidants from beverage samples, and disc analyzer was introduced to operate the on-disc optical detection. For microalgae lipid quantification, fully automated lab-on-a-disc platform was developed for rapid on-site quantification of lipids from microalgal samples. The whole serial process for microalgal lipid quantification was integrated on lab-on-a-disc platform. Lab-on-a-disc operation time was 13 min. To integrate liquid-solvent extraction of microalgal lipid on a disc, the organic solvent compatible (for ethanol and n-hexane) lab-on-a-disc was newly fabricated. Fabrication technique was developed by combining thermal fusion bonding and laser printed carbon dot based valving. For natural antioxidant determination from beverage samples, lab-on-a-disc platform was developed to integrate the complicated determination steps of antioxidant activity (AA) and the total phenolic content (TPC) from beverage sample. Different beverage samples including wine, beer, various fruit juices and tea were analyzed using our lab-on-a-disc platform. For disc analyzer, on-disc optical signal detecting instrument was developed. It can detect on-disc signal of absorbance and chemiluminescence from result solution. The trials and errors to develop the disc analyzer for following researchers was demonstrated. From both lab-on-a-disc quantification methods, it is proved that lab-on-a-disc can be effectively used for quantification of microalgal biofuel and food antioxidants. It is expected that our lab-on-a-disc based microalgal lipid quantification works give considerable contribution to the commercial production of microalgal-based biofuels by providing rapid, cost-effective, user-friendly on-site quantification of microalgal lipids. Also, our on-disc natural antioxidant detection works would also contribute to functional beverage industry as providing fast, simple determination of natural antioxidants from beverage sample. Both lab-on-a-disc was fabricated by the organic solvent compatible (for ethanol and n-hexane) manner, and it can be used for chemical engineering fields for quality control.clos

원심력 기반 유체 시스템의 생물의학적 응용에 관한 연구

학위논문 (박사)-- 서울대학교 대학원 : 바이오엔지니어링전공, 2017. 2. 김희찬.This dissertation focuses on the design, fabrication, evaluation, and application of a centrifugal force-based fluidic system based on macro and micro scale engineering disciplines. Unlike other fluid control forces including electrical force, compression force, magnetic force, etc., centrifugal force is capable of manipulating fluids ranging from macro- to micro-scales with high efficiencies regardless of fluid properties. Accordingly, centrifugal force has been extensively used for a great number of biomedical applications. However, the design optimization of such centrifugal force-based fluidic system for practical use is still under investigation due to the inadequate integrating technique, especially for clinical settings, and the strong dependency on geometric designs within spatially varying three different rotational forces (centrifugal, Coriolis, and Euler forces) to precisely regulate the flow of the fluid. Therefore, this dissertation aims to develop a centrifugal force-based fluidic system appropriate for either clinical or biological research environment based on thorough investigations of the fluid flow, the environments created by the rotational forces, and the geometric designs of the system at both the macro- and micro-scale. The macro-scale study involves the evaluation of design strategies for developing a smart all-in-one cardiopulmonary circulatory support device (CCSD) applicable to diverse clinical environments (emergency room (ER), intensive care unit (ICU), operation room (OR), etc.) (Chapter 2, Section 2.1), the evaluation of hemolytic characteristics of centrifugal blood pump (Chapter 2, Section 2.2), and the evaluation of drug sequestration (Chapter 2, Section 2.3) in CCSD component. Smart all-in-one CCSD equipped with a qualified low hemolytic centrifugal blood pump developed in this study resulted in low hemolysis with a free plasma hemoglobin level far less than 50 mg/dL, and an oxygenator membrane made of polyurethane fibers was turned out to be especially susceptible to the analgesic drug loss (41.8%). The micro-scale study involves the numerical evaluation of the Coriolis effects on fluid flow inside a rotating microchannel (Chapter 3, Section 3.1), the feasibility study for the development of a centrifugal microfluidic-based viscometer (Chapter 3, Section 3.2), the evaluation of hypergravity-induced spheroid formation (Chapter 3, Section 3.3), and the cellular adaptation study to hypergravity conditions using human adipose derived stem cell (hASC) and human lung fibroblast (MRC-5) (Chapter 3, 3.4). Application studies performed under fundamental understanding of the microfluidic flows in rotating platform demonstrated new potential uses for centrifugal microfluidic technologies especially for cell research, revealing that hypergravity conditions can be an important environmental cues affecting cellular interactions. Through evaluating various types of centrifugal force-based fluidic system designs for both practical applications and bench-scale experiments, considerable potential of centrifugal force-based fluidic system for introducing new paradigms in the development of medical devices and biomedical research has been demonstrated. The unprecedented integration technique to further miniaturize and improve usability of the centrifugal force-based system might facilitate product innovations, fostering its wide acceptance in the future (Chapter 4).Chapter 1. Introduction 1 1.1 Centrifugal force 1 1.2 Centrifugal force-based biomedical system 2 1.2.1 Cardiopulmonary support system: Macro-scale 3 1.2.2 Centrifugal micro-fluidic biochip: Micro-scale 5 1.3 Research Aims 8 Chapter 2. Macro scale centrifugal-fluidic system for biomedical application 10 2.1 Development of a smart all-in-one cardiopulmonary circulatory support device 10 2.1.1 Introduction 11 2.1.2 Materials and Methods 12 2.1.3 Results and Discussion 14 2.1.4 Conclusion 15 2.2 Evaluation of hemolytic characteristics of centrifugal blood pump 22 2.2.1 Introduction 23 2.2.2 Materials and Methods 26 2.2.3 Results and Discussion 29 2.2.4 Conclusion 33 2.3 Evaluation of drug sequestration in the extracorporeal membrane oxygenation (ECMO) circuit 45 2.3.1 Introduction 45 2.3.2 Materials and Methods 47 2.3.3 Results 50 2.3.4 Discussion 51 2.3.5 Conclusion 54 Chapter 3. Micro scale centrifugal-fluidic system for biomedical application 60 3.1 A numerical study of the Coriolis effect in centrifugal microfluidics with different channel arrangements 60 3.1.1 Introduction 61 3.1.2 Model problem 64 3.1.3 Analytical solution 69 3.1.4 Numerical solution 71 3.1.5 Results 75 3.1.6 Discussion 79 3.1.7 Summary and Conclusion 83 3.2 Centrifugal microfluidic-based viscometer 103 3.2.1 Introduction 103 3.2.2 Materials and Methods 104 3.2.3 Results 105 3.2.4 Discussion 105 3.2.5 Conclusion 106 3.3 Hypergravity-induced multicellular spheroid generation 110 3.3.1 Introduction 111 3.3.2 Materials and Methods 114 3.3.3 Results and Discussion 119 3.2.4 Conclusion 125 3.4 A study on adipose-derived stem cells adaptions to hypergravity environment 144 3.4.1 Introduction 144 3.4.2 Materials and Methods 147 3.4.3 Results 150 3.4.4 Discussion 151 3.4.5 Conclusion 152 Chapter 4. Conclusion and Perspective 161 References 168 Abstract in Korean 193Docto

Development of a microfluidic atmospheric - pressure plasma reactor for water treatment

Conventional water treatment methodologies are often incapable of eliminating chemical and biological pollutants from water sources leaving residual contaminants in treated water. These contaminants are of growing concern due to their potential for adverse health effects from chronic exposure. Non-thermal plasma generated in a dielectric barrier microfluidic plasma reactor, operated at atmospheric pressure, was studied for its potential to treat organic contaminants and pathogenic microorganisms in water. In this thesis, non-thermal plasma generated in a microfluidic reactor was investigated for the degradation of contaminants in water. The overall aim of this thesis is to optimize treatment efficiency of an organic contaminant, i.e. methylene blue, and biological contaminants, i.e. E. coli and P. aeruginosa, in non-thermal plasma by investigating the key process parameters. The microfluidic device in this work incorporated a dielectric barrier discharge generated in a continuous gas flow stream of a two-phase annular flow regime generated in the microchannel of the device. Using air as the carrier gas, low concentrations of long-lived chemicals generated in plasma such as nitrates were detected in plasma treated water. The relative degradation rates of MB were influenced by the residence time of the sample solution in the discharge zone, type of gas applied, channel depth and flow rate. Increasing the residence time inside the plasma region led to higher levels of degradation. Using a 100 μm deep device, oxygen was found to be the most effective gas for promoting MB degradation and by reducing the channel depth to 50 μm, the highest results were obtained, achieving more than a 97% level of degradation with air as the applied gas at a flow rate of 4 ml/min. Effective disinfection of water was achieved using air as the carrier gas. Full inactivation of both bacteria (108 CFU/mL maximum number of each bacteria treated) as monocultures and mixed cultures in water was achieved after 5 seconds of residence time in the plasma zone. The microfluidic system presented here demonstrates proof–of-concept that plasma technology can be utilised as an advanced oxidation process for water treatment, with the potential to achieve total mineralization of organics and hence eliminate water treatment consumables such as filters and disinfectants. A summary of the findings of this work is presented in Chapter 7 including further works