Construction of artificial skin tissue with placode-like structures in well-defined patterns using dielectrophoresis

During embryonic development of animal skin tissue, the skin cells form regular patterns of high cell density (placodes) where hair or feathers will be formed. These placodes are thought to be formed by the aggregation of dermal cells into condensates. The aggregation process is thought to be controlled by a reaction-diffusion mechanism of activator and inhibitor molecules, and involve mechanical forces between cells and cells with the matrix. In this project, placode formation in chicken embryonic skin cells was used as a model system for the study of the mechanism by which the placodes are formed. Artificial aggregates of chicken embryonic skin cells were created by suspending them in a 300 mM low conductivity sorbitol solution and attracting them by positive dielectrophoresis to high field regions within microelectrode arrays by applying a 10 - 20 Vpk-pk 1 MHz signal across the microelectrodes. It was demonstrated that using this method aggregates can be produced in a large variety of patterns and that the distance between the aggregates and aggregate size and shape within the pattern can be controlled effectively. Custom-built image analysis tools were developed in LabVIEW to analyze the patterns formed. The formation of aggregates by dielectrophoresis was followed by an immobilization phase of the resulting patterns inside a gel matrix, forming an artificial skin. Nutrients and oxygen were supplied externally. Long-term incubation of the artificial skin shows that embryonic skin cells in the aggregates were viable and showed behavior similar to that of developing embryonic skin, including further aggregation of the cells and the formation of cell condensates. The domain size was shown to have an influence on the condensation process, with cells in small aggregates forming only one condensate near the centre of the aggregate, and several condensates in larger aggregates. Whilst the distribution of cell condensates within the aggregates in round large aggregates is predominantly random, some line formation could be observed in linear aggregations, indicating some self-organization may be occurring

Thermal Bimorph Micro-Cantilever Based Nano-Calorimeter for Sensing of Energetic Materials

The objective of this study is to develop a robust portable nano-calorimeter sensor for detection of energetic materials, primarily explosives, combustible materials and propellants. A micro-cantilever sensor array is actuated thermally using bi-morph structure consisting of gold (Au: 400 nm) and silicon nitride (Si3N4: 600 nm) thin film layers of sub-micron thickness. An array of micro-heaters is integrated with the microcantilevers at their base. On electrically activating the micro-heaters at different actuation currents the microcantilevers undergo thermo-mechanical deformation, due to differential coefficient of thermal expansion. This deformation is tracked by monitoring the reflected ray from a laser illuminating the individual microcantilevers (i.e., using the optical lever principle). In the presence of explosive vapors, the change in bending response of microcantilever is affected by the induced thermal stresses arising from temperature changes due to adsorption and combustion reactions (catalyzed by the gold surface). A parametric study was performed for investigating the optimum values by varying the thickness and length in parallel with the heater power since the sensor sensitivity is enhanced by the optimum geometry as well as operating conditions for the sensor (e.g., temperature distribution within the microcantilever, power supply, concentration of the analyte, etc.). Also, for the geometry present in this study the nano-coatings of high thermal conductivity materials (e.g., Carbon Nanotubes: CNTs) over the microcantilever surface enables maximizing the thermally induced stress, which results in the enhancement of sensor sensitivity. For this purpose, CNTs are synthesized by post-growth method over the metal (e.g., Palladium Chloride: PdCl2) catalyst arrays pre-deposited by Dip-Pen Nanolithography (DPN) technique. The threshold current for differential actuation of the microcantilevers is correlated with the catalytic activity of a particular explosive (combustible vapor) over the metal (Au) catalysts and the corresponding vapor pressure. Numerical modeling is also explored to study the variation of temperature, species concentration and deflection of individual microcantilevers as a function of actuation current. Joule-heating in the resistive heating elements was coupled with the gaseous combustion at the heated surface to obtain the temperature profile and therefore the deflection of a microcantilever by calculating the thermally induced stress and strain relationship. The sensitivity of the threshold current of the sensor that is used for the specific detection and identification of individual explosives samples - is predicted to depend on the chemical kinetics and the vapor pressure. The simulation results showed similar trends with the experimental results for monitoring the bending response of the microcantilever sensors to explosive vapors (e.g., Acetone and 2-Propanol) as a function of the actuation current

Microelectromechanical Systems and Devices

The advances of microelectromechanical systems (MEMS) and devices have been instrumental in the demonstration of new devices and applications, and even in the creation of new fields of research and development: bioMEMS, actuators, microfluidic devices, RF and optical MEMS. Experience indicates a need for MEMS book covering these materials as well as the most important process steps in bulk micro-machining and modeling. We are very pleased to present this book that contains 18 chapters, written by the experts in the field of MEMS. These chapters are groups into four broad sections of BioMEMS Devices, MEMS characterization and micromachining, RF and Optical MEMS, and MEMS based Actuators. The book starts with the emerging field of bioMEMS, including MEMS coil for retinal prostheses, DNA extraction by micro/bio-fluidics devices and acoustic biosensors. MEMS characterization, micromachining, macromodels, RF and Optical MEMS switches are discussed in next sections. The book concludes with the emphasis on MEMS based actuators

Biofabrication of three-dimensional liver cell-embedded tissue constructs for in vitro drug metabolism models

In their normal in vivo matrix milieu, tissues assume complex well-organized threedimensional architectures. Therefore, a primary aim in the tissue engineering design process is to fabricate an optimal analog of the in vivo scenario. This challenge can be addressed by applying emerging layered biofabrication approaches in which the precise configuration and composition of cells and bioactive matrix components can recapitulate the well-defined three-dimensional biomimetic microenvironments that promote cell-cell and cell-matrix interactions. Furthermore, the advent of and refinements in microfabricated systems can present physical and chemical cues to cells in a controllable and reproducible fashion unrealizable with conventional tissue culture, resulting in highfidelity, high-throughput in vitro models. As such, the convergence of layered solid freeform fabrication (SFF) technologies along with microfabrication techniques, a threedimensional micro-organ device can serve as an in vitro platform for cell culture, drug screening, or to elicit further biological insights, particularly for NASA’s interest of a flight-suitable high-fidelity microscale platform to study drug metabolism in space and planetary environments. A proposed model in this thesis involves the combinatorial setup of an automated syringe-based, layered direct cell writing bioprinting process with micropatterning techniques to fabricate a microscale in vitro device housing a chamber of bioprinted three-dimensional cell-encapsulated hydrogel-based tissue constructs in defined design patterns that biomimics the cell’s natural microenvironment for enhanced performance and functionality. In order to assess the structural formability and biological feasibility of such a micro-organ, reproducibly fabricated tissue constructs arebiologically characterized for both viability and cell-specific function. Another key facet of the in vivo microenvironment that is recapitulated with the in vitro system is the necessary dynamic perfusion of the three-dimensional microscale liver analog with cells probed for their collective drug metabolic function and suitability as a drug metabolism model. This thesis details the principles, methods, and engineering science basis that undergird the direct cell writing fabrication process development and adaptation of microfluidic devices for the creation of a drug screening model, thereby establishing a novel drug metabolism study platform for NASA’s interest to adopt a microfluidic microanalytical device with an embedded three-dimensional microscale liver tissue analog to assess drug pharmacokinetic profiles in planetary environments.Ph.D., Mechanical Engineering -- Drexel University, 200

A Review on Mechanics and Mechanical Properties of 2D Materials - Graphene and Beyond

Since the first successful synthesis of graphene just over a decade ago, a variety of two-dimensional (2D) materials (e.g., transition metal-dichalcogenides, hexagonal boron-nitride, etc.) have been discovered. Among the many unique and attractive properties of 2D materials, mechanical properties play important roles in manufacturing, integration and performance for their potential applications. Mechanics is indispensable in the study of mechanical properties, both experimentally and theoretically. The coupling between the mechanical and other physical properties (thermal, electronic, optical) is also of great interest in exploring novel applications, where mechanics has to be combined with condensed matter physics to establish a scalable theoretical framework. Moreover, mechanical interactions between 2D materials and various substrate materials are essential for integrated device applications of 2D materials, for which the mechanics of interfaces (adhesion and friction) has to be developed for the 2D materials. Here we review recent theoretical and experimental works related to mechanics and mechanical properties of 2D materials. While graphene is the most studied 2D material to date, we expect continual growth of interest in the mechanics of other 2D materials beyond graphene

Polymer Micro- and Nanofluidic Systems for In Vitro Diagnostics: Analyzing Single Cells and Molecules

Polymer micro- and nanofluidic systems, with their critical dimensions, offer a potential to outperform conventional analysis techniques and diagnostic methods by enhancing speed, accuracy, sensitivity and specificity. In this work, applications of microfluidics have been demonstrated to address the existing challenges in stroke diagnosis, by mRNA expression profiling from whole blood within \u3c20 min. A brief overview of various biomarkers for stroke diagnosis is given in chapter 1 followed by design and testing of individual microfluidic modules (chapter 2 and 3) required for the development of POC diagnostic strategy for stroke. We have designed and evaluated the performance of polymer microfluidic devices for the isolation of leukocyte subsets, known for their differential gene expression in the event of stroke. Target cells (T-cells and neutrophils) were selected from with greater purities, from 50 µl whole human blood by using affinity based capture in COC devices within a 6.6 min processing time. In addition, we have also demonstrated the ability to isolate and purify total RNA by using UV activated polycarbonate solid phase extraction platform. Polymer-based nanofluidic devices were used to study the effects of surface charge on the electrodynamic transport dynamics of target molecules. In this work, we report the fabrication of mixed-scale micro- and nanofluidic networks in poly(methylmethacrylate), PMMA, using thermal nanoimprint lithography using a resin stamp and surface modification of polymer nanoslits and nanochannels for the assessment of the associated electrokinetic parameters – surface charge density, zeta potential and electroosmotic flow. This study provided information on possible routes that can be adopted to engineer proper wall chemistry of polymer nanochannels for the enhancement or reduction of solute/wall interactions in a variety of relevant single-molecule studies

Glioma on Chips Analysis of glioma cell guidance and interaction in microfluidic-controlled microenvironment enabled by machine learning

In biosystems, chemical and physical fields established by gradients guide cell migration, which is a fundamental phenomenon underlying physiological and pathophysiological processes such as development, morphogenesis, wound healing, and cancer metastasis. Cells in the supportive tissue of the brain, glia, are electrically stimulated by the local field potentials from neuronal activities. How the electric field may influence glial cells is yet fully understood. Furthermore, the cancer of glia, glioma, is not only the most common type of brain cancer, but the high-grade form of it (glioblastoma) is particularly aggressive with cells migrating into the surrounding tissues (infiltration) and contribute to poor prognosis. In this thesis, I investigate how electric fields in the microenvironment can affect the migration of glioblastoma cells using a versatile microsystem I have developed. I employ a hybrid microfluidic design to combine poly(methylmethacrylate) (PMMA) and poly(dimethylsiloxane) (PDMS), two of the most common materials for microfluidic fabrication. The advantages of the two materials can be complemented while disadvantages can be mitigated. The hybrid microfluidics have advantages such as versatile 3D layouts in PMMA, high dimensional accuracy in PDMS, and rapid prototype turnaround by facile bonding between PMMA and PDMS using a dual-energy double sided tape. To accurately analyze label-free cell migration, a machine learning software, Usiigaci, is developed to automatically segment, track, and analyze single cell movement and morphological changes under phase contrast microscopy. The hybrid microfluidic chip is then used to study the migration of glioblastoma cell models, T98G and U-251MG, in electric field (electrotaxis). The influence of extracellular matrix and chemical ligands on glioblastoma electrotaxis are investigated. I further test if voltage-gated calcium channels are involved in glioblastoma electrotaxis. The electrotaxes of glioblastoma cells are found to require optimal laminin extracellular matrices and depend on different types of voltage-gated calcium channels, voltage-gated potassium channels, and sodium transporters. A reversiblysealed hybrid microfluidic chip is developed to study how electric field and laminar shear can condition confluent endothelial cells and if the biomimetic conditions affect glioma cell adhesion to them. It is found that glioma/endothelial adhesion is mediated by the Ang1/Tie2 signaling axis and adhesion of glioma is slightly increased to endothelial cells conditioned with shear flow and moderate electric field. In conclusion, robust and versatile hybrid microsystems are employed for studying glioma biology with emphasis on cell migration. The hybrid microfluidic tools can enable us to elucidate fundamental mechanisms in the field of the tumor biology and regenerative medicine.Okinawa Institute of Science and Technology Graduate Universit

Shaping surface waves for diagnostics

Infectious diseases continue to kill millions of people every year and are a significant burden on the socio-economic development of developing countries. After many years of international policy aimed at containing diseases, it has recently become an explicit aim to move towards elimination of infectious diseases. However, if this is to occur, it will be necessary to have highly eficacious diagnostic tools to ensure infected individuals are identified and treated. However, the diagnosis of infectious diseases in the developed and developing world requires the full integration of complex assays in easy-to-use platforms with robust analytical performances at low cost. Many relevant bioanalytical technologies have been developed for use in laboratories and clinics, including the current gold standard for the diagnosis of tuberculosis and malaria. The miniaturization and integration of complex functions into lab-on-a-chip (LOC)technologies using microfluidics have only had limited success in translating diagnosis assays out of a centralized laboratory to point-of-care (POC) settings, because they still remain constrained due to chip interconnection and they are either not likely to go out of research laboratories or are not appropriate for low resource settings. In this thesis, a new microfluidic platform was developed that reduced the dependency of the diagnostic procedure on large laboratory instruments providing simplicity of use, enabling the patient sample to be processed and diagnosed on a low cost, disposable biochip. Surface acoustic wave (SAW) devices, which are commonly used in mobile phone technologies, were adapted to provide controlled microfluidic functions by shaping the SAW using particular designs of electrodes and phononic structures. The control of lateral positioning of the SAW was demonstrated using a slanted finger interdigitated transducer (IDT) in a frequency tuneable manner allowing microfluidic functions such as mixing, moving and merging, sequentially performed using a single IDT both on the substrate and on a disposable chip. Alternatively, phononic bandgaps were designed to break the symmetry of the SAW in a tuneable manner and gradient index phononic crystals (GRIN-PC) lenses were designed to focus the SAW and successfully increased the amplitude of the wave by a factor 3 while the focal position could be tuned with the frequency. The potential of these techniques was demonstrated by controlling the amplitude and direction of water jet towers by the use of a phononic horn structure that allowed the enhancement of energy at defined positions and by propelling and directing a macrometer scale object in water using a slanted IDT. As proof of concepts of diagnostic devices for the developing world, an immunoassay for tuberculosis using only mobile phone technologies (SAW, light-emitting diode(LED) and complementary metaloxidesemiconductor (CMOS) camera) was demonstrated with a limit of detection of 1 pM, which is the limit required in an interferon-release assay. This limit of detection was only achievable because of the ability of SAW to increase the mixing and to reduce the non-specific binding. Furthermore, a method to enrich malaria infected cells, based on SAW and isopycnic gradient, was also demonstrated and showed an enrichment up to 100x in the equivalent of a fingerprick of blood in 3 seconds. This technique will allow to reduce the limit of detection of the current gold standard. This platform not only opens a clear road toward POC diagnostics due to its size, cost, versatility and ease in integration, but has also the potential to provide useful tools in laboratory settings for large scale, high throughput technologies