Manipulation of Cell and Particle Trajectory in Microfluidic Devices

Microfluidics, the manipulation of fluid samples on the order of nanoliters and picoliters, is rapidly emerging as an important field of research. The ability to miniaturize existing scientific and medical tools, while also enabling entirely new ones, positions microfluidic technology at the forefront of a revolution in chemical and biological analysis. There remain, however, many hurdles to overcome before mainstream adoption of these devices is realized. One area of intense study is the control of cell motion within microfluidic channels. To perform sorting, purification, and analysis of single cells or rare populations, precise and consistent ways of directing cells through the microfluidic maze must be perfected. The aims of this study focused on developing novel and improved methods of controlling the motion of cells within microfluidic devices, while simultaneously probing their physical and chemical properties. To this end we developed protein-patterned smart surfaces capable of inducing changes in cell motion through interaction with membrane-bound ligands. By linking chemical properties to physical behavior, protein expression could then be visually identified without the need for traditional fluorescent staining. Tracking and understanding motion on cytotactic surfaces guided our development of new software tools for analyzing this motion. To enhance these cell-surface interactions, we then explored methods to adjust and measure the proximity of cells to the channel walls using electrokinetic forces and 3D printed microstructures. Combining our work with patterned substrates and 3-dimensional microfabrication, we created micro-robots capable of rapid and precise movements via magnetic actuation. The micro-robots were shown to be effective tools for mixing laminar flows, capturing or transporting individual cells, and selectively isolating cells on the basis of size. In the course of development of these microfluidic tools we gained valuable new insights into the differences and limitations of planar vs. 3D lithography, especially for fabrication of magnetic micro-machines. This work as a whole enables new mechanisms of control within microfluidics, improving our ability to detect, sort, and analyze cells in both a high throughput and high resolution manner

Microfluidics and Nanofluidics Handbook

The Microfluidics and Nanofluidics Handbook: Two-Volume Set comprehensively captures the cross-disciplinary breadth of the fields of micro- and nanofluidics, which encompass the biological sciences, chemistry, physics and engineering applications. To fill the knowledge gap between engineering and the basic sciences, the editors pulled together key individuals, well known in their respective areas, to author chapters that help graduate students, scientists, and practicing engineers understand the overall area of microfluidics and nanofluidics. Topics covered include Finite Volume Method for Numerical Simulation Lattice Boltzmann Method and Its Applications in Microfluidics Microparticle and Nanoparticle Manipulation Methane Solubility Enhancement in Water Confined to Nanoscale Pores Volume Two: Fabrication, Implementation, and Applications focuses on topics related to experimental and numerical methods. It also covers fabrication and applications in a variety of areas, from aerospace to biological systems. Reflecting the inherent nature of microfluidics and nanofluidics, the book includes as much interdisciplinary knowledge as possible. It provides the fundamental science background for newcomers and advanced techniques and concepts for experienced researchers and professionals

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

All-in-one microsystem for long-term cell culturing and real-time chip-level lensless microscopy

The study is to concept, develop and evaluate an all-in-one microsystem with combined long-term animal cell culturing and real-time chip-level lensless microscopy functions suitable for applications in cell biology studies, and at the point-of-use. The microsystem consists of a 5 megapixel CMOS image sensor, a disposable microchip for cell culture, heating elements and LED illumination. The overall size is only 40 mm x 40 mm x 50 mm. The disposable microchip for cell culture is composed of a polymer microfluidic interface and a silicon micro-cavity chip with a 1 µm thick 1 mm x 1 mm transparent Si3N4 bottom membrane, which is directly placed onto the image sensor surface. Under the collimated LED illumination, the optical resolution of the lensless imaging module is only dependent on the digital resolution of the image sensor, which amounts to 3.5 µm (double pixel pitches). The imaging quality is proven comparable to a 4x optical microscope without image computation or processing. Both the morphologies of different cell cultures (L929, A549 and T47D) and the single cells with colorimetric staining can be clearly visualized in real time. With the additional deposition of an interference filter on the image sensor surface, fluorescence spreading cells in culture are observed on the chip under a common blue LED illumination. The temperature for the incubating module is controlled at 37±0.2°C in the room environment. Mammalian cells (L929 and A549) are cultured with conventional culture medium and monitored under the time-lapse lensless microscopy by the all-in-one microsystem up to 5 days outside a laboratory incubator. Very fast operational processes, such as cell loading, passaging and staining, have been readily carried out and monitored in real-time by the platform. Besides cell cultures in monolayer, the formation of 3D clusters of L929 cells has also been demonstrated and recorded under time-lapse lensless microscopy by using the all-in-one microsystem.Das Ziel der vorliegenden Dissertation ist die Konzeption, Entwicklung und Evaluation eines All-in-One-Mikrosystems mit der Kombination aus Langzeit Kultivierung von Tierzellen und Echtzeit Linsenloser Mikroskopie Funktionen auf Chip Level, die für Anwendungen in zellbiologischen Studien sowie für Point-of-use geeignet sind. Das Mikrosystem besteht aus einem 5 Megapixel CMOS-Bildsensor, einem Einweg-Mikrochip für die Zellkultur, Heizelementen sowie einer LED-Beleuchtung Die Gesamtgröße beträgt nur 40 mm x 40 mm x 50 mm. Der Einweg-Mikrochip für die Zellkultur besteht aus einer polymeren, mikrofluidischen Grenzfläche und einem Silizium-Mikrohohlraum-Chip mit einer 1 µm dicken und 1 mm x 1 mm transparenten Si3N4-Bodenmembran, die direkt auf die Bildsensoroberfläche aufgesetzt wird. Unter der kollimierten LED-Beleuchtung ist die optische Auflösung des linsenlosen Abbildungsmoduls nur von der digitalen Auflösung des Bildsensors abhängig, was 3,5 µm beträgt (Doppelpixelabstände). Die Bildqualität ist vergleichbar mit einem 4x optischen Mikroskop ohne Bildberechnung oder Verarbeitung. Sowohl die Morphologien verschiedener Zellkulturen (L929, A549 und T47D) als auch die einzelnen Zellen mit farbmetrischer Färbung können in Echtzeit deutlich sichtbar gemacht werden. Mit der zusätzlichen Abscheidung eines Interferenzfilters auf der Bildsensoroberfläche werden fluoreszenzverteilende Zellen in Kultur auf dem Chip unter einer gemeinsamen blauen LED-Beleuchtung beobachtet. Die Temperatur für das Inkubationsmodul wird bei 37 ± 0,2 ° C in der Raumumgebung festgelegt. Säugetierzellen (L929 und A549) werden mit herkömmlichem Kulturmedium kultiviert und unter der Zeitrafferlinsenmikroskopie durch das All-in-One-Mikrosystem bis zu 5 Tage außerhalb eines Laborinkubators überwacht. Sehr schnelle operative Prozesse wie z. B. Zellbeladung, Durchfluss und Färbung wurden in Echtzeit durch die Plattform durchgeführt und überwacht. Neben den Zellkulturen in der Monoschicht wurde auch die Bildung von 3D-Clustern von L929-Zellen in Zeitraffer bei lichtempfindlicher Mikroskopie unter Verwendung des All-in-One-Mikrosystems nachgewiesen und aufgezeichnet

BioMEMS

As technological advancements widen the scope of applications for biomicroelectromechanical systems (BioMEMS or biomicrosystems), the field continues to have an impact on many aspects of life science operations and functionalities. Because BioMEMS research and development require the input of experts who use different technical languages and come from varying disciplines and backgrounds, scientists and students can avoid potential difficulties in communication and understanding only if they possess a skill set and understanding that enables them to work at the interface of engineering and biosciences. Keeping this duality in mind throughout, BioMEMS: Science and Engineering Perspectives supports and expedites the multidisciplinary learning involved in the development of biomicrosystems. Divided into nine chapters, it starts with a balanced introduction of biological, engineering, application, and commercialization aspects of the field. With a focus on molecules of biological interest, the book explores the building blocks of cells and viruses, as well as molecules that form the self-assembled monolayers (SAMs), linkers, and hydrogels used for making different surfaces biocompatible through functionalization. The book also discusses: Different materials and platforms used to develop biomicrosystems Various biological entities and pathogens (in ascending order of complexity) The multidisciplinary aspects of engineering bioactive surfaces Engineering perspectives, including methods of manufacturing bioactive surfaces and devices Microfluidics modeling and experimentation Device level implementation of BioMEMS concepts for different applications. Because BioMEMS is an application-driven field, the book also highlights the concepts of lab-on-a-chip (LOC) and micro total analysis system (μTAS), along with their pertinence to the emerging point-of-care (POC) and point-of-need (PON) applications

BioMEMS

As technological advancements widen the scope of applications for biomicroelectromechanical systems (BioMEMS or biomicrosystems), the field continues to have an impact on many aspects of life science operations and functionalities. Because BioMEMS research and development require the input of experts who use different technical languages and come from varying disciplines and backgrounds, scientists and students can avoid potential difficulties in communication and understanding only if they possess a skill set and understanding that enables them to work at the interface of engineering and biosciences. Keeping this duality in mind throughout, BioMEMS: Science and Engineering Perspectives supports and expedites the multidisciplinary learning involved in the development of biomicrosystems. Divided into nine chapters, it starts with a balanced introduction of biological, engineering, application, and commercialization aspects of the field. With a focus on molecules of biological interest, the book explores the building blocks of cells and viruses, as well as molecules that form the self-assembled monolayers (SAMs), linkers, and hydrogels used for making different surfaces biocompatible through functionalization. The book also discusses: Different materials and platforms used to develop biomicrosystems Various biological entities and pathogens (in ascending order of complexity) The multidisciplinary aspects of engineering bioactive surfaces Engineering perspectives, including methods of manufacturing bioactive surfaces and devices Microfluidics modeling and experimentation Device level implementation of BioMEMS concepts for different applications. Because BioMEMS is an application-driven field, the book also highlights the concepts of lab-on-a-chip (LOC) and micro total analysis system (μTAS), along with their pertinence to the emerging point-of-care (POC) and point-of-need (PON) applications

New Trends and Applications in Femtosecond Laser Micromachining

This book contains the scientific contributions to the Special Issue entitled: "New Trends and Applications in Femtosecond Laser Micromachining". It covers an array of subjects, from the basics of femtosecond laser micromachining to specific applications in a broad spectra of fields such biology, photonics and medicine