Microdevices and Microsystems for Cell Manipulation

Microfabricated devices and systems capable of micromanipulation are well-suited for the manipulation of cells. These technologies are capable of a variety of functions, including cell trapping, cell sorting, cell culturing, and cell surgery, often at single-cell or sub-cellular resolution. These functionalities are achieved through a variety of mechanisms, including mechanical, electrical, magnetic, optical, and thermal forces. The operations that these microdevices and microsystems enable are relevant to many areas of biomedical research, including tissue engineering, cellular therapeutics, drug discovery, and diagnostics. This Special Issue will highlight recent advances in the field of cellular manipulation. Technologies capable of parallel single-cell manipulation are of special interest

Biophysical investigations of single cells with optically actuated microtools

I demonstrated that optical tweezer-based techniques are capable to explore living cells physical and biochemical properties. For example the measurement of Young’s modulus of endothelial cells or determination of adhesion force of preliminary functionalized microtools. These experiments could be carried out with precisely manipulated two-photon polymerized, purpose designed microstructures. In my thesis I investigated the glutathione's adhesion forces to brain endothelial cells using our novel holographic optical tweezer-based binding force measurement technique. In this series of experiments the used micromanipulators could prevent the cells from photodamage, furthermore the laser microfabrication made it possible to easily change the geometry of the micromanipulator’s probe as the experimental methodology required. In both type of measurement arrangements, we used a cell culturing method where the cells were grown on mask-lithography made walls, which were parallel to the optical axis what enabled us to measure the adhesion force and the stiffness in a direction perpendicular to the cell membrane by approximation of the cell via lateral movement of the trapped microstructure. Our method could be extended in the future to differentiate between multiple and single binding events, to characterize other BBB targeting ligands with the adhesion force on living cells or even to select novel targeting molecules. The functionalization protocol could be easily adapted to immobilize those molecules with covalent bonds, thanks to the variety of PEG-linkers

Soft Tactile Sensors for Mechanical Imaging

Tactile sensing aims to electronically capture physical attributes of an object via mechanical contact. It proves indispensable to many engineering tasks and systems, in areas ranging from manufacturing to medicine and autonomous robotics. Biological skin, which is highly compliant, is able to perform sensing under challenging and highly variable conditions with levels of performance that far exceed what is possible with conventional tactile sensors, which are normally fabricated with non-conforming materials. The development of stretchable, skin-like tactile sensors has, as a result, remained a longstanding goal of engineering. However, to date, artificial tactile sensors that might mimic both the mechanical and multimodal tactile sensory capabilities of biological skin remain far from realization, due to the challenges of fabricating spatially dense, mechanically robust, and compliant sensors in elastic media. Inspired by these demands, this dissertation addresses many aspects of the challenging problem of engineering skin-like electronic sensors. In the first part of the thesis, new methods for the design and fabrication of thin, highly deformable, high resolution tactile sensors are presented. The approach is based on a novel configuration of arrays of microfluidic channels embedded in thin elastomer membranes. To form electrodes, these channels are filled with a metal alloy, eutectic Gallium Indium, that remains liquid at room temperature. Using capacitance sensing techniques, this approach achieves sensing resolutions of 1 mm$^{-1}$. To fabricate these devices, an efficient and robust soft lithography method is introduced, based on a single step cast. An analytical model for the performance of these devices is derived from electrostatic theory and continuum mechanics, and is demonstrated to yield excellent agreement with measured performance. This part of the investigation identified fundamental limitations, in the form of nonmonotonic behavior at low strains, that is demonstrated to generically affect solid cast soft capacitive sensors. The next part of the thesis is an investigation of new methods for designing soft tactile sensors based on multi-layer heterogeneous 3D structures that combine active layers, containing embedded liquid metal electrodes, with passive and mechanically tunable layers, containing air cavities and micropillar geometric supports. In tandem with analytical and computational modeling, these methods are demonstrated to facilitate greater control over mechanical and electronic performance. A new soft lithography fabrication method is also presented, based on the casting, alignment, and fusion of multiple functional layers in a soft polymer substrate. Measurements indicate that the resulting devices achieve excellent performance specifications, and avoid the limiting nonmonotonic behavior identified in the first part of the thesis. In order to demonstrate the practical utility of the devices, we used them to perform dynamic two-dimensional tactile imaging under distributed indentation loads. The results reflect the excellent static and dynamic performance of these devices. The final part of the thesis investigates the utility of the tactile sensing methods pursued here for imaging lumps embedded in simulated tissue. In order to facilitate real-time sensing, an electronic system for fast, array based measurement of small, sub-picofarad (pF) capacitance levels was developed. Using this system, we demonstrated that it is possible to accurately capture strain images depicting small lumps embedded in simulated tissue with either an electronic imaging system or a sensor worn on the finger, supporting the viability of wearable sensors for tactile imaging in medicine. In conclusion, this dissertation confronts many of the most vexing problems arising in the pursuit of skin-like electronic sensors, including fundamental operating principles, structural and functional electronic design, mechanical and electronic modeling, fabrication, and applications to biomedical imaging. The thesis also contributes knowledge needed to enable applications of tactile sensing in medicine, an area that has served as a key source of motivation for this work, and aims to facilitate other applications in areas such as manufacturing, robotics, and consumer electronics.Ph.D., Electrical Engineering -- Drexel University, 201

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

Application and Development of Mechanoresponsive Polymer Structures

Mechanoresponsive Systeme antworten auf mechanische Reize mit einer Eigenschaftsänderung. Diese Dissertation umfasst die Arbeiten mit zwei mechanoresponsiven Systemen, die optisch auf mechanische Reize antworten. Sie basieren auf polymeren Strukturen, einer Polymerbürste und einem Hydrogelnetzwerk. Ihr optischer Antwortmechanismus ermöglicht die Beobachtung wirkender Kräfte als ein Ansatz zur in situ-Kraftmessung. Im ersten Teil wird ein existierendes, mechanoresponsives System zur Anwendung gebracht, das auf einer mit Fluoreszenzfarbstoff markierten Polyelektrolytbürste basiert. Die Ladungen des Polyelektrolyts können die Fluoreszenz des Farbstoffs unterdrücken, sodass lokale Kompression und Zugspannung über die Fluoreszenzintensität unterschieden werden können. Die mechanoresponsive Polymerbürste wurde als mechanosensitive Oberflächenbeschichtung angewandt, um Unterschiede in der Kontaktspannungsverteilung von Gecko-inspirierten adhäsiven Mikrostempelstrukturen aufzuklären. Die erarbeiteten Ergebnisse und daraus abgeleiteten Ablösemechanismen der Mikrostempeltypen deckten sich qualitativ mit Vorhersagen aus theoretischen Ansätzen. Aufgrund geometrischer Einschränkungen einer planaren Oberflächenbeschichtung zielt der zweite Teil darauf ab, dieses mechanoresponsive Prinzip in ein dreidimensionales Netzwerk zu überführen und ein mechanoresponsives Hydrogelnetzwerk als Plattform zur Kraftmessung zu entwickeln. Konzeptionell besitzt ein homogenes Netzwerk vorhersagbare mechanische Eigenschaften, sodass lokale optische Antworten auf mechanische Kräfte ermöglichen könnten, die wirkenden Kräfte zu lokalisieren und quantifizieren. Basierend auf einer Gestaltung nach der Flory-Rehner-Theorie wurden Präkursoren mit vordefinierter Größe und Architektur für die Hydrogelherstellung eingesetzt, um auf ein homogenes Netzwerk abzuzielen. Zu diesem Zweck wurde das Mischungsvolumen durch Tropfenmikrofluidik reduziert. Für den optischen Antwortmechanismus wurden die Hydrogelnetzwerk-Präkursoren mit zwei verschiedenen Fluorophoren markiert, die sich durch abstandsabhängige Emission über Förster-Resonanzenergietransfer auszeichnen. Die Funktionalität des optischen Antwortmechanismus wurde auf globaler Ebene durch Kollabieren und kontrolliertes Quellen des Netzwerks, dann auf lokalisierter Ebene durch definierte mechanische Belastung mit Rasterkraftmikroskopie gezeigt. Durch ihre Anpassbarkeit könnte die Hydrogelplattform zukünftig verschiedenste Anwendungen im Bereich intrisischer Kraftmessung weicher Materie bedienen.Mechanoresponsive systems respond to mechanical triggers by changes in a certain property. This thesis covers the work conducted with two mechanoresponsive systems that respond optically to mechanical triggers. These two systems are based on polymer structures, a polymer brush and a hydrogel network. Thus, the optical response mechanism allows observing acting forces as an approach to force sensing in situ. In the first part, an existing mechanoresponsive system based on a polyelectrolyte brush labeled with a fluorescent dye is engaged in application. The charges of the polyelectrolyte are able to quench the fluorescence of the dye so that local compression or tension can be distinguished from the local fluorescence intensity. The mechanoresponsive polymer brush was applied as mechanosensitive surface coating to elucidate differences in the contact stress distributions of gecko-inspired adhesive micropillar structures. The determined results and the derived detachment mechanisms of the micropillar types were in qualitative accordance with predictions from theoretical approaches. Overcoming the geometrical limitations of a planar surface coating, the second part aims at translating the mechanoresponse principle to a three-dimensional network and developing a mechanoresponsive hydrogel as a platform for force sensing. Conceptually, a homogeneous network allows to predict mechanical properties so that localized optical mechanoresponses could enable locating and quantifying acting forces. Based on network design principles from the Flory-Rehner theory, precursors with predefined size and architecture were utilized in hydrogel preparation, aiming for a homogeneous network. Further in this regard, the mixing volume was reduced by employing droplet microfluidics. As optical response mechanism, the hydrogel network precursors were labeled with two kinds of fluorophore, featuring distance-dependent emission from Förster Resonance Energy Transfer. The functionality of the optical response mechanism was demonstrated on global level by collapsing and controlled swelling of the network, and on a localized level by defined mechanical stress, applied with Atomic Force Microscopy. Owing to its adjustability, the hydrogel platform might be employed in various applications that require intrinsic force sensing of soft matter in future

A Polymer-Based Microfluidic Device with Electrolyte-Enabled Distributed Transducers (EEDT) for Distributed Load Detection

The capability of detecting distributed static and dynamic loads is indispensable in a wide variety of applications, such as examining anatomical structures of biological tissues in tissue health analysis and minimally invasive surgery (MIS) and determining the texture of an object in robotics. This dissertation presents a comprehensive study of a polymer-based microfluidic device with electrolyte-enabled distributed transducers and demonstrates a new concept on using a single microfluidic device for distributed-load detection, which takes advantage of the low-cost microfluidic fabrication technology and the low modulus and biocompatibility of polymer. The core of the device is a single deformable polymer microstructure integrated with electrolyte-enabled transducers. While distributed loads are converted to different levels of deflections by the polymer microstructure, the deflections of the microstructure are translated to resistance changes by the five pairs of distributed transducers underneath the microstructure. Firstly, the design and working principle of the device is described. Then, due to its simple but efficient configuration, a standard fabrication process well developed for polydimethylsiloxane(PDMS)-based microfluidic devices is detailed and employed to fabricate this device. After that, the experimental setups for characterizing the device performance in static, step and sinusoidal inputs are illustrated. The experimental data then are collected and processed by using custom-built electronic circuits and custom LabVIEW/Matlab program to characterize the device performance. Lastly, the performance analysis of the device is conducted to obtain the performance parameters such as device sensitivity and load resolution. In summary, this polymer-based microfluidic device not only demonstrates the new concept and the capability of detecting distributed static and dynamic loads with a single device, with a thorough experimental study on the performance and characterization of this PDMS-based microfluidic device to correlate the device performance to its design parameters, but also the potential application of directly adopting this experimental method to measure the elasticity/viscoelasticity of a soft tissue

Microfabricated platforms to investigate cell mechanical properties

Mechanical stimulation has been imposed on living cells using several approaches. Most early investigations were conducted on groups of cells, utilizing techniques such as substrate deformation and flow-induced shear. To investigate the properties of cells individually, many conventional techniques were utilized, such as AFM, optical traps/optical tweezers, magnetic beads, and micropipette aspiration. In specific mechanical interrogations, microelectro- mechanical systems (MEMS) have been designed to probe single cells in different interrogation modes. To exert loads on the cells, these devices often comprise piezo-electric driven actuators that attach directly to the cell or move a structure on which cells are attached. Uniaxial and biaxial pullers, micropillars, and cantilever beams are examples of MEMS devices. In this review, the methodologies to analyze single cell activity under external loads using microfabricated devices will be examined. We will focus on the mechanical interrogation in three different regimes: compression, traction, and tension, and discuss different microfabricated platforms designed for these purposes

Microfabricated tactile sensors for biomedical applications: a review

During the last decades, tactile sensors based on different sensing principles have been developed due to the growing interest in robotics and, mainly, in medical applications. Several technological solutions have been employed to design tactile sensors; in particular, solutions based on microfabrication present several attractive features. Microfabrication technologies allow for developing miniaturized sensors with good performance in terms of metrological properties (e.g., accuracy, sensitivity, low power consumption, and frequency response). Small size and good metrological properties heighten the potential role of tactile sensors in medicine, making them especially attractive to be integrated in smart interfaces and microsurgical tools. This paper provides an overview of microfabricated tactile sensors, focusing on the mean principles of sensing, i.e., piezoresistive, piezoelectric and capacitive sensors. These sensors are employed for measuring contact properties, in particular force and pressure, in three main medical fields, i.e., prosthetics and artificial skin, minimal access surgery and smart interfaces for biomechanical analysis. The working principles and the metrological properties of the most promising tactile, microfabricated sensors are analyzed, together with their application in medicine. Finally, the new emerging technologies in these fields are briefly described

Engineering patterned and dynamic surfaces for the spatio-temporal control of cell behaviour

Stem cell shape and mechanical properties in vitro can be directed by geometrically defined micropatterned adhesion substrates. However conventional methods are limited by the fixed micropattern design, which cannot recapitulate the dynamic changes of the natural cell microenvironment. Recent advancements in microfabrication technologies in combination with the use of light-responsive materials, allow to manipulate the shape of living cells in real-time in a non-invasive Spatio-temporal controlled way to introduce additional geometrically defined adhesion sites and to study relative cell behaviour. Here, the confocal laser technique is exploited for dynamically evaluate the variation over time of the tensional and morphological cell state. This method allows the precise control of specific actin structures that regulate cell architecture. Actin filament bundles, initially randomly organized in circular-shaped cells, are induced to align and distribute to form a rectangular-shaped cell in response to specific dynamic changes in the cell adhesion pattern. The changes in morphology also reflect dramatic changes in FAs distribution, cell mechanics, nuclear morphology, and chromatin conformation. The reported strategy is convenient to explore the cell-substrate interface and the mechanisms through which cell geometry regulates cell signalling in a facile and cost-effective manner and it open new routes to understand how the field of dynamic platforms should potentially contribute to unveil complex biological events such as the modulation of cell shape