PLANNING FOR AUTOMATED OPTICAL MICROMANIPULATION OF BIOLOGICAL CELLS

Optical tweezers (OT) can be viewed as a robot that uses a highly focused laser beam for precise manipulation of biological objects and dielectric beads at micro-scale. Using holographic optical tweezers (HOT) multiple optical traps can be created to allow several operations in parallel. Moreover, due to the non-contact nature of manipulation OT can be potentially integrated with other manipulation techniques (e.g. microfluidics, acoustics, magnetics etc.) to ensure its high throughput. However, biological manipulation using OT suffers from two serious drawbacks: (1) slow manipulation due to manual operation and (2) severe effects on cell viability due to direct exposure of laser. This dissertation explores the problem of autonomous OT based cell manipulation in the light of addressing the two aforementioned limitations. Microfluidic devices are well suited for the study of biological objects because of their high throughput. Integrating microfluidics with OT provides precise position control as well as high throughput. An automated, physics-aware, planning approach is developed for fast transport of cells in OT assisted microfluidic chambers. The heuristic based planner employs a specific cost function for searching over a novel state-action space representation. The effectiveness of the planning algorithm is demonstrated using both simulation and physical experiments in microfluidic-optical tweezers hybrid manipulation setup. An indirect manipulation approach is developed for preventing cells from high intensity laser. Optically trapped inert microspheres are used for manipulating cells indirectly either by gripping or pushing. A novel planning and control approach is devised to automate the indirect manipulation of cells. The planning algorithm takes the motion constraints of the gripper or pushing formation into account to minimize the manipulation time. Two different types of cells (Saccharomyces cerevisiae and Dictyostelium discoideum) are manipulated to demonstrate the effectiveness of the indirect manipulation approach

Active particles in complex and crowded environments

Differently from passive Brownian particles, active particles, also known as self-propelled Brownian particles or microswimmers and nanoswimmers, are capable of taking up energy from their environment and converting it into directed motion. Because of this constant flow of energy, their behavior can be explained and understood only within the framework of nonequilibrium physics. In the biological realm, many cells perform directed motion, for example, as a way to browse for nutrients or to avoid toxins. Inspired by these motile microorganisms, researchers have been developing artificial particles that feature similar swimming behaviors based on different mechanisms. These man-made micromachines and nanomachines hold a great potential as autonomous agents for health care, sustainability, and security applications. With a focus on the basic physical features of the interactions of self-propelled Brownian particles with a crowded and complex environment, this comprehensive review will provide a guided tour through its basic principles, the development of artificial self-propelling microparticles and nanoparticles, and their application to the study of nonequilibrium phenomena, as well as the open challenges that the field is currently facing. © 2016 American Physical Society

Magnetically Driven Micro and Nanorobots

Manipulation and navigation of micro and nanoswimmers in different fluid environments can be achieved by chemicals, external fields, or even motile cells. Many researchers have selected magnetic fields as the active external actuation source based on the advantageous features of this actuation strategy such as remote and spatiotemporal control, fuel-free, high degree of reconfigurability, programmability, recyclability, and versatility. This review introduces fundamental concepts and advantages of magnetic micro/nanorobots (termed here as "MagRobots") as well as basic knowledge of magnetic fields and magnetic materials, setups for magnetic manipulation, magnetic field configurations, and symmetry-breaking strategies for effective movement. These concepts are discussed to describe the interactions between micro/nanorobots and magnetic fields. Actuation mechanisms of flagella-inspired MagRobots (i.e., corkscrew-like motion and traveling-wave locomotion/ciliary stroke motion) and surface walkers (i.e., surface-assisted motion), applications of magnetic fields in other propulsion approaches, and magnetic stimulation of micro/nanorobots beyond motion are provided followed by fabrication techniques for (quasi)spherical, helical, flexible, wire-like, and biohybrid MagRobots. Applications of MagRobots in targeted drug/gene delivery, cell manipulation, minimally invasive surgery, biopsy, biofilm disruption/eradication, imaging-guided delivery/therapy/surgery, pollution removal for environmental remediation, and (bio)sensing are also reviewed. Finally, current challenges and future perspectives for the development of magnetically powered miniaturized motors are discussed

REGULATION OF SHAPE DYNAMICS AND ACTIN POLYMERIZATION DURING COLLECTIVE CELL MIGRATION

This thesis aims to understand how cells coordinate their motion during collective migration. As previously shown, the motion of individually migrating cells is governed by wave-like cell shape dynamics. The mechanisms that regulate these dynamic behaviors in response to extracellular environment remain largely unclear. I applied shape dynamics analysis to Dictyostelium cells migrating in pairs and in multicellular streams and found that wave-like membrane protrusions are highly coupled between touching cells. I further characterized cell motion by using principle component analysis (PCA) to decompose complex cell shape changes into a serial shape change modes, from which I found that streaming cells exhibit localized anterior protrusion, termed front narrowing, to facilitate cell-cell coupling. I next explored cytoskeleton-based mechanisms of cell-cell coupling by measuring the dynamics of actin polymerization. Actin polymerization waves observed in individual cells were significantly suppressed in multicellular streams. Streaming cells exclusively produced F-actin at cell-cell contact regions, especially at cell fronts. I demonstrated that such restricted actin polymerization is associated with cell-cell coupling, as reducing actin polymerization with Latrunculin A leads to the assembly of F-actin at the side of streams, the decrease of front narrowing, and the decoupling of protrusion waves. My studies also suggest that collective migration is guided by cell-surface interactions. I examined the aggregation of Dictyostelim cells under distinct conditions and found that both chemical compositions of surfaces and surface-adhesion defects in cells result in altered collective migration patterns. I also investigated the shape dynamics of cells suspended on PEG-coated surfaces, which showed that coupling of protrusion waves disappears on touching suspended cells. These observations indicate that collective migration requires a balance between cell-cell and cell-surface adhesions. I hypothesized such a balance is reached via the regulation of cytoskeleton. Indeed, I found cells actively regulate cytoskeleton to retain optimal cell-surface adhesions on varying surfaces, and cells lacking the link between actin and surfaces (talin A) could not retain the optimal adhesions. On the other hand, suspended cells exhibited enhanced actin filament assembly on the periphery of cell groups instead of in cell-cell contact regions, which facilitates their aggregation in a clumping fashion

Program and Proceedings: The Nebraska Academy of Sciences 1880-2009

PROGRAM FRIDAY, APRIL 17, 2009 REGISTRATION FOR ACADEMY, Lobby of Lecture wing, Olin Hall Aeronautics and Space Science, Olin 249 Collegiate Academy, Biology Session A, Olin B Earth Science, Olin 224 Collegiate Academy, Chemistry and Physics, Session A, Olin 324 Biological and Medical Sciences, Session A, Olin 112 Biological and Medical Sciences, Session B, Smith Callen Conference Center Junior Academy, Senior High REGISTRATION, Olin Hall Lobby NWU Health and Sciences Graduate School Fair, Olin and Smith Curtiss Halls Junior Academy, Senior High Competition, Olin 124, Olin 131 Aeronautics and Space Science, Poster Session, Olin 249 History and Philosophy of Science, Olin 325, combined section Teaching of Science and Math, Olin 325, combined section MAIBEN MEMORIAL LECTURE, OLIN B Dr. Donald Frey, Chair, Department of Family Practice, Creighton University Medical Center LUNCH, PATIO ROOM, STORY STUDENT CENTER (pay and carry tray through cafeteria line, or pay at NAS registration desk) Policy and Program Committee Luncheon, Roundup Room Emeriti Luncheon, Presidents Room Aeronautics Group, Conestoga Room Anthropology, Olin 111 Biological and Medical Sciences, Session C, Olin 112 Biological and Medical Sciences, Session D, Smith Callen Conference Center Chemistry and Physics, Section A, Chemistry, Olin A Chemistry and Physics, Section B, Physics, Planetarium Collegiate Academy, Biology Session A, Olin B Collegiate Academy, Biology Session B, Olin 249 Collegiate Academy, Chemistry and Physics, Session A, Olin 324 Junior Academy, Junior High REGISTRATION, Olin Hall Lobby Junior Academy, Senior High Competition, (Final), Olin 110 Junior Academy, Junior High Competition, Olin 124, Olin 131 NJAS Board/Teacher Meeting, Olin 219 Junior Academy, General Awards Presentations, Smith Callen Conference Center BUSINESS MEETING, OLIN B SOCIAL HOUR for Members, Spouses, and Guests First United Methodist Church, 2723 N 50th Street, Lincoln, NE ANNUAL BANQUET and Presentation of Awards and Scholarships First United Methodist Church, 2723 N 50th Street, Lincoln, N

Enzyme Powered Nanomotors Towards Biomedical Applications

[eng] The advancements in nanotechnology enabled the development of new diagnostic tools and drug delivery systems based on nanosystems, which offer unique features such as large surface area to volume ratio, cargo loading capabilities, increased circulation times, as well as versatility and multifunctionality. Despite this, the majority of nanomedicines do not translate into clinics, in part due to the biological barriers present in the body. Synthetic nano- and micromotors could be an alternative tool in nanomedicine, as the continuous propulsion force and potential to modulate the medium may aid tissue penetration and drug diffusion across biological barriers. Enzyme-powered motors are especially interesting for biomedical applications, owing to their biocompatibility and use of bioavailable substrates as fuel for propulsion. This thesis aims at exploring the potential applications of urease-powered nanomotors in nanomedicine. In the first work, we evaluated these motors as drug delivery systems. We found that active urease- powered nanomotors showed active motion in phosphate buffer solutions, and enhanced in vitro drug release profiles in comparison to passive nanoparticles. In addition, we observed that the motors were more efficient in delivering drug to cancer cells and caused higher toxicity levels, due to the combination of boosted drug release and local increase of pH produced by urea breakdown into ammonia and carbon dioxide. One of the major goals in nanomedicine is to achieve localized drug action, thus reducing side-effects. A commonly strategy to attain this is the use moieties to target specific diseases. In our second work, we assessed the ability of urease-powered nanomotors to improve the targeting and penetration of spheroids, using an antibody with therapeutic potential. We showed that the combination of active propulsion with targeting led to a significant increase in spheroid penetration, and that this effect caused a decrease in cell proliferation due to the antibody’s therapeutic action. Considering that high concentrations of nanomedicines are required to achieve therapeutic efficiency; in the third work we investigated the collective behavior of urease-powered nanomotors. Apart from optical microscopy, we evaluated the tracked the swarming behavior of the nanomotors using positron emission tomography, which is a technique widely used in clinics, due to its noninvasiveness and ability to provide quantitative information. We showed that the nanomotors were able to overcome hurdles while swimming in confined geometries. We observed that the nanomotors swarming behavior led to enhanced fluid convection and mixing both in vitro, and in vivo within mice’s bladders. Aiming at conferring protecting abilities to the enzyme-powered nanomotors, in the fourth work, we investigated the use of liposomes as chassis for nanomotors, encapsulating urease within their inner compartment. We demonstrated that the lipidic bilayer provides the enzymatic engines with protection from harsh acidic environments, and that the motility of liposome-based motors can be activated with bile salts. Altogether, these results demonstrate the potential of enzyme-powered nanomotors as nanomedicine tools, with versatile chassis, as well as capability to enhance drug delivery and tumor penetration. Moreover, their collective dynamics in vivo, tracked using medical imaging techniques, represent a step-forward in the journey towards clinical translation.[spa] Recientes avances en nanotecnología han permitido el desarrollo de nuevas herramientas para el diagnóstico de enfermedades y el transporte dirigido de fármacos, ofreciendo propiedades únicas como encapsulación de fármacos, el control sobre la biodistribución de estos, versatilidad y multifuncionalidad. A pesar de estos avances, la mayoría de nanomedicinas no consiguen llegar a aplicaciones médicas reales, lo cual es en parte debido a la presencia de barreras biológicas en el organismo que limitan su transporte hacia los tejidos de interés. En este sentido, el desarrollo de nuevos micro- y nanomotores sintéticos, capaces de autopropulsarse y causar cambios locales en el ambiente, podrían ofrecer una alternativa para la nanomedicina, promoviendo una mayor penetración en tejidos de interés y un mejor transporte de fármacos a través de las barreras biológicas. En concreto, los nanomotores enzimáticos poseen un alto potencial para aplicaciones biomédicas gracias a su biocompatibilidad y a la posibilidad de usar sustancias presentes en el organismo como combustible. Los trabajos presentados en esta tesis exploran el potenical de nanomotores, autopropulsados mediante la enzima ureasa, para aplicaciones biomédicas, y investigan su uso como vehículos para transporte de fármacos, su capacidad para mejorar penetración de tejidos diana, su versatilidad y movimiento colectivo. En conjunto, los resultados presentados en esta tesis doctoral demuestran el potencial del uso de nanomotores autopropulsados mediante enzimas como herramientas biomédicas, ofreciendo versatilidad en su diseño y una alta capacidad para promover el transporte de fármacos y la penetración en tumores. Por último, su movimiento colectivo observado in vivo mediante técnicas de imagen médicas representan un significativo avance en el viaje hacia su aplicación en medicina

Etude et réalisation d'un prototype avancé de plateforme de mesure de micro et nanoforce par lévitation diamagnétique

La mesure de micro- et nano-force fait partie des mesures nécessaires à la caractérisation des interactionsou des propriétés mécaniques intervenant à l échelle micrométrique. Dans cette optique,nous avons poursuivi un travail de conception initié au Laboratoire d Automatique de Besançon en2002 pour développer un prototype avancé de capteur de micro- et nano-force par lévitation diamagnétique.Le transducteur force-déplacement de ce capteur est un microcapillaire rigide en verred une dizaine de centimètres. Ce microcapillaire est en lévitation passive stable dans l espace grâceà l action conjuguée de forces magnétiques et diamagnétiques créant ainsi un ressort magnétiquevirtuel. La mesure d une force externe appliquée à l extrémité du capillaire est rendue possible grâceà la connaissance de la mesure du déplacement du capillaire et de la raideur du ressort magnétique.La plage de mesure de ce capteur varie entre +-40 N avec une résolution de l ordre du nanonewton.Les avancées présentées dans ce manuscrit ont porté sur la détermination des efforts diamagnétiquesengendrés par l utilisation des plaques de graphite. Ce travail a permis une optimisation dudesign global du dispositif et son transfert à l entreprise STIL SA. De plus un processus d estimationpar déconvolution a été développé pour tenir compte du comportement dynamique du micro capillaireet des bruits de mesure du déplacement. Les domaines d applications potentiels de ce dispositifconcernent notamment la caractérisation des interactions mécaniques quasi-statiques pouvant intervenirentre deux micro-objets et la détermination de propriétés mécaniques propres à un micro-objet.The measurement of micro- and nano-force is necessary to characterize the mechanical propertiesand interactions occurring at micrometer scale. In this context, we work on an advanced design ofmicro- and nano-force sensor based on diamagnetic levitation initiated at Laboratoire d Automatiquede Besançon in 2002. The force-displacement transducer of this sensor is a ten centimeter long glassmicro capillary. This micro capillary levitates passively and stably thanks to the combined action ofmagnetic and diamagnetic forces which create a virtual magnetic spring. Measuring an external forceapplied to the end of the capillary is made possible through knowledge of the capillary displacementand the stiffness of magnetic spring. The measuring range of the sensor varies between +-40 N witha resolution about a nanonewton. Advances presented in this thesis focused on the determinationof the diamagnetic forces generated by the use of graphite plates. This work has allowed us tooptimize the overall design of the device and transfer it to our industrial partner STIL SA. Moreovera deconvolution estimation process was developed to take into account the dynamic behavior of themicro capillary and measurement noises. Potential fields of application of this device concern themechanical characterization of quasistatic interactions which may occur between two micro-objectsand the determination of intrinsic mechanical properties of a micro-objectBESANCON-Bib. Electronique (250560099) / SudocSudocFranceF

The dynamics of phototaxis in photosynthetic microorganisms

The motility strategies of M. pusilla are characterised for the first time, establishing a new variant of run-tumble motion: stop, run or reverse. This pattern bears remarkable similarities to the run-reverse motion of marine bacteria, suggesting the size of the organism dominates the choice of motility strategy. The phototaxis of M. pusilla is then described for the first time at both population and single cell scales. The proposed method of phototaxis - an extension of the run length when the cell is orientated towards the light stimulus - is verified with a series of jump-diffusion numerical simulations. The phototactic study was extended by demonstrating the first recorded intensity dependent step-up photophobic response of M. pusilla, followed by a stronger step-down response. During these responses cells switch from typically stationary behaviour to continuous swimming to escape the harmful environment. The step-up response can also be triggered chemically during an apparent cell death, leading to burst events where cells attempt to escape from a spherically diffusing source of pollutant radiating from a single cell. The similarities in these independent responses suggest there is a global avoidance strategy present in the organism to escape from harmful environments. Finally, a new experimental system is proposed to investigate phototaxis in more complicated optical landscapes and channel confinements using the model organism C. reinhardtii. Photoaccumulation to two Gaussian stripes is observed to impede the transport through a channel, opening up the question of what influence phototaxis could have in porous media. In whole, the motility and phototactic behaviour of the most globally dominant pico-eukaryote M. pusilla has been investigated and characterised for the first time, as well the discovery of an apparently universal avoidance strategy from a variety of harmful environments