Dissertations / Theses on the topic 'Single Particle Tracking (SPT'
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Robson, Alex J. "Single particle tracking as a tool to investigate the dynamics of integrated membrane complexes in vivo." Thesis, University of Oxford, 2012. http://ora.ox.ac.uk/objects/uuid:7769f80c-a56d-4513-9123-1d65ef8c9911.
Mawoussi, Kodjo. "Effet de l'encombrement des protéines sur la diffusion des lipides et des protéines membranaires." Thesis, Paris 6, 2017. http://www.theses.fr/2017PA066541/document.
Lateral diffusion of lipids and transmembrane proteins is essential for biological functions. In the cellular context, the surface fraction of membrane proteins is high, reaching approximately 50 to 70% depending on the membrane type. Therefore, diffusion occurs in a very crowded environment. The aim of this work is to study in vitro the effect of protein crowding on their own diffusion and on those of the surrounding lipids. So far, lateral diffusion measurements generally have been carried out at low protein density, and the effect of proteins crowding has not been much studied experimentally. We used a single particle tracking (SPT) method to track the trajectories of the Bacterorhodopsin (BR) proton pump and of lipids labeled with quantum dots at the bottom of giant unilamellar vesicles (GUVs) as a function of the total surface fraction (Ф) of BR reconstituted in 1,2-Dioleoyl-sn-glycero-3-phosphocholine (DOPC) membrane
Mereghetti, Alessio. "Performance evaluation of the SPS scraping system in view of the high luminosity LHC." Thesis, University of Manchester, 2015. https://www.research.manchester.ac.uk/portal/en/theses/performance-evaluation-of-the-sps-scraping-system-in-view-of-the-high-luminosity-lhc(600579c0-0877-415d-bf8d-32896497b5ff).html.
Relich, Peter Kristopher II. "Single Particle Tracking| Analysis Techniques for Live Cell Nanoscopy." Thesis, The University of New Mexico, 2017. http://pqdtopen.proquest.com/#viewpdf?dispub=10251887.
Single molecule experiments are a set of experiments designed specifically to study the properties of individual molecules. It has only been in the last three decades where single molecule experiments have been applied to the life sciences; where they have been successfully implemented in systems biology for probing the behaviors of sub-cellular mechanisms. The advent and growth of super-resolution techniques in single molecule experiments has made the fundamental behaviors of light and the associated nano-probes a necessary concern amongst life scientists wishing to advance the state of human knowledge in biology. This dissertation disseminates some of the practices learned in experimental live cell microscopy. The topic of single particle tracking is addressed here in a format that is designed for the physicist who embarks upon single molecule studies. Specifically, the focus is on the necessary procedures to generate single particle tracking analysis techniques that can be implemented to answer biological questions. These analysis techniques range from designing and testing a particle tracking algorithm to inferring model parameters once an image has been processed. The intellectual contributions of the author include the techniques in diffusion estimation, localization filtering, and trajectory associations for tracking which will all be discussed in detail in later chapters. The author of this thesis has also contributed to the software development of automated gain calibration, live cell particle simulations, and various single particle tracking packages. Future work includes further evaluation of this laboratory's single particle tracking software, entropy based approaches towards hypothesis validations, and the uncertainty quantification of gain calibration.
Sanamrad, Arash. "Biological Insights from Single-Particle Tracking in Living Cells." Doctoral thesis, Uppsala universitet, Beräknings- och systembiologi, 2014. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-229342.
Guerrier, Mark Paul. "The development and evaluation of phosphorescent particle tracking." Thesis, University College London (University of London), 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.324887.
Woringer, Maxime. "Tools to analyze single-particle tracking data in mammalian cells." Electronic Thesis or Diss., Sorbonne université, 2019. http://www.theses.fr/2019SORUS419.
This work aims at providing tools to dissect the regulation of transcription in eukaryotic cells, with a focus on single-particle tracking of transcription factors in mammalian cells. The nucleus of an eukeryotic cell is an extremely complex medium, that contains a high concentration of macromolecules (DNA, RNA, proteins) and other small molecules (ATP, etc). How these molecules interact with transcription factors, and thus influence transcription rates is an area of intense investigations. Although some of these interactions can be captured by regular biochemistry, many of them, including weak, non-covalent interactions remain undetected by these methods. Live-cell imaging and single-particle tracking (SPT) techniques are increasingly used to characterize such effects. The inference of biophysical parameters of a given transcription factor (TF), such as its diffusion constant, the number of subpopulations or its residence time on DNA, are crucial to understanding how TF dynamics and transcription intertwine. Accurate and validated SPT analysis tools are needed. To be used by the community, SPT tools should not only be carefully validated, but also be easily accessible to non-programmers. They should also be designed to take into account known biases of the imaging techniques. In this work, we first propose a tool, accessible through a web interface, based on the modeling of the diffusion propagator. We validate it extensively and show that it exhibits state-of-the art performance. We apply this tool to two experimental settings: (1) the study of catalysis-enhanced diffusion in-vitro and (2) the analysis of the dynamics of the c-Myc transcription factor in mammalian cells
Piette, Nathalie. "Micropatterning subcellulaire pour étudier la connectivité neuronale." Electronic Thesis or Diss., Bordeaux, 2024. http://www.theses.fr/2024BORD0034.
Micropatterning was initially employed to replicate and understand the influence of the extracellular matrix on cells and some of their components. Over the past decade, subcellular printing has emerged, enabling the study of protein interactions and their role in signaling pathways as well as in the formation of synaptic, immunological, or neuronal pathways.The synaptic connection is mediated by synaptic adhesion proteins present on each side of the synapse. Due to the complexity of the synaptic environment and the lack of in vitro models to study synaptic connection in a biomimetic and controlled environment, the exact roles of these proteins in synaptogenesis remain uncertain. Subcellular protein printing presents a potential solution to address this gap. For this purpose, we have developed two biomimetic models based on protein printing: a first one using heterologous cells, providing insights into the interaction kinetics of protein pairs and linking them to their potential function. And a second one using primary neurons, allowing the formation of artificial synapses to study synaptic nano-organization during development.The protein printing system PRIMO, commercialized by Alvéole, which is co-funding this thesis, is underutilized by neuroscientists. Besides these biological objectives, the industrial aim of this thesis is to develop methodologies and proofs of concept to demonstrate the advantages and feasibility of the PRIMO technology in neuroscience.By coupling our first model, based on heterologous cells, with live-cell imaging techniques (sptPALM and FRAP), we differentiated interaction kinetics among various synaptic adhesion protein pairs and also for interactions with scaffold proteins. A labile interaction was observed for SynCAM1, known for its role in synaptic morphology. A strong and stable interaction was evident for Neuroligin1/Neurexine1β due to Neuroligin1's dimerization, which is essential for synaptic functionality.With the second model using primary hippocampal neurons, we demonstrated, in the presence of LRRTM2, the specific formation of artificial synapses. These hemi-synapses exhibited morphological and functional characteristics close to native synapses, including the presence of vesicles and spontaneous calcium activity. However, we were unable to form artificial postsynapses with Neurexine1β. Based on our observations and bibliographic analysis, we hypothesize that the postsynapse could be the initiating compartment for synaptogenesis.In conclusion, this study demonstrates: (1) that subcellular printing is an excellent model to study synaptic connectivity and adhesion from both a functional and organizational perspective. (2) That models of hemi-synapses using micropatterning are more specific than previous models. (3) That the PRIMO system opens numerous perspectives in neuroscience through its quantitative printing capabilities
Naeem, Asad. "Single and multiple target tracking via hybrid mean shift/particle filter algorithms." Thesis, University of Nottingham, 2010. http://eprints.nottingham.ac.uk/12699/.
Zelman-Femiak, Monika [Verfasser], and Gregory [Akademischer Betreuer] Harms. "Single Particle Tracking ; Membrane Receptor Dynamics / Monika Zelman-Femiak. Betreuer: Gregory Harms." Würzburg : Universitätsbibliothek der Universität Würzburg, 2012. http://d-nb.info/1026414768/34.
Spille, Jan-Hendrik [Verfasser]. "Three-dimensional single particle tracking in a light sheet microscope / Jan-Hendrik Spille." Bonn : Universitäts- und Landesbibliothek Bonn, 2014. http://d-nb.info/1052581986/34.
Ernst, Dominique [Verfasser], and Jürgen [Akademischer Betreuer] Köhler. "Single-Particle Orbit Tracking - Setup, Characterisation and Application / Dominique Ernst. Betreuer: Jürgen Köhler." Bayreuth : Universität Bayreuth, 2013. http://d-nb.info/1059353709/34.
Liu, Honghui Liu. "Using Tunable Lens to Extend the Range of the Single-Particle Tracking Microscop." The Ohio State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=osu1525655236634443.
Hävermark, Tora. "Single-particle tracking for direct measurements of Trigger Factor ribosome binding in live cells." Thesis, Uppsala universitet, Molekylärbiologi, 2021. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-445886.
Ali, Rehan. "An Automated Analysis Of Single Particle Tracking Data For Proteins That Exhibit Multi Component Motion." ScholarWorks @ UVM, 2018. https://scholarworks.uvm.edu/graddis/870.
Trenkmann, Ines, Daniela Täuber, Michael Bauer, Jörg Schuster, Sangho Bok, Shubhra Gangopadhyay, and Christian von Borczyskowski. "Investigations of solid liquid interfaces in ultra-thin liquid films via single particle tracking of silica particles." Universitätsbibliothek Leipzig, 2015. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-191734.
Trenkmann, Ines, Jörg Schuster, Shubhra Gangopadhyay, and Christian von Borczyskowski. "Investigation of solid liquid interface in ultra-thin liquid films via single particle tracking of colloidal particles." Universitätsbibliothek Leipzig, 2015. http://nbn-resolving.de/urn:nbn:de:bsz:15-qucosa-191812.
Trenkmann, Ines, Daniela Täuber, Michael Bauer, Jörg Schuster, Sangho Bok, Shubhra Gangopadhyay, and Christian von Borczyskowski. "Investigations of solid liquid interfaces in ultra-thin liquid films via single particle tracking of silica particles." Diffusion fundamentals 11 (2009) 108, S. 1-12, 2009. https://ul.qucosa.de/id/qucosa%3A14082.
Trenkmann, Ines, Jörg Schuster, Shubhra Gangopadhyay, and Christian von Borczyskowski. "Investigation of solid liquid interface in ultra-thin liquid films via single particle tracking of colloidal particles." Diffusion fundamentals 11 (2009) 115, S. 1-2, 2009. https://ul.qucosa.de/id/qucosa%3A14089.
Barry, Zachary Thomas. "Single-particle tracking and fluorescence correlation spectroscopy for systems-level analysis of molecular dynamics in diverse biological systems." Thesis, Massachusetts Institute of Technology, 2017. http://hdl.handle.net/1721.1/112494.
Cataloged from PDF version of thesis.
Includes bibliographical references.
Fluorescence microscopy has proven to be immensely powerful for the study of biological systems at both the cellular and systems biological levels. The ability to specifically label a single molecular species fluorescently has enabled the study of complex cellular structures through the visualization of their constituent components both individually as well as in context of the overall structure. Since the advent of engineered fluorescent proteins (such as GFP) and other proteins capable of being genetically encoded as fusion constructs, the utility of fluorescence microscopy has increased exponentially in terms of the ability to efficiently, specifically label desired molecules while limiting perturbations to the biology under study. With this enhanced ability of visualization came a hand-in-hand evolution of computational techniques to extract quantitative information from microscopy images. In this thesis, I focus on the application of fluorescence imaging at the biophysical level in living cells: analyzing the motion/dynamics of single molecules and complexes, which are small relative to the structures of the cell, in order to elucidate their molecular function and mechanism. The motion of these "particles" within living cells is necessarily related to their functions as well as their interacting partners, which can vary dynamically during their lifetimes. Observation and analysis of this motion using a combination of fluorescence microscopy and robust quantitative analysis allows one to infer these characteristics. Here, I study three diverse biological systems in the context of live-cell fluorescence microscopy and biophysical analysis: 1) the transport of 0-actin mRNA particles in primary mouse neurons, 2) kinetochore motion during cell division, specifically focusing on anaphase dynamics, and 3) the motion of cell-growth-implicated membrane proteins in Bacillus subtilis.
Funded by the NSF Physics of Living Systems PHY 1305537.
by Zachary Thomas Barry.
Ph. D.
Takeyama, Mao. "Convective heat transfer of saturation nucleate boiling induced by single and multi-bubble dynamics." Kyoto University, 2021. http://hdl.handle.net/2433/261621.
Sharma, Gaurav. "Direct numerical simulation of particle-laden turbulence in a straight square duct." Thesis, Texas A&M University, 2003. http://hdl.handle.net/1969.1/155.
Leung, Siu Ling. "Fabrication of Multimodal Organic-Inorganic Hybrid Nanovesicles and Study on their Intracellular Fates in Cancer Cells by Single Particle Tracking." Diss., The University of Arizona, 2013. http://hdl.handle.net/10150/312657.
Subburaj, Yamunadevi [Verfasser], and Ana J. [Akademischer Betreuer] García-Sáez. "Single Particle Tracking to Characterize the Mechanism of Pore Formation by Pore Forming Proteins / Yamunadevi Subburaj ; Betreuer: Ana J. García-Sáez." Tübingen : Universitätsbibliothek Tübingen, 2014. http://d-nb.info/1162897333/34.
Zanetti, Domingues Laura Carolina. "Single-particle tracking of the human epidermal growth factor receptors (HER) 1-3 in a breast cancer model : the effect of tyrosine kinase inhibitors." Thesis, King's College London (University of London), 2014. http://kclpure.kcl.ac.uk/portal/en/theses/singleparticle-tracking-of-the-human-epidermal-growth-factor-receptors-her-13-in-a-breast-cancer-model-the-effect-of-tyrosine-kinase-inhibitors(929e12cf-48bc-484f-b304-3f09d74203c3).html.
El, Beheiry Mohamed Hossam. "Towards whole-cell mapping of single-molecule dynamics." Thesis, Paris 6, 2015. http://www.theses.fr/2015PA066618/document.
Imaging of single molecules inside living cells confers insight to biological function at its most granular level. Single molecules experience a nanoscopic environment that is complicated, and in general, poorly understood. The modality of choice for probing this environment is live-cell localisation microscopy, where trajectories of single molecules can be captured. For many years, the great stumbling block in comprehension of physical processes at this scale was the lack of information accessible; statistical significance and robust assertions are hardly possible from a few dozen trajectories. It is the onset of high-density single-particle tracking that has dramatically reframed the possibilities of such studies. Importantly, the consequential amounts of data it provides invites the use of powerful statistical tools that assign probabilistic descriptions to experimental observations. In this thesis, Bayesian inference tools have been developed to elucidate the behaviour of single molecules via the mapping of motion parameters. As a readout, maps describe heterogeneities at local and whole-cell scales. Importantly, they grant quantitative details into basic cellular processes. This thesis uses the mapping approach to study receptor-scaffold interactions inside neurons and non-neuronal cells. A promising system in which interactions are patterned is also examined. It is shown that interactions of different types of chimeric glycine receptors to the gephyrin scaffold protein may be described and distinguished in situ. Finally, the prospects of whole-cell mapping in three-dimensions are evaluated based on a discussion of state-of-the-art volumetric microscopy techniques
Bhat, Siddharth. "Design and characterization in depleted CMOS technology for particle physics pixel detector." Thesis, Aix-Marseille, 2019. http://www.theses.fr/2019AIXM0267.
The ATLAS experiment will start operating at the High Luminosity LHC accelerator (HL-LHC) in 2026 to increase the probability of new discoveries. Depleted CMOS monolithic pixel detector technology has been one of the options considered for the outer layer of an upgraded ATLAS pixel detector and is a high potential technology for future pixel detectors. In this thesis, several prototypes have been developed using different depleted CMOS technologies, for instance, LFoundry (LF) 150 nm, TowerJazz (TJ) 180 nm and austriamicrosystems AG (AMS) 180 nm. In a high-energy environment like HL-LHC, Single Event Upsets (SEU), which become of concern for reliable circuit operation. Several test-chips in AMS, TowerJazz and LFoundry technologies with different SEU tolerant structures have been prototyped and tested. The SEU tolerant structures were designed with appropriate electronics simulations using Computer Aided Design (CAD) tools in order to study the sensitivity of injected charge to upset a memory state. An alternative powering scheme named Serial Powering scheme is foreseen for the future Inner Tracker (ITk) detector of the ATLAS experiment. To meet the requirements ofthe ATLAS experiment to the environment of a pixelated layer in a high radiation collider environment, new developments with depleted CMOS sensors have been made in Shunt-LDO regulator and sensor biasing which are designed in modified TowerJazz 180 nm CMOS imaging technology. In the TowerJazz modified process, two different voltage levels are used for the purpose of sensor depletion. The bias voltages are generated by using a negative charge pump circuit
Butler, Corey. "Quantitative single molecule imaging deep in biological samples using adaptive optics." Thesis, Bordeaux, 2017. http://www.theses.fr/2017BORD0632/document.
Optical microscopy is an indispensable tool for research in neurobiology and medicine, enabling studies of cells in their native environment. However, subcellular processes remain hidden behind the resolution limits of diffraction-limited optics which makes structures smaller than ~300nm impossible to resolve. Recently, single molecule localization (SML) and tracking has revolutionized the field, giving nanometer-scale insight into protein organization and dynamics by fitting individual fluorescent molecules to the known point spread function of the optical imaging system. This fitting process depends critically on the amount of collected light and renders SML techniques extremely sensitive to imperfections in the imaging path, called aberrations, that have limited SML to cell cultures on glass coverslips. A commercially available adaptive optics system is implemented to compensate for aberrations inherent to the microscope, and a workflow is defined for depth-dependent aberration correction that enables 3D SML in complex biological environments. A new SML technique is presented that employs a dual-objective approach to detect the emission spectrum of single molecules, enabling 5-dimensional single particle imaging and tracking (x,y,z,t,λ) without compromising spatiotemporal resolution or field of view. These acquisitions generate ~GBs of data, containing a wealth of information about the localization and environment of individual proteins. To facilitate quantitative acquisition and data analysis, the development of biochemical, software and hardware tools are presented. Together, these approaches aim to enable quantitative SML in complex biological samples
Lanoiselée, Yann. "Revealing the transport mechanisms from a single trajectory in living cells." Thesis, Université Paris-Saclay (ComUE), 2018. http://www.theses.fr/2018SACLX081/document.
This thesis is dedicated to the analysis and modeling of experiments where the position of a tracer in the cellular medium is recorded over time. The goal is to be able to extract as much information as possible from a single experimentally observed trajectory. The main challenge is to identify the transport mechanisms underlying the observed movement. The difficulty of this task lies in the analysis of individual trajectories, which requires the development of new statistical analysis tools. In the first chapter, an overview is given of the wide variety of dynamics that can be observed in the cellular medium. In particular, a review of different models of anomalous and non-Gaussian diffusion is carried out. In the second chapter, a test is proposed to reveal weak ergodicity breaking from a single trajectory. This is a generalization of the approach of M. Magdziarz and A. Weron based on the time-averaged characteristic function of the process. This new estimator is able to identify the ergodicity breaking of continuous random walking where waiting times are power law distributed. By calculating the average of the estimator for several subdiffusion models, the applicability of the method is demonstrated. In the third chapter, an algorithm is proposed to recognize the different phases of an intermittent process from a single trajectory (e.g. active/passive transport within cells, etc.).This test assumes that the process alternates between two distinct phases but does not require any hypothesis on the dynamics of each phase. Phase changes are captured by calculating quantities associated with the local convex hull (volume, diameter) evaluated along the trajectory. It is shown that this algorithm is effective in distinguishing states from a large class of intermittent processes (6 models tested). In addition, this algorithm is robust at high noise levels due to the integral nature of the convex hull. In the fourth chapter, a diffusion model in a heterogeneous medium where the diffusion coefficient evolves randomly is introduced and solved analytically. The probability density function of the displacements presents exponential tails and converges towards a Gaussian one at long time. This model generalizes previous approaches and thus makes it possible to study dynamic heterogeneities in detail. In particular, it is shown that these heterogeneities can drastically affect the accuracy of measurements made by time averages along a trajectory. In the last chapter, single-trajectory based methods are used for the analysis of two experiments. The first analysis carried out shows that the tracers exploring the cytoplasm show that the probability density of displacements has exponential tails over periods of time longer than the second. This behavior is independent of the presence of both microtubules and the actin network in the cell. The trajectories observed therefore show fluctuations in diffusivity, indicating for the first time the presence of dynamic heterogeneities within the cytoplasm. The second analysis deals with an experiment in which a set of 4mm diameter discs was vibrated vertically on a plate, inducing random motion of the disks. Through an in-depth statistical analysis, it is demonstrated that this experiment is close to a macroscopic realization of a Brownian movement. However, the probability densities of disks’ displacements show deviations from Gaussian which are interpreted as the result of inter-disk shocks. In the conclusion, the limits of the approaches adopted as well as the future research orientation opened by this thesis are discussed in detail
Stirnnagel, Kristin. "Herstellung autofluoreszierender retroviraler Partikel zur Analyse der zellulären Aufnahmemechanismen von Foamyviren." Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2016. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-84198.
Haas, Kalina. "Nanoscale co-organization of AMPAR and Neuroligin probed with single-molecule based microscopy." Thesis, Bordeaux 2, 2013. http://www.theses.fr/2013BOR22134/document.
The brain is made of complex networks of interconnected neuronal cells. All our mental activities are underlain by electrochemical signals passing through dedicated neuronal circuits. Climbing further up on the complexity ladder, information processing by neurons is performed by multiple molecules assembling and interacting together. It is well accepted that the understanding of the molecular structuring inside neuronal cells is essential to apprehend functioning of the brain. For this reason, study of the organization of the key neuronal and synaptic molecules greatly contributes to understand the mystery of the brain. AMPA receptors (AMPARs) are ionotropic glutamate receptors that play a central role in synaptic plasticity and basal synaptic transmission in the central nervous system. The distribution of AMPARs on the neuronal membrane is remarkably heterogeneous. They are organized in distinct functional aggregates, called nanodomains. Previous work demonstrate that the postsynaptic adhesion molecule Neuroligin (NRLG) anchors AMPARs through PSD-95 in the postsynaptic membrane while simultaneously forming a trans-synaptic adhesion complex with presynaptic Neurexin (NRX), and recruiting vesicular release machinery at the presynaptic site. In this way, NRLG functionally organizes synapses by recruiting post and pre-synaptic molecules essential for regulation of synaptic responses. Here we studied the effect of NRLG modulation (modification of expression level or activity) on AMPAR nano-dynamics and nano-organization at individual synapses. Our hypothesis is that the NRX-NRLG complex could be involved in the precise localization of postsynaptic receptors and their apposition with the neurotransmitter release sites in the presynaptic active zone, thus playing important role in proper signal transduction. The size of the postsynaptic density (PSD) is in the order of 500 nm, whereas the average diameter of AMPAR nanodomains 100 nm. Such small dimension necessitated the application of super-resolution microscopy techniques, whose resolution in the range of 20-40 nm is almost an order of magnitude better than diffraction limited fluorescence microscopy. Probe-based far-field fluorescence nanoscopies allow visualizing cells down to almost molecular level. To achieve my goals, I implemented different single-molecule localization nanoscopies which rely on the detection of selected populations of fluorescence probes that are separated in space and time. It was proposed that membrane trafficking of neurotransmitter receptors may contribute to modulation of synaptic efficacy. I have probed diffusional properties of AMPARs with single particle tracking, which has long been applied to probe heterogeneity of the cell membrane. Relative localization of biomolecules provides the basis for understanding their functional relationship. It is well accepted that the juxtaposition of two objects, as well as their colocalization, may give evidence of their association. With the recent developments in multi-color acquisition of single molecule and ensemble based super resolution images, it is now possible to explore the colocalization at the nanoscale between biomolecules in live and fixed cells. Despite the popularity and wide spread application of super resolution imaging, there exist only a few quantitative analysis paradigms for the colocalization of multicolor super-resolution images. Here, with the aid of conventional colocalization measurement paradigms and multivariate statistics, we analyze and report in detail the scale segregation and proximity of macromolecules within functional zones of synapses. Furthermore, we use these paradigms to evaluate fluorescent tags involved in the routine generation of single molecule based super-resolution images. We extend our analysis to elucidate in depth the co-aggregation and clustering of two key synaptic molecules, PSD95 and AMPARs, which are involved in basal synaptic organization and transmission
Gross, Linda C. M. "Applications of droplet interface bilayers : specific capacitance measurements and membrane protein corralling." Thesis, University of Oxford, 2011. http://ora.ox.ac.uk/objects/uuid:0b7ffba6-b86d-499c-a93f-3b2fc46a427b.
Gennerich, Arne. "Fluoreszenzkorrelationsspektroskopie und Rasterkorrelationsmikroskopie molekularer Prozesse in Nervenzellen." Doctoral thesis, [S.l.] : [s.n.], 2003. http://deposit.ddb.de/cgi-bin/dokserv?idn=971029008.
Haziza, Simon. "Quantification du transport intraneuronal par suivi de nanodiamants fluorescents. Application à l’étude de l’impact fonctionnel de facteurs de risque génétiques associés aux maladies neuropsychiatriques." Thesis, Université Paris-Saclay (ComUE), 2015. http://www.theses.fr/2015SACLN013/document.
The identification of molecular biomarkers of brain diseases as diverse as autism, schizophrenia and Alzheimer’s disease, is of crucial importance not only for an objective diagnosis but also to monitor response to treatments. The establishment and maintenance of sub-cellular neuronal functions, such as synaptic plasticity, are highly dependent on intracellular transport, which is essential to deliver important materials to specific locations. Abnormalities in such active transport are thought to be partly responsible for synaptic plasticity and neuronal morphology impairment found in many neuropsychiatric and neurodegenerative diseases. This thesis reports (i) the development of a quantification technic of intraneuronal transport based on fluorescent nanodiamonds (fNDs) tracking; (ii) the application of this simple and minimally invasive approach to the functional analysis of neuropsychiatric disease-related genetic variants.This manuscript falls into four chapters. The first one details the complex polygenic architecture of mental disorders and demonstrates the disease relevance of monitoring the intraneuronal transport. The second and the third chapters are dedicated to the nanodiamond-tracking assay and describe the fNDs internalisation strategies, the spatiotemporal quantitative readouts and the validation of the technic. The high brightness, the perfect photostability and the absence of cytotoxicity make fNDs a tool of choice to perform high throughput long-term bioimaging at high spatiotemporal resolution. Finally, in the fourth chapter, we apply this new functional analysis method to study the effect of genetic variants associated to autism and schizophrenia. We established transgenic mouse lines in which MARK1 and SLC25A12 genes were slightly overexpressed, and AAV-shRNA to induce AUTS2 gene haploinsufficiency. Our molecular diagnosis assay proves sufficiently sensitive to detect fine changes in intraneuronal transport dynamic, paving the way for future development in translational nanomedicine
Kohram, Maryam. "A Combined Microscopy and Spectroscopy Approach to Study Membrane Biophysics." University of Akron / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=akron1436530389.
Constals, Audrey. "Etude de la régulation glutamate dépendante de la mobilité des récepteurs AMPA et de son rôle physiologique." Thesis, Bordeaux 2, 2013. http://www.theses.fr/2013BOR22058.
AMPA receptors (AMPAR) are ionotropic glutamate receptors which are responsible for the vast majority of fast excitatory synaptic currents in fast transmission. Upon release of glutamate, AMPAR undergo three main conformational states: pore closed/agonist unbound, pore open/agonist bound and pore closed/agonist bound. Controlling the number of AMPAR and their organization in the synapse, through a combination of lateral diffusion and endo/exocytosis, is essential to regulate the intensity of synaptic transmission. The interactions between proteins of the post-synaptic density and accessory receptor proteins regulate the distribution of receptors, controlling their number and organization in the post-synapse. During my PhD, I studied the impact of AMPAR activation on their mobility and organization in the post-synapse. Indeed, the binding of glutamate to AMPAR and their following desensitization lead to major structural changes on the receptor which impacts on their interactions with scaffolding proteins and accessory proteins. The impact of such modifications on the lateral diffusion and sub-synaptic organization of AMPAR was not known yet. My findings show a mobilization of synaptic AMPAR following their activation by glutamate. At the molecular level, I suggest that the transition from the activated state to the desensitized state of AMPAR leads to a change in affinity of the receptor for their partner protein: Stargazin. This glutamate dependent regulation of AMPAR diffusion participates in maintaining the fidelity of fast synaptic transmission
Orré, Thomas. "Mécanismes moléculaires d’activation des intégrines par la kindline-2 lors de l’adhésion cellulaire." Thesis, Bordeaux, 2017. http://www.theses.fr/2017BORD0824/document.
Focal adhesions (FAs) are adhesive structures linking the cell to the extracellular matrix (ECM) and constitute molecular platforms for biochemical and mechanical signals controlling cell adhesion, migration, differentiation and survival. Integrin transmembrane receptors are core components of FAs, connecting the ECM to the actin cytoskeleton. During the early 2000s, the intracellular protein talin, which directly binds to the cytoplasmic tail of β-integrins, was considered as the main integrin activator. Nevertheless, it has been shown that kindlin, another intracellular protein that bind to β-integrin, is also a critical integrin activator. In fact, several studies have shown that kindlin and talin play complementary and synergistic roles during integrin activation. The molecular basis of these phenomena remains to determine. Moreover, most studies focusing on the role of kindlin during integrin activation and cell adhesion have been performed with suspended cells and/or with the platelet integrin αIIbβ3. Here we combined PALM microscopy with single protein tracking to decipher the role and behavior of kindlin during key molecular events occurring outside and inside FAs at the plasma membrane and leading to integrin activation, as we have done previously for talin (Rossier et al., 2012). We found that beta1 and beta3-integrins with a point mutation inhibiting binding to kindlin show reduced immobilization inside FAs. We also found that kindlin-2, which is enriched inside FAs, displayed free diffusion at the plasma membrane outside and inside FAs. This constitutes a major difference with talin, which, at the plasma membrane level, is observed almost exclusively in FAs, where it is immobile, which shows that talin is recruited into FAs directly from the cytosol without lateral diffusion along the plasma membrane (Rossier et al. 2012). To determine the molecular basis of kindlin membrane recruitment and diffusion, we used a kindlin variant known to decrease binding to integrins (kindlin-2- QW614/615AA). This mutant displayed increased membrane diffusion, suggesting that kindlin-2 can freely diffuse at the plasma membrane without interacting with integrins. Moreover, the kindlin-2-QW mutant showed decreased immobilization inside FA, showing that part of kindlin immobilization depends on interaction with integrins. This suggests that kindlin can form an immobile complex with integrins inside focal adhesions. Deletion of the kindlin pleckstrin homology (PH) domain strongly reduced the membrane recruitment and diffusion of kindlin. We assessed the functional role of kindlin membrane recruitment and diffusion by re-expressing different kindlin-2 mutants in kindlin-1/kindlin-2 double KO cells. Those experiments demonstrated that kindlin-2 membrane recruitment and diffusion are crucial for integrin activation during cell spreading and favor adhesion formation. This suggests that kindlin uses a different route from talin to reach integrins and trigger their activation, providing a possible molecular basis for their complementarity during integrin activation
Linarès-Loyez, Jeanne. "Développement de la microscopie par auto-interférences pour l'imagerie super-résolue tridimensionnelle au sein de tissus biologiques épais." Thesis, Bordeaux, 2019. http://www.theses.fr/2019BORD0167/document.
The work of this thesis was devoted to the development of a new technique SELFI (for self-interferences). This method unlocks the three-dimensional localization of individual fluorescent emitters. We have demonstrated that this allows 3D super-resolved imaging and 3D tracking of single molecules deep into dense and complex biological samples. The SELFI technique is based on the use of self-referenced interference to go back to the 3D location of a emitter in a single measurement. These interferences are generated using a diffraction grating placed at the exit of the fluorescence microscope: the fluorescence signal diffracts on the grating and, after a short propagation, the orders interfere on the detector. The formed interferences are digitally decoded to extract the 3D location of a fluorescent molecule within the sample. A single molecule can thus be localized with a precision of approximatively ten nanometers up to a depth of at least 50 µm in a thick living biological sample (for example a biological tissue).By combining the SELFI method with different super-resolution techniques (PALM, dSTORM and uPAINT), we show that this three-dimensional localization method grants the access to the hierarchy and organization of proteins in biological objects. By performing SELFI-PALM, we observed different proteins of the adhesion focal points (talin C-terminal and paxilin) and found the expected elevation differences, and those within living cell samples. These results confirm the resolution capability of the SELFI technique (about 25 nm) even for a small number of photons collected (about 500photons per molecule).We highlight the robustness of the SELFI technique by reconstructing 3D super-resolution images of dense structures at depth in complex tissue samples. By performing SELFI-dSTORM, we observed the actin network in cells grown on the surface of the coverslip at first, and at different depths (25 and 50 microns) within artificial tissues in a second time.3D single particle tracking has also been performed in living biological tissues. We observed the free diffusion of quantum dots at different depths (up to 50 microns) in living brain slices.We applied the SELFI technique to the detection of NMDA postsynaptic receptors. We observed, in primary culture of neurons but also within slices of rat brains, a difference in organization between the two subunits GluN2A and GluN2B of this glutamate receptor.Finally, we show the importance of following the evolution of the living biological sample environment during the acquisition of images leading to detections of single molecules. Thanks to the additional and simultaneous use of quantitative phase imaging, we were able to study cell membrane dynamics during the activation by a growth factor. The correlative analysis between white light quantitative phase images and single fluorescent molecule detections provides new relevant information on the sample under study
Renancio, Cédric. "Étude du trafic vésiculaire des récepteurs glutamatergiques de type AMPA : caractérisation d’une nouvelle protéine auxiliaire." Thesis, Bordeaux 2, 2013. http://www.theses.fr/2013BOR22139/document.
AMPA-type glutamate receptors (AMPAR) are the main actors of the fast excitatory synaptic transmission. Their abundance at the postsynaptic density is essential for the establishment and maintenance of synaptic function, and is the result of a highly dynamic trafficking. Many studies have characterized the membrane diffusion mechanisms involved in the AMPAR synaptic localization, and revealed the critical role of the AMPAR auxiliary proteins in the modulation of this trafficking. Furthermore, it is suggested that AMPAR synaptic localization is also regulated during the early steps of the intracellular trafficking, from the Golgi apparatus to the plasma membrane via the post-Golgi vesicles. However, the post-Golgi vesicular trafficking of AMPAR has never been visualized and therefore remains poorly understood. In collaboration with the Guus Smit team (Amsterdam), I participated in the caracterization of a novel AMPAR auxiliary protein called Shisa6. As part of this project, I studied the role of this protein on the AMPAR membrane diffusion, using a method of single particle tracking (Quantum dot) developed in the laboratory. My main thesis project was to study the post-Golgi vesicular trafficking of AMPAR through the development of a new experimental protocol. Indeed, the failure in the dynamic visualization of the receptor vesicular trafficking could be explained by a low signal/noise ratio resulting of a poor AMPAR vesicular concentration, combined with a high background noise due to receptors localized both in the endoplasmic reticulum (ER) and at the plasma membrane. In order to overcome this difficulty, we have used an ingenious tool (ARIAD system) so as to block AMPAR into the ER and, by adding a ligand, control their trafficking from the ER to the plasma membrane. Thanks to this tool we have not only significantly increased the AMPAR concentration in the post-Golgi vesicles, but also eliminated the plasma membrane background noise. The FRAP imaging technique was used in order to remove the ER background noise. Such methodological approach combined with imaging techniques in living neurons, allowed us to clearly visualize for the first time the post-Golgi vesicular trafficking of AMPAR, and to study the mechanisms involved in this trafficking
Octeau, Vivien. "Microscopie de nano-objets individuels : étude de la diffusion des intégrines dans les sites d'adhésion focales de cellules vivantes." Thesis, Bordeaux 1, 2010. http://www.theses.fr/2010BOR14047/document.
Gold nanoparticles may be detected with optical far-field microscopy by use of the photothermal effect due to their strong light absorbance. With no photophysic issues, gold nanoparticles are an alternative to fluorescent probes for use in biological systems. The PhACS method (Photothermal Absorption Correlation Spectroscopy) is used to study diffusion by measuring the autocorrelation of photothermal signal fluctuations due to nanoparticles passing through the detection volume. This method is sensitive enough to mesure the precise hydrodynamic diameter of functionalised nanoparticles. The SnaPT method (Single Nano-Particle Tracking) can track 2-dimensional motion of individual nanoparticles by pinpointing the localization with a triangulation method. The SNaPT method was used to study motion of alphaV-beta3 integrins that were bound to a 5 nm gold nanoparticle inside focal adhesion, where the cell cytoskeleton is linked to the extracullular matrix. The integrin was found to organize into clusters oscillating between the bound and diffuse states. These observations require new working models where integrins would be constantly redistributed
Salomon, Antoine. "Modélisations statistiques pour l'analyse de la diffusion des molécules et du trafic intracellulaire en microscopie de fluorescence." Electronic Thesis or Diss., Université de Rennes (2023-....), 2023. http://www.theses.fr/2023URENS125.
Fluorescence microscopy is a tool of primary interest in biomedical research as it allows to selectively visualize particle dynamics within the cell. Hence, there is a high demand for algorithmic tools capable of automatically analyzing raw microscopic data. After a presentation of the theory and techniques surrounding particle dynamics, fluorescence microscopy, tracking methods and motion classification, we present in this thesis a new mapping method based on spatiotemporal kernel estimators that robustly estimate intracellular diffusion and drift from tracking data. We evaluate it in an extensive set of experiments using simulated, real, 2D and 3D data and show that our method provides precise and accurate diffusion and drift maps while outperforming existing methods. As such, it allows biologists to study intracellular dynamics of specifically tagged particles with a wider range of acquisitions and fluorescence microscopy techniques. In addition, we present a confinement domain detection method, using particle tracks and motion classification as well
Simon, Apolline A. "Décryptage des paramètres physico-chimiques critiques favorisant la diffusion efficace des nanoparticules dans des modèles tumoraux." Electronic Thesis or Diss., Bordeaux, 2024. http://www.theses.fr/2024BORD0046.
Bioimaging of complex and heterogeneous biological environments using nanoparticles is only relevant if one controls their intrinsic and surface properties to promote their diffusion in depth. Indeed, the shape (i.e. aspect ratios, nanotubes, nanospheres), the dimension (from a few nanometers up to a few tens of nanometers), the surface charges and the surface interactions with the surrounding environment are key parameters. They regulate for instance the mobility and the future of nanoparticles inside the biological milieu, such as tumoral microenvironments. In this PhD thesis, we mainly focused on semiconducting single-walled carbon nanotubes with the aim to tailor and apply their diffusion in bio-environments by controlling their surface properties. This choice was motivated by their exceptional advantages for bio-imaging applications. Their emission wavelength is in the short-wave infrared region (SWIR), which corresponds to the second window of biological transparency. In addition, they are photostable and it has been proved that they show a high tissue penetration ability due to their nanoscale 1D morphology. To study the mobility of nanotubes in complex environments, we tracked their trajectories at the single particle level and applied super-resolution fluorescence microscopy approaches. We first detected morphological modifications associated with early-stage fibrosis on murine liver slices. To that end, we employed a correlative microscopy strategy to identify the in situ biological environment (cell membranes and nuclei) surrounding the nanotubes in addition to the study of their mobilities. This first work motivated us to explore a second strategy to suspend the nanotubes to enhance their brightness while maintaining their stealth behaviours. We investigated how changing the coating around the nanotubes (PEG molecular size or presence of an insaturation) impacted their brightness and diffusivity. Diffusion has been tested within various models with growing complexity from an agarose gel to extracts of the extracellular matrix. We distinguished two molecular sizes of PEG rising to suspensions of nanotubes suitable for our studies. Finally, with the aim of expanding the library of SWIR-emitting nanoparticles for biological imaging, we investigated another type of luminescent nanoobjects: gold nanoclusters and polymeric nanoparticles loaded with such clusters. The analysis of their luminescence as well as their potential for single particle tracking were evaluated. Single gold cluster analysis has been conducted showing excellent brightness, but only in a dried environment. In addition, the polymeric nanoparticles were shown to be detectable at the single particle level diffusing within an aqueous media constituting promising candidates for bioimaging applications
Höfer, Chris Tina. "Influenza virus assembly." Doctoral thesis, Humboldt-Universität zu Berlin, Lebenswissenschaftliche Fakultät, 2015. http://dx.doi.org/10.18452/17251.
Influenza A viruses have a segmented single-stranded RNA genome, which is packed in form of viral ribonucleoprotein (vRNP) complexes. While the viral genome is replicated and transcribed in the host cell nucleus, assembly and budding of mature virus particles take place at the apical plasma membrane. Efficient virus formation requires delivery of all viral components to this site. While intrinsic apical targeting signals of the viral transmembrane proteins have been identified, it still remains poorly understood how the viral genome is transported and targeted into progeny virus particles. In this study, potential targeting mechanisms were investigated like the ability of vRNPs to associate with lipid membranes and the intrinsic ability of the viral nucleoprotein (NP) – which is the major protein component of vRNPs – for subcellular targeting. It could be shown that vRNPs are not able to associate with model membranes in vitro, which was demonstrated by flotation of purified vRNPs with liposomes of different lipid compositions. Results indicated, however, that the matrix protein M1 can mediate binding of vRNPs to negatively charged lipid bilayers. Intrinsic subcellular targeting of NP was further investigated by expression of fluorescent NP fusion protein and fluorescence photoactivation, revealing that NP by itself does not target cytoplasmic structures. It was found to interact extensively with the nuclear compartment instead and to target specific nuclear domains with high affinity, in particular nucleoli and small interchromatin domains that frequently localized in close proximity to Cajal bodies and PML bodies. An experimental approach was finally established that allowed monitoring the transport of vRNP-like complexes in living infected cells by fluorescence detection. It was possible to perform single particle tracking and to describe different stages of vRNP transport between the nucleus and the plasma membrane. A model of three-stage transport is suggested.
Glushonkov, Oleksandr. "Imagerie de fluorescence à haute résolution : étude de la localisation nucléolaire de la protéine de la nucléocapside du VIH." Thesis, Strasbourg, 2018. http://www.theses.fr/2018STRAJ028/document.
During this experimental thesis work, we investigated the nuclear and nucleolar localization of the nucleocapsid protein (NC) of HIV-1. Previous studies performed in our laboratory evidenced a strong accumulation of NC in a subnuclear structure called nucleolus. Playing role in multiple cellular processes, nucleolus is often targeted by viruses to promote their replication. Electron microscopy revealed three nucleolar components (fibrillar centers, dense fibrillar component and granular component) associated to specific steps of the ribosome biogenesis. To characterize the distribution of the NC in these three sub-compartments and therefore shed light on the nucleolar localization of NC during the replication cycle, we developed a high-resolution optical microscopy approach. After having minimized the optical aberrations and corrected the mechanical drifts inherent to the imaging setup, the NC-mEos2 fusion protein overexpressed in HeLa cells was visualized simultaneously with immunolabeled nucleolar markers. The use of high-resolution fluorescence microscopy enabled us to resolve for the first time the three nucleolar compartments and to demonstrate the preferential localization of NC in the granular compartment of nucleolus. Finally, preliminary experiments performed with living cells showed that NC is actively transported in the nucleus and therefore may interact directly with nucleolar proteins
FANTACCI, CLAUDIO. "Distributed multi-object tracking over sensor networks: a random finite set approach." Doctoral thesis, 2015. http://hdl.handle.net/2158/1003256.
Zelman-Femiak, Monika. "Single Particle Tracking ; Membrane Receptor Dynamics." Doctoral thesis, 2011. https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-65420.
Die Einzelmolekül-Mikroskopie, das Verfolgen der Diffusion einzelner, mikroskopischer Partikel, welche an relevanten biologischen Molekülen gekoppelt sind, ist eine der entscheidenden Verfahren zur räumlichen und zeitlichen Quantifizierung der Zellsignalisierung und hat eine Genauigkeit im Nanometerbereich. Die so gewonnene Trajektorienanalyse ermöglicht nicht nur die Bestimmung der Mechanismen, die der Bewegung der Partikel zugrunde liegen, sondern liefert auch wichtige Informationen über die molekulare Wechselwirkungen, Bewegungsfreiheit und Stöchiometrie sowie über alle existierenden Subpopulationen und besondere Funktionen der einzelnen Moleküle. Die Wirksamkeit dieser Technik hängt von der Verwendung des geeigneten Flurophors und der Art seiner biochemischen Anhaftung ab. Das Ziel dieser Arbeit war die Entwicklung eines hochspezifischen Markierungsverfahrens, das zur Verwendung der Einzelmolekül-Mikroskopie für Studien im Bereich Endozytose geeignet ist und gleichzeitig eine Fluorophore-Rezeptor Stöchiometrie von 1:1 erreicht. Eine kovalente Anhaftung von Quantenpunkten an Membranrezeptoren wurde erfolgreich in einer Methode realisiert, die ACP-Systeme (Engl. Acyl-Carrier-Protein) mit Koenzym A (CoA-) funktionalisierten Quantenpunkten amalgamiert. Die notwendige Optimierung der Verwendung von Quantenpunkten mit dem Ziel einer genaueren Berechnung der Stöchiometrie von Membranproteinen sehr großer Anzahl führte zu weiteren Studien. In diesem Zusammenhang wurden Methoden zur Maximierung der Signalanzahl und Beobachtungszeiten diverser Quantenpunktentypen untersucht. Im nächsten Schritt wurden die optimierten Verfahren angewendet, um das Verhalten von IL-5Rßc (Engl. Interleukin-5 ß-common chain receptor) Rezeptoren, die endogen auf niedriger Stufe auf lebende differenzierte eosinophile-ähnlichen HL-60 Zellen existieren, zu analysieren. Die gewonnenen Daten haben gezeigt, dass die Rezeptoren sich in stabilen Oligomeren hoher Ordnung bilden, was zusätzlich mit den Ergebnissen der Analyse der Mobilität, die auf einer hohen Anzahl unterbrochener 1000-Schritt Trajektorien basiert, zwei abgegrenzte Bewegungsmuster ergab. Daraufhin wurden Methoden entwickelt, die eine Bestimmung der Stöchiometrie von Zelloberflächen-Proteinkomplexen und die Erfassung umfangreicher Trajektorien zur Bewegungsanalyse ermöglichen. Im Weiteren wurden die zuvor genannten Methoden zur genauen Überprüfung der Mobilität, Endozytose und der Charakterisierung der rückläufigen Dynamik der repräsentativen Rezeptoren von zwei verschiedenen Membranrezeptoren Klassen, des Parathormon-Rezeptors (Engl. the parathyroid hormone receptor), der zu der G-Protein-gekoppelter Rezeptor Gruppe (GPCRs) gehört und der Rezeptoren der knochenmorphogenetischen Proteine (BMPs) verwendet. BMPs aktivieren SMAD- und non-SMAD Signalkaskaden und als ein Bestandteil des TGF-β-Signalszstem sind sie in die Proliferation, die Differenyiation, die Chemotaxis und die Apoptose involviert. Zwei BMP Rezeptor Typen, BMP Typ I und BMP Typ II (BMPRI und BMPRII) sind nötig für die effektive Signalwirkung. Offenbar sind die Bewegungsmuster für BMPRI und BMPRII sehr unterschiedlich, was hier die Genauigkeit des Signals festlegt. Non-SMAD Kaskade und die nachfolgende Differenzierung von den Osteoblastenzellen benötigt das abgegrenzte Bewegungsmuster von BMPRI. Daraus folgert, dass die laterale Mobilität ein Hauptmechanismus in der SMAD gegen non-SMAD Signalwirkung während der Differenziation ist. Das abgegrenzte Bewegungsmuster war auch für den Parathormon Rezeptor (Engl. the parathyroid hormone receptor) (PTHR1), der in die Calcium Homeostase und den Knochenumbau involviert ist, in den Studien zu beobachten. In diesen Studien wurden fünf Peptide Ligande, spezifische Teile von dem PTH: hPTH(1–34), hPTHrP(107–111)NH2; PTH(1–14); PTH(1–28) G1R19, bPTH(3–34), von denen die ersten vier zu der Agonistengruppe und der Letzte zu der Antagonistengruppe gehören, in verschiedenen Konzentrationen mit lebenden COS-1 und AD293 Zellen verwendet. (oder aufgebracht) Eine der Hauptfragen war die Festlegung der Rate der PTHR1 Internalisierung und des Recycling in dieser Forschung. Im Allgemeinen reduziert Internalisierung die Stärke der Signale, die von den G Proteinen kommen und durch die Rezeptoren übermittelt (die Desensibilisierung) werden. Durch den Rücklauf werden die Rezeptoren wieder sensibilisiert, degradiert und können somit an anderen Signalkaskaden ankoppeln (zB. MAP-Kinase ). Die Determinanten der Internalisierung sind das Hauptthema in den aktuellen Studien, da sie der Schlüssel zum besseren Verständnis der Internalisierung und zu den nachfolgenden biologischen Antworten sind. Die Internalisierung von dem PTH Rezeptor verläuft entsprechend des Clathrin-coated Pit Weges mit der Teilnahme von β-arrestin2 und ist durch den Ligand eingeleitet, der zur Aktivierung von adenylyl cyclase (via Gs), und phosphatidylinositol-specific phospholipase Cβ (via Gq) führt. Zusammenfassend ist diese Arbeit unter Verwendung von Einzelmolekül-Mikroskopie mit der neuen ACP-Quantumpunktmethoden sowie standard Markierungsmethoden ein komplexes Studium über die IL-5Rßc Rezeptoren, die BMP Rezeptoren und den PTH Rezeptor
Li, Zhenning. "Single View Human Pose Tracking." Thesis, 2013. http://hdl.handle.net/10012/7543.
Chu, Jia-Yin, and 朱佳音. "Ratiometric Fluorescence pH Sensing and Single-particle Tracking in Cancer Cell." Thesis, 2012. http://ndltd.ncl.edu.tw/handle/25319646567471306974.
國立臺灣大學
化學研究所
100
The intracellular pH is an important modulator of cell functions. Since the activities of most proteins are affected by very small changes of the proton concentration, there are a number of cellular mechanisms that finely regulate the intracellular pH values (pHi). Therefore, monitoring pHi with high spatial resolution could help us to elucidate many physiological or pathogenic processes taking place within cells. We synthesized 100 nm mesoporous silica nanoparticles with a pH-sensitive dye, FITC and a pH-insensitive dye, RITC. This nanoparticle was employed to estimate the pHi by the fluorescence intensity ratio of FITC to RITC. We conjugated high amount of dyes to improve the photostability of the dyed nanoparticles. Besides, we modified the surface by post-modification to form two different nanoparticles, positively charged TA-hMSN and negatively charged THPMP-hMSN. The nanoparticles were characterized with various physical methods. Then, we used the single-particle tracking (SPT) system to examine the interaction between the nanoparticles and HeLa cells. We obtained the trajectories of nanoparticles during the uptake process and were able to estimate the fluorescence ratio in the local environment simultaneously.
Epperla, Chandra Prakash, and 艾培仁. "Single Particle Tracking and Nanothermometry of Fluorescent Nanodiamonds in Membrane Nanotubes." Thesis, 2016. http://ndltd.ncl.edu.tw/handle/t7992w.
國立清華大學
化學系
105
Fluorescent nanodiamond (FND) is a novel carbon based material that has drawn much attention in recent years due to its uniquely embedded defect centers named nitrogen-vacancy (N-V) centers. Most notable is the negatively charged nitrogen-vacancy (NV ̶ ) color center which emits a highly photostable far-red fluorescence emission. Since it does not photobleach or photoblink, it can be used to track for longer times. The FND also exhibits a very good biocompatibility and its surface can be easily functionalized through covalent or non-covalent interactions with biomolecules. The NV ̶ center has been used to sense environmental variables such as temperature and electric or magnetic fields by studying the shifts in their electronic transitions or spin transitions at electronic ground state. All these characteristics make FND a promising fluorescent probe for biological applications. Cell-to-cell communication is essential for the development and maintenance of multicellular organisms. Recently discovered membrane nanotubes (MNTs) are capable of creating intercellular communication pathways through which transport of proteins and other cytoplasmic components occurs. These cellular connections are very heterogeneous in both structure, function, and have been found to be formed in numerous cell types. MNTs are also known to participate in pathogenesis of many diseases such as Alzheimer’s, Parkinson’s and HIV. Hence, it is important to understand the dynamics of transport along these nanotubes and to explore the potential of MNTs as drug delivery channels. This doctoral thesis presents several applications of variously functionalized FNDs in membrane nanotubes. We applied protein functionalized FNDs as a photostable tracker, as well as a protein carrier, to illustrate the transport events in MNTs of human cells. Proteins, including bovine serum albumin and green fluorescent protein, were coated on 100-nm FNDs by physical adsorption. Then single-particle tracking of the bio-conjugates in the transient membrane connections was carried out by fluorescence microscopy. We observed different types of motions and velocity distribution of cargos that took takes place inside the MNTs. Our results demonstrate the promising applications of this novel carbon-based nanomaterial for intercellular delivery of biomolecular cargo down to the single-particle level. Further, we have studied the thermostability of both MNTs and cell membrane. We have developed gold nanorods (GNR) functionalized FNDs as a two-in-one optical nanodevice that can heat and sense the temperature simultaneously. We used all-optical method to study the nanothermometry of GNR-FNDs. We also demonstrated the photoporation on MNTs using GNR-FND nanohybrids to selectively deliver drugs to cytoplasm. Finally, we performed hyperlocalized hyperthermia on cell membrane for the treatment of cancer cell. During such a treatment, cancer cells can be killed selectively, while healthy cells remain unaffected. Our results demonstrate promising applications of this novel carbon-based nanomaterial for intercellular delivery of biomolecular cargo down to the single-particle level and a new paradigm for hyperthermia research and application.
Shih, Hsien-Kang, and 石憲剛. "3D Human Motion Tracking Based on Single Video Input and Particle Filtering." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/50924743055631440780.
國立中正大學
電機工程所
97
This study uses monocular video sequence as input for the tracking of humans’ 3D motion pose, enabling accurate estimation of 3D motion parameters. Our method adopts silhouette, edge, and color features for matching the input images with the projected 3D human model. We also adopts an improved annealed particle filter to accurately estimate the system states (i.e., the human 3D pose).To have the particles search in solution space more efficient, we estimate motion vectors information between consecutive frames to control the search range of the particles in every degree of freedom of the humans 3D model and the search range of the particles at every time instance. Besides that, this study also proposes several algorithms to reduce the number of particles, but still maintains the tracking accuracy. Compared with the traditional particle filter and the annealed particle filter, experiments show that our method makes particles more clustering around true solutions with higher weights, thus having better tracking results than the other two methods. Compared with the annealed particle filter, experiments also show that our method can reduce the required number of particles by about 43%, without sacrificing the tracking accuracy.