Thematische Bibliographien / Quantitative live-imaging

Inhaltsverzeichnis

  1. Zeitschriftenartikel
  2. Dissertationen
  3. Buchteile
  4. Konferenzberichte

Auswahl der wissenschaftlichen Literatur zum Thema „Quantitative live-imaging“

Autor: Grafiati
Veröffentlicht am 25. Mai 2024

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Zeitschriftenartikel zum Thema "Quantitative live-imaging"

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Tany, Ryosuke, Yuhei Goto, Yohei Kondo und Kazuhiro Aoki. „Quantitative live-cell imaging of GPCR downstream signaling dynamics“. Biochemical Journal 479, Nr. 8 (21.04.2022): 883–900. http://dx.doi.org/10.1042/bcj20220021.

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G-protein-coupled receptors (GPCRs) play an important role in sensing various extracellular stimuli, such as neurotransmitters, hormones, and tastants, and transducing the input information into the cell. While the human genome encodes more than 800 GPCR genes, only four Gα-proteins (Gαs, Gαi/o, Gαq/11, and Gα12/13) are known to couple with GPCRs. It remains unclear how such divergent GPCR information is translated into the downstream G-protein signaling dynamics. To answer this question, we report a live-cell fluorescence imaging system for monitoring GPCR downstream signaling dynamics. Genetically encoded biosensors for cAMP, Ca2+, RhoA, and ERK were selected as markers for GPCR downstream signaling, and were stably expressed in HeLa cells. GPCR was further transiently overexpressed in the cells. As a proof-of-concept, we visualized GPCR signaling dynamics of five dopamine receptors and 12 serotonin receptors, and found heterogeneity between GPCRs and between cells. Even when the same Gα proteins were known to be coupled, the patterns of dynamics in GPCR downstream signaling, including the signal strength and duration, were substantially distinct among GPCRs. These results suggest the importance of dynamical encoding in GPCR signaling.
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Grossmann, Guido, Melanie Krebs, Alexis Maizel, Yvonne Stahl, Joop E. M. Vermeer und Thomas Ott. „Green light for quantitative live-cell imaging in plants“. Journal of Cell Science 131, Nr. 2 (20.12.2017): jcs209270. http://dx.doi.org/10.1242/jcs.209270.

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Youn, Yeoan, Yongjae Lee, Gloria W. Lau und Paul R. Selvin. „Quantitative DNA-paint imaging of AMPA receptors in live neurons“. Biophysical Journal 121, Nr. 3 (Februar 2022): 141a. http://dx.doi.org/10.1016/j.bpj.2021.11.2028.

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Woehler, Andrew. „Simultaneous Quantitative Live Cell Imaging of Multiple FRET-Based Biosensors“. PLoS ONE 8, Nr. 4 (16.04.2013): e61096. http://dx.doi.org/10.1371/journal.pone.0061096.

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Lambert, Talley J., und Jennifer C. Waters. „Choosing a Fluorescence Microscopy Imaging Modality for Live Quantitative Experiments“. Microscopy and Microanalysis 20, S3 (August 2014): 2122–23. http://dx.doi.org/10.1017/s1431927614012343.

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Trivedi, Vikas, Yuwei Li, Thai V. Truong, David Koos, Chuong Cheng-Ming, Rex Moats und Scott E. Fraser. „How Embryonic Cartilage Grows: Insights Gained from Quantitative Live Imaging“. Biophysical Journal 106, Nr. 2 (Januar 2014): 575a. http://dx.doi.org/10.1016/j.bpj.2013.11.3188.

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Burgess, Andrew, Thierry Lorca und Anna Castro. „Quantitative Live Imaging of Endogenous DNA Replication in Mammalian Cells“. PLoS ONE 7, Nr. 9 (20.09.2012): e45726. http://dx.doi.org/10.1371/journal.pone.0045726.

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Rudkouskaya, Alena, Nattawut Sinsuebphon, Jamie Ward, Kate Tubbesing, Xavier Intes und Margarida Barroso. „Quantitative imaging of receptor-ligand engagement in intact live animals“. Journal of Controlled Release 286 (September 2018): 451–59. http://dx.doi.org/10.1016/j.jconrel.2018.07.032.

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Plant, Anne L., Michael Halter und Jeffrey Stinson. „Probing pluripotency gene regulatory networks with quantitative live cell imaging“. Computational and Structural Biotechnology Journal 18 (2020): 2733–43. http://dx.doi.org/10.1016/j.csbj.2020.09.025.

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Sengupta, Kheya, Eric Moyen, Magali Macé, Anne-Marie Benoliel, Anne Pierres, Frank Thibaudau, Laurence Masson, Laurent Limozin, Pierre Bongrand und Margrit Hanbücken. „Large-Scale Ordered Plastic Nanopillars for Quantitative Live-Cell Imaging“. Small 5, Nr. 4 (20.02.2009): 449–53. http://dx.doi.org/10.1002/smll.200800836.

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Dissertationen zum Thema "Quantitative live-imaging"

1

Kim, Jang-Mi. „Quantitative live imaging analysis of proneural factor dynamics during lateral inhibition in Drosophila“. Electronic Thesis or Diss., Sorbonne université, 2022. https://accesdistant.sorbonne-universite.fr/login?url=https://theses-intra.sorbonne-universite.fr/2022SORUS585.pdf.

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L’inhibition latérale par Notch est un mécanisme bien conservé au sein des espèces qui permet la formation de pattern de destins cellulaires1. Dans de nombreux tissus, la signalisation intercellulaire entre Delta et Notch coordonne dans le temps et l’espace des décisions de destin cellulaire binaires dont l’origine est proposée stochastique. Dans le contexte du développement des organes sensoriels chez la Drosophile, il a été proposé que la rupture de symétrie entre cellules équipotentes dépendait de fluctuations aléatoires dans le niveau d’expression de Delta/Notch2 (ou d’un de ses régulateurs en amont, par exemple YAP1 dans l’intestin de la souris3), avec des petites différences qui sont amplifiées et stabilisées pour générer des destins distincts. La décision cellulaire stochastique médiée par Notch peut aussi être biaisée par des facteurs intrinsèques (par exemple, l’histoire de la cellule4) ou des facteurs extrinsèques. Bien que l’inhibition latérale ait été largement étudiée dans de nombreux contextes développementaux, il manque toujours une analyse détaillée in vivo de la dynamique de l’acquisition du destin cellulaire et des signaux régulant cette décision. Ici, nous avons utilisé une approche quantitative d’imagerie en temps réel pour étudier la dynamique de spécification des organes sensoriels dans l’abdomen de la drosophile. Pour suivre la compétence des cellules à s’engager dans le destin neural et devenir une cellule précurseur des organes sensoriels (SOP), nous avons utilisé l’accumulation du facteur de transcription Scute, un régulateur majeur de la formation des organes sensoriels dans l’abdomen. Pour visionner Scute directement dans les pupes en développement, nous avons utilisé des pupes exprimant la protéine Scute taguée par une GFP. Nous avons généré des films haute résolution dans le temps et l’espace puis nous avons segmenté et traqué tous les noyaux grâce un pipeline personnalisé. Nous avons ainsi pu étudier quantitativement la dynamique de l’expression de Scute dans toutes les cellules. Après avoir défini un index de différence de destin cellulaire (FDI), nous avons trouvé que la rupture de symétrie était détectée tôt, quand les cellules exprimaient encore un niveau faible et hétérogène de Scute. Quelques rares cas de résolution tardive ont été observés c’est-à-dire quand deux cellules voisines accumulent toutes les deux un fort niveau de Scute avant d’être séparées. Il est aussi intéressant de noter que le niveau de Scute n’a pas rapidement diminué dans les cellules non sélectionnées, immédiatement après la rupture de symétrie. D’autre part, nous avons trouvé une corrélation positive entre la pente du FDI après la rupture de symétrie et l’hétérogénéité intercellulaire mesurée dans le niveau de Scute mais il reste à démontrer si l’augmentation de l’hétérogénéité est causalement liée à la rupture de symétrie. Nous avons ensuite voulu savoir si cette décision cellulaire stochastique était biaisée par l’ordre de naissance (comme proposé dans le contexte de décision AC/VU chez le C. elegans4) ou par la taille et la géométrie des contacts cellulaires (comme suggéré par une modélisation5). Nous avons trouvé qu’aucun des deux biais ne semblait influencer la décision cellulaire binaire médiée par Notch dans l’abdomen de la Drosophile. En conclusion, nos données d’imagerie fournissent une analyse quantitative détaillée de la dynamique des proneuraux pendant l’inhibition latérale chez la Drosophile
Lateral inhibition by Notch is a conserved mechanism that regulates the formation of regular patterns of cell fates1. In many tissues, intercellular Delta-Notch signaling coordinates in time and space binary fate decisions thought to be stochastic. In the context of sensory organ development in Drosophila, it has been proposed that fate symmetry breaking between equipotent cells relies on random fluctuations in the level of Delta/Notch2 (or one of their upstream regulators, e.g. YAP1 in the mouse gut3), with small differences being amplified and stabilized to generate distinct fates. Notch-mediated stochastic fate choices may also be biased by intrinsic, i.e. cell history4, or extrinsic factors. Although lateral inhibition has been extensively studied in many developmental contexts, a detailed in vivo analysis of fate and signaling dynamics is still lacking. Here, we used a quantitative live imaging approach to study the dynamics of sensory organ fate specification in the Drosophila abdomen. The accumulation of the transcription factor Scute (Sc), a key regulator of sensory organ formation in the abdomen, was used as a proxy to monitor proneural competence and SOP fate acquisition in developing pupae expressing GFP-tagged Sc. We generated high spatial and temporal resolution movies and segmented/tracked all nuclei using a custom-made pipeline. This allowed us to quantitatively study Sc dynamics in all cells. Having defined a fate difference index (FDI), we found that symmetry breaking can be detected early, when cells expressed very low and heterogeneous levels of Sc. We also observed rare cases of late fate resolution, e.g. when two cells close to each other accumulate high levels of GFP-Scute before being pulled away from each other. Interestingly, we did not observe a rapid decrease in GFP-Sc levels in non-selected cells right after symmetry breaking. Also, the rate of change of FDI values after symmetry breaking appeared to positively correlate with cell-to-cell heterogeneity in Sc levels. Whether increased heterogeneity is causally linked to symmetry breaking remains to be tested. We next addressed if this stochastic fate decision is biased by birth order (as proposed in the context of the AC/VU decision in worms4) or by the size and geometry of cell-cell contacts (as modeling suggested5). We found that neither appeared to significantly influence Notch-mediated binary fate decisions in the Drosophila abdomen. In conclusion, our live imaging data provide a detailed analysis of proneural dynamics during lateral inhibition in Drosophila
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Boni, Andrea [Verfasser], und Jan [Akademischer Betreuer] Ellenberg. „Inner nuclear membrane protein targeting studied by quantitative live cell imaging and RNAi screening / Andrea Boni ; Betreuer: Jan Ellenberg“. Heidelberg : Universitätsbibliothek Heidelberg, 2016. http://d-nb.info/1180608046/34.

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Monypenny, James Edward. „Development of quantitative live cell imaging techniques and their applications in the study of inter-cellular communication and Sarcoma cell motility“. Thesis, University College London (University of London), 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.406165.

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Lu, Yi-Ju [Verfasser], und Jane [Akademischer Betreuer] Parker. „Live-cell imaging reveals subcellular localization of plant membrane compartments during oomycete infections and quantitative high-throughput imaging identifies endocytic trafficking mutants / Yi-Ju Lu. Gutachter: Jane Parker“. Köln : Universitäts- und Stadtbibliothek Köln, 2012. http://d-nb.info/1038225981/34.

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Torrano, Adriano de Andrade [Verfasser], und CHRISTOPH [Akademischer Betreuer] BRAEUCHLE. „Quantitative live-cell imaging studies on the biological effects of nanoparticles at the cellular level / Adriano de Andrade Torrano. Betreuer: Christoph Bräuchle“. München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2015. http://d-nb.info/110469798X/34.

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Chamiolo, Jasmine. „Verbesserte FIT-Sonden für die selektive und quantitative RNA-Visualisierung in lebenden Zellen“. Doctoral thesis, Humboldt-Universität zu Berlin, 2020. http://dx.doi.org/10.18452/20889.

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In dieser Arbeit wurden die von Seitz et al. entwickelten forced intercalation (FIT)-Sonden zur mRNA-Charakterisierung in lebenden Zellen eingesetzt. Es erfolgte die Synthese verbesserter FIT-Sonden für die systematische Untersuchung der Aufnahme durch lebende Flp-In™ 293 T-REx™-Zellen. Dafür wurden sowohl hergestellte FIT-Sonden-Konjugate/Aggregate als auch kommerziell erhältliche Reagenzien, wie z.B. die Palmitinsäure und das porenbildende Enzym Streptolysin-O auf ihre Effizienz untersucht. Die optimalen Bedingungen für das Einbringen von DNA- und PNA-FIT-Sonden in Flp-In™ 293 T-REx™-Zellen lieferte das Enzym Streptolysin-O. Durch den simultanen Einsatz von drei unterschiedlichen Sonden (BO-, TO- und CB-markiert), komplementär zu drei verschiedenen Zielsequenzen, gelang es erstmals eine Dreifarben-Lebendzell-Bildgebung mit FIT-Sonden durchzuführen. Des Weiteren wurden TO-FIT-Sonden zur Unterscheidung verschiedener T-Zelllinien eingesetzt. Mithilfe eines kompetitiven Hybridisierungsexperiments konnte die spezifische Fluoreszenzemission der Sonden in den Zellen belegt werden. Untersuchungen mit zwei T-Zelllinien zeigten, dass TO-FIT-Sonden sowie terminal Cy7-markierte TO-FIT-Sonden eine erhöhte TO-Emission bei Vorhandensein der komplementären TCR-mRNA-Zielsequenz in den Zellen aufwiesen. Der terminale Cy7-Farbstoff bot mit einem zweiten Detektionskanal die Möglichkeit die Cy7-Intensität und die vorhandene TO-Intensität ins Verhältnis zu setzen, sodass Signale von ungebundener Sonde leichter ausgeschlossen werden konnten. Dies ermöglichte eine spezifische Markierung der T-Zellen. Es folgte die Synthese CB-markierter FIT-Sonden zur Aufklärung biologischer Fragestellungen, wie dem Verlauf einer Influenza A Infektion und die Synthese und Evaluation neuer Farbstoffe mit einem Absorptionsmaximum bei 590/596 nm. Zudem wurde der Einbau eines zyklischen PNA- Monomers bezüglich der Verbesserung von Responsivität und Helligkeit von PNA-FIT-Sonden analysiert.
In this work forced Intercalation (FIT) probes, developed by Seitz et al. were used for the mRNA characterization in living cells. The synthesis of improved FIT probes as well as the systematic study on the uptake of FIT probes by living Flp-In™ 293 T-REx™ cells was performed. Therefore FIT probe conjugates/aggregates as well as commercially available reagents, e.g. palmitic acid and the pore-forming enzyme Streptolysin-O were investigated under various conditions. Furthermore, the transfection was tested using an electroporator. The optimal transfection condition for the introduction of DNA and PNA FIT probes into Flp-In™ 293 T-REx™ cells was achieved using Streptolysin-O. Multicolor live cell imaging with the simultaneous use of three different FIT probes (BO, TO and QB) against three different target sequences was performed successfully. In addition, FIT probes were used for the differentiation between T cell lines. A competitive hybridization experiment with cells confirmed the specific fluorescence emission of the probes. Further studies with two cell lines and TO-FIT probes as well as terminal Cy7-labeled TO-FIT probes showed an increased TO emission in the presence of the complementary TCR mRNA target sequence in the cells. A second detection channel of the terminal Cy7 dye provided the advantage of comparing the Cy7- and TO-intensity ratio, thereby making it easier to exclude signals from unbound probe. This enabled the specific tagging of t cells. This was followed by the synthesis of QB-DNA-based FIT probes for the use in various biological applications e.g. as a pan selective marker for Influenza A infection. Moreover, the synthesis and evaluation of new dyes with an absorption maximum at 590/596 nm was performed. The incorporation of a cyclic PNA monomer next to the TO dye has also been realized to improve responsiveness and brightness in PNA-FIT probes.
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Verkhedkar, Ketki Dinesh. „Quantitative Analysis of DNA Repair and p53 in Individual Human Cells“. Thesis, Harvard University, 2012. http://dissertations.umi.com/gsas.harvard:10660.

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The goal of my research was to obtain a quantitative understanding of the mechanisms of DNA double-strand break (DSB) repair, and the activation of the tumor suppressor p53 in response to DSBs in human cells. In Chapter 2, we investigated how the kinetics of repair, and the balance between the alternate DSB repair pathways, nonhomologous end-joining (NHEJ) and homologous recombination (HR), change with cell cycle progression. We developed fluorescent reporters to quantify DSBs, HR and cell cycle phase in individual, living cells. We show that the rates of DSB repair depend on the cell cycle stage at the time of damage. We find that NHEJ is the dominant repair mechanism in G1 and in G2 cells even in the presence of a functional HR pathway. S and G2 cells use both NHEJ and HR, and higher use of HR strongly correlates with slower repair. Further, we demonstrate that the balance between NHEJ and HR changes gradually with cell cycle progression, with a maximal use of HR at the peak of active replication in mid-S. Our results establish that the presence of a sister chromatid does not affect the use of HR in human cells. Chapter 3 examines the sensitivity of the p53 pathway to DNA DSBs. We combined our fluorescent reporter for DSBs with a fluorescent reporter for p53, to quantify the level of damage and p53 activation in single cells. We find that the probability of inducing a p53 pulse increases linearly with the amount of damage. However, cancer cells do not have a distinct threshold of DSBs above which they uniformly induce p53 accumulation. We demonstrate that the decision to activate p53 is potentially controlled by cell-specific factors. Finally, we establish that the rates of DSB repair do not affect the decision to activate p53 or the dynamical properties of the p53 pulse. Collectively, this work emphasizes the importance of collecting quantitative dynamic information in single cells in order to gain a comprehensive understanding of how different DNA damage response pathways function in a coordinated manner to maintain genomic integrity.
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Wang, Renjie. „Quantitative analysis of chromatin dynamics and nuclear geometry in living yeast cells“. Thesis, Toulouse 3, 2016. http://www.theses.fr/2016TOU30122/document.

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L'analyse de l'organisation à grande échelle des chromosomes, par des approches d'imagerie et de biologie moléculaire, constitue un enjeu important de la biologie. Il est maintenant établi que l'organisation structurelle du génome est un facteur déterminant dans tous les aspects des " transactions " génomiques: transcription, recombinaison, réplication et réparation de l'ADN. Bien que plusieurs modèles aient été proposés pour décrire l'arrangement spatial des chromosomes, les principes physiques qui sous-tendent l'organisation et la dynamique de la chromatine dans le noyau sont encore largement débattus. Le noyau est le compartiment de la cellule dans lequel l'ADN chromosomique est confiné. Cependant, la mesure quantitative de l'influence de la structure nucléaire sur l'organisation du génome est délicate, principalement du fait d'un manque d'outils pour déterminer précisément la taille et la forme du noyau. Cette thèse est organisée en deux parties: le premier axe de mon projet était d'étudier la dynamique et les propriétés physiques de la chromatine dans le noyau de la levure S. cerevisiae. Le deuxième axe visait à développer des techniques pour détecter et quantifier la forme et la taille du noyau avec une grande précision. Dans les cellules de levure en croissance exponentielle, j'ai étudié la dynamique et les propriétés physiques de la chromatine de deux régions génomiques distinctes: les régions codant les ARN ribosomiques regroupés au sein d'un domaine nucléaire, le nucléole, et la chromatine du nucléoplasme. Le mouvement de la chromatine nucléoplasmique peut être modélisé par une dynamique dite de " Rouse ". La dynamique de la chromatine nucléolaire est très différente et son déplacement caractérisé par une loi de puissance d'exposant ~ 0,7. En outre, nous avons comparé le changement de la dynamique de la chromatine nucléoplasmique dans une souche sauvage et une souche porteuse d'un allèle sensible à la température (ts) permettant une inactivation conditionnelle de la transcription par l'ARN polymérase II. Les mouvements chromatiniens sont beaucoup plus importants après inactivation transcriptionnelle que dans la souche témoin. Cependant, les mouvements de la chromatine restent caractérisés par une dynamique dite de " Rouse ". Nous proposons donc un modèle biophysique prenant en compte ces résultats : le modèle de polymère dit "branched-Rouse". Dans la deuxième partie, j'ai développé "NucQuant", une méthode d'analyse d'image permettant la localisation automatique de la position de l'enveloppe nucléaire du noyau de levures. Cet algorithme comprend une correction post-acquisition de l'erreur de mesure due à l'aberration sphérique le long de l'axe Z. "NucQuant" peut être utilisée pour déterminer la géométrie nucléaire dans de grandes populations cellulaires. En combinant " NucQuant " à la technologie microfluidique, nous avons pu estimer avec précision la forme et la taille des noyaux en trois dimensions (3D) au cours du cycle cellulaire. "NucQuant" a également été utilisé pour détecter la distribution des regroupements locaux de complexes de pore nucléaire (NPCs) dans des conditions différentes, et a révélé leur répartition non homogène le long de l'enveloppe nucléaire. En particulier, nous avons pu montrer une distribution particulière sur la région de l'enveloppe en contact avec le nucléole. En conclusion, nous avons étudié les propriétés biophysiques de la chromatine, et proposé un modèle dit "branched Rouse-polymer" pour rendre compte de ces propriétés. De plus, nous avons développé "NucQuant", un algorithme d'analyse d'image permettant de faciliter l'étude de la forme et la taille nucléaire. Ces deux travaux combinés vont permettre l'étude des liens entre la géométrie du noyau et la dynamique de la chromatine
Chromosome high-order architecture has been increasingly studied over the last decade thanks to technological breakthroughs in imaging and in molecular biology. It is now established that structural organization of the genome is a key determinant in all aspects of genomic transactions. Although several models have been proposed to describe the folding of chromosomes, the physical principles governing their organization are still largely debated. Nucleus is the cell’s compartment in which chromosomal DNA is confined. Geometrical constrains imposed by nuclear confinement are expected to affect high-order chromatin structure. However, the quantitative measurement of the influence of the nuclear structure on the genome organization is unknown, mostly because accurate nuclear shape and size determination is technically challenging. This thesis was organized along two axes: the first aim of my project was to study the dynamics and physical properties of chromatin in the S. cerevisiae yeast nucleus. The second objective I had was to develop techniques to detect and analyze the nuclear 3D geomtry with high accuracy. Ribosomal DNA (rDNA) is the repetitive sequences which clustered in the nucleolus in budding yeast cells. First, I studied the dynamics of non-rDNA and rDNA in exponentially growing yeast cells. The motion of the non-rDNA could be modeled as a two-regime Rouse model. The dynamics of rDNA was very different and could be fitted well with a power law of scaling exponent ~0.7. Furthermore, we compared the dynamics change of non-rDNA in WT strains and temperature sensitive (TS) strains before and after global transcription was actived. The fluctuations of non-rDNA genes after transcriptional inactivation were much higher than in the control strain. The motion of the chromatin was still consistent with the Rouse model. We propose that the chromatin in living cells is best modeled using an alternative Rouse model: the “branched Rouse polymer”. Second, we developed “NucQuant”, an automated fluorescent localization method which accurately interpolates the nuclear envelope (NE) position in a large cell population. This algorithm includes a post-acquisition correction of the measurement bias due to spherical aberration along Z-axis. “NucQuant” can be used to determine the nuclear geometry under different conditions. Combined with microfluidic technology, I could accurately estimate the shape and size of the nuclei in 3D along entire cell cycle. “NucQuant” was also utilized to detect the distribution of nuclear pore complexes (NPCs) clusters under different conditions, and revealed their non-homogeneous distribution. Upon reduction of the nucleolar volume, NPCs are concentrated in the NE flanking the nucleolus, suggesting a physical link between NPCs and the nucleolar content. In conclusion, we have further explored the biophysical properties of the chromatin, and proposed that chromatin in the nucleoplasm can be modeled as "branched Rouse polymers". Moreover, we have developed “NucQuant”, a set of computational tools to facilitate the study of the nuclear shape and size. Further analysis will be required to reveal the links between the nucleus geometry and the chromatin dynamics
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Kawahira, Naofumi. „Quantitative analysis of 3D tissue deformation reveals key cellular mechanism associated with initial heart looping“. Kyoto University, 2020. http://hdl.handle.net/2433/254507.

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Chen, Huiyi. „System-Wide Studies of Gene Expression in Escherichia coli by Fluorescence Microscopy and High Throughput Sequencing“. Thesis, Harvard University, 2011. http://dissertations.umi.com/gsas.harvard:10044.

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Gene expression is a fundamental process in the cell and is made up of two parts – the information flow from DNA to RNA, and from RNA to protein. Here, we examined specific sub-processes in Escherichia coli gene expression using newly available tools that permit genome-wide analysis. We begin our studies measuring mRNA and protein abundances in single cells by single-molecule fluorescence microscopy, and then focus our attention to studying RNA generation and degradation by high throughput sequencing. The details of the dynamics of gene expression can be observed from fluctuations in mRNA and protein copy numbers in a cell over time, or the variations in copy numbers in an isogenic cell population. We constructed a yellow fluorescent fusion protein library in E. coli and measured protein and mRNA abundances in single cells. At below ten proteins per cell, a simple model of gene expression is sufficient to explain the observed distributions. At higher expression levels, the distributions are dominated by extrinsic noise, which is the systematic heterogeneity between cells. Unlike proteins which can be stable over many hours, mRNA is made and degraded on the order of minutes in E. coli. To measure the dynamics of RNA generation and degradation, we developed a protocol using high throughput sequencing to measure steady-state RNA abundances, RNA polymerase elongation rates and RNA degradation rates simultaneously with high nucleotide-resolution genome-wide. Our data shows that RNA has similar lifetime at all positions throughout the length of the transcript. We also find that our polymerase elongation rates measured in vivo on a chromosome are generally slower than rates measured on plasmids by other groups. Studying nascent RNA will allow further understanding of RNA generation and degradation. To this end, we have developed a labeling protocol with a nucleoside analog that is compatible with high throughput sequencing.
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Buchteile zum Thema "Quantitative live-imaging"

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Le Marois, Alix, und Klaus Suhling. „Quantitative Live Cell FLIM Imaging in Three Dimensions“. In Advances in Experimental Medicine and Biology, 31–48. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/978-3-319-67358-5_3.

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Schöler, Ulrike, Anna-Lena Merten, Sebastian Schürmann und Oliver Friedrich. „Quantitative Live-Cell Ca2+ Imaging During Isotropic Cell Stretch“. In Methods in Molecular Biology, 155–76. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-3052-5_10.

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3

Kemper, Björn, und Jürgen Schnekenburger. „Digital Holographic Microscopy for Quantitative Live Cell Imaging and Cytometry“. In Advanced Optical Flow Cytometry, 211–37. Weinheim, Germany: Wiley-VCH Verlag GmbH & Co. KGaA, 2011. http://dx.doi.org/10.1002/9783527634286.ch8.

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4

Rakymzhan, Asylkhan, Helena Radbruch und Raluca A. Niesner. „Quantitative Imaging of Ca2+ by 3D–FLIM in Live Tissues“. In Advances in Experimental Medicine and Biology, 135–41. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/978-3-319-67358-5_9.

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5

Kaminski, Clemens F., Eric J. Rees und Gabriele S. Kaminski Schierle. „A Quantitative Protocol for Intensity-Based Live Cell FRET Imaging“. In Methods in Molecular Biology, 445–54. Totowa, NJ: Humana Press, 2013. http://dx.doi.org/10.1007/978-1-62703-649-8_19.

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6

Marquet, Pierre, Benjamin Rappaz und Nicolas Pavillon. „Quantitative Phase-Digital Holographic Microscopy: A New Modality for Live Cell Imaging“. In New Techniques in Digital Holography, 169–217. Hoboken, NJ, USA: John Wiley & Sons, Inc., 2015. http://dx.doi.org/10.1002/9781119091745.ch5.

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Jelcic, Mark, Balázs Enyedi und Philipp Niethammer. „Quantitative Imaging of Endogenous and Exogenous H2O2 Gradients in Live Zebrafish Larvae“. In Methods in Molecular Biology, 283–99. New York, NY: Springer New York, 2019. http://dx.doi.org/10.1007/978-1-4939-9424-3_17.

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8

Kemper, Björn, Patrik Langehanenberg, Sebastian Kosmeier, Frank Schlichthaber, Christian Remmersmann, Gert von Bally, Christina Rommel, Christian Dierker und Jürgen Schnekenburger. „Digital Holographic Microscopy: Quantitative Phase Imaging and Applications in Live Cell Analysis“. In Handbook of Coherent-Domain Optical Methods, 215–57. New York, NY: Springer New York, 2012. http://dx.doi.org/10.1007/978-1-4614-5176-1_6.

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9

Amin, Viren, Mercedes Izquirdo, Doyle Wilson, Gene Rouse und Ronald Roberts. „Ultrasonic Evaluation of Quality Attributes in Live Beef Animals Using Real-Time B-Mode Ultrasound Imaging“. In Review of Progress in Quantitative Nondestructive Evaluation, 1329–34. Boston, MA: Springer US, 1996. http://dx.doi.org/10.1007/978-1-4613-0383-1_173.

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Badal, Kerriann, Yibo Zhao, Kyle E. Miller und Sathyanarayanan V. Puthanveettil. „Live Imaging and Quantitative Analysis of Organelle Transport in Sensory Neurons of Aplysia Californica“. In Methods in Molecular Biology, 23–48. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-1990-2_2.

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Konferenzberichte zum Thema "Quantitative live-imaging"

1

Wax, Adam. „Phase imaging of mechanical properties of live cells (Conference Presentation)“. In Quantitative Phase Imaging III, herausgegeben von Gabriel Popescu und YongKeun Park. SPIE, 2017. http://dx.doi.org/10.1117/12.2255903.

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2

Choi, Wonshik, Yongjin Sung, Yongkeun Park, Christopher Fang-Yen, Kamran Badizadegan, Ramachandra R. Dasari und Michael S. Feld. „Quantitative live cell imaging with tomographic phase microscopy“. In Novel Techniques in Microscopy. Washington, D.C.: OSA, 2009. http://dx.doi.org/10.1364/ntm.2009.ntua1.

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3

Shu, Xin, Yi Zhang, Mengxuan Niu, Wei Luo und Renjie Zhou. „Compact and ease-of-use quantitative phase microscopy for real-time live-cell imaging“. In Quantitative Phase Imaging VIII, herausgegeben von Gabriel Popescu, YongKeun Park und Yang Liu. SPIE, 2022. http://dx.doi.org/10.1117/12.2610489.

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4

Slabý, Tomáš, Aneta Křížová, Martin Lošt'ák, Jana Čolláková, Veronika Jůzová, Pavel Veselý und Radim Chmelík. „Coherence-controlled holographic microscopy for live-cell quantitative phase imaging“. In SPIE BiOS, herausgegeben von Gabriel Popescu und YongKeun Park. SPIE, 2015. http://dx.doi.org/10.1117/12.2080128.

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5

Kemper, Bjorn. „Multi-parameter quantitative live cell imaging with digital holographic microscopy“. In 2013 IEEE Photonics Conference (IPC). IEEE, 2013. http://dx.doi.org/10.1109/ipcon.2013.6656487.

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6

Saintoyant, Anaïs, Sherazade Aknoun, Fabrice Valentino, Antoine Federici und Benoit Wattellier. „High-Definition Quantitative Phase Imaging System applied to live cell samples“. In Digital Holography and Three-Dimensional Imaging. Washington, D.C.: OSA, 2019. http://dx.doi.org/10.1364/dh.2019.th4a.6.

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7

Pandiyan, Vimal Prabhu, und Renu John. „Quantitative Phase imaging of Live Yeast cells using Digital Holographic Microscopy“. In International Conference on Fibre Optics and Photonics. Washington, D.C.: OSA, 2014. http://dx.doi.org/10.1364/photonics.2014.s5a.4.

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8

Zuo, Chao, Yefeng Shu und Jiasong Sun. „Adaptive optical quantitative phase imaging with annular illumination Fourier ptychographic microscopy“. In Digital Holography and Three-Dimensional Imaging. Washington, D.C.: Optica Publishing Group, 2022. http://dx.doi.org/10.1364/dh.2022.w7a.3.

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Annotation:
We propose an adaptive optical QPI method to solve time-varying aberrations in long-term imaging based on Fourier ptychographic microscopy. Only a few images captured under the annular matched illumination are required to realize the aberration-free live-cell observation.
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9

Ling, Zhi, Keyi Han, Wenhao Liu, Xuanwen Hua und Shu Jia. „Volumetric Live-Cell Autofluorescence Imaging Using Fourier Light-Field Microscopy“. In Frontiers in Optics. Washington, D.C.: Optica Publishing Group, 2023. http://dx.doi.org/10.1364/fio.2023.jm4a.79.

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This study introduces Fourier light-field microscopy technique for rapid, volumetric live-cell autofluorescence imaging. The method enables multicolor imaging and quantitative analysis of lysosomal and mitochondrial interactions, facilitating insights into native cellular states with minimized photodamage.
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10

Pandiyan, Vimal Prabhu, Kedar Khare und Renu John. „High resolution quantitative phase imaging of live cells with constrained optimization approach“. In SPIE BiOS, herausgegeben von Gabriel Popescu und YongKeun Park. SPIE, 2016. http://dx.doi.org/10.1117/12.2209289.

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