Relevant bibliographies by topics / Fluorescence Recovery while photobleaching

Academic literature on the topic 'Fluorescence Recovery while photobleaching'

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Published: 10 December 2022
Last updated: 28 January 2023

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Journal articles on the topic "Fluorescence Recovery while photobleaching"

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Combs, Christian A., and Robert S. Balaban. "Enzyme-Dependant Fluorescence Recovery After Photobleaching (ED-FRAP): Application to Imaging Dehydrogenase Activity in Living Single Cells." Microscopy and Microanalysis 7, S2 (August 2001): 18–19. http://dx.doi.org/10.1017/s1431927600026167.

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Fluorescent recovery from photobleaching coupled with confocal microscopy was explored as a potential high-resolution method of imaging the distribution of enzyme activity in single living cardiac myocytes without relying on steady state measurements of fluorescence. On a fundamental level, much remains to be determined regarding how local conditions within a cell affect metabolism. Many studies have suggested that energy metabolism in muscle cells cannot be accurately described assuming a homogenous system of enzymatic reactions [1,2]. The autofluorescence of NADH has been used in many studies as a quantitative assay of mitochondrial energy metabolism [3,4], but studies of steady state fluorescence cannot distinguish between changes in energy production or utilization.In this study conditions were created where the fluorescent recovery of a probe would be solely dependent on cellular enzymatic activity (Enzyme Dependent Fluorescence Recovery after Photobleaching (ED-FRAP)). Experiments examining the inherent fluorescence of NADH (351nm excitation, 450 nm emission) were conducted on small droplets (less than 325 μm diameter) containing NADH alone, in droplets containing an enzyme system capable of synthesis of NADH (Figure 1A) and on isolated rabbit cardiac myocytes (Figure 1D). Photobleaching of the entire cell or droplet eliminated diffusion or bulk transport of NADH from non-bleached regions. Droplets containing NADH alone did not recover, while droplets containing enzyme were shown to recover exponentially (Figure 1B) with a rate constant of fluorescent recovery (kf) that was proportional to enzyme concentration (Figure 1C).
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Wüstner, Daniel. "Dynamic Mode Decomposition of Fluorescence Loss in Photobleaching Microscopy Data for Model-Free Analysis of Protein Transport and Aggregation in Living Cells." Sensors 22, no. 13 (June 23, 2022): 4731. http://dx.doi.org/10.3390/s22134731.

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The phase separation and aggregation of proteins are hallmarks of many neurodegenerative diseases. These processes can be studied in living cells using fluorescent protein constructs and quantitative live-cell imaging techniques, such as fluorescence recovery after photobleaching (FRAP) or the related fluorescence loss in photobleaching (FLIP). While the acquisition of FLIP images is straightforward on most commercial confocal microscope systems, the analysis and computational modeling of such data is challenging. Here, a novel model-free method is presented, which resolves complex spatiotemporal fluorescence-loss kinetics based on dynamic-mode decomposition (DMD) of FLIP live-cell image sequences. It is shown that the DMD of synthetic and experimental FLIP image series (DMD-FLIP) allows for the unequivocal discrimination of subcellular compartments, such as nuclei, cytoplasm, and protein condensates based on their differing transport and therefore fluorescence loss kinetics. By decomposing fluorescence-loss kinetics into distinct dynamic modes, DMD-FLIP will enable researchers to study protein dynamics at each time scale individually. Furthermore, it is shown that DMD-FLIP is very efficient in denoising confocal time series data. Thus, DMD-FLIP is an easy-to-use method for the model-free detection of barriers to protein diffusion, of phase-separated protein assemblies, and of insoluble protein aggregates. It should, therefore, find wide application in the analysis of protein transport and aggregation, in particular in relation to neurodegenerative diseases and the formation of protein condensates in living cells.
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DeBiasio, R. L., L. L. Wang, G. W. Fisher, and D. L. Taylor. "The dynamic distribution of fluorescent analogues of actin and myosin in protrusions at the leading edge of migrating Swiss 3T3 fibroblasts." Journal of Cell Biology 107, no. 6 (December 1, 1988): 2631–45. http://dx.doi.org/10.1083/jcb.107.6.2631.

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The formation of protrusions at the leading edge of the cell is an essential step in fibroblast locomotion. Using fluorescent analogue cytochemistry, ratio imaging, multiple parameter analysis, and fluorescence photobleaching recovery, the distribution of actin and myosin was examined in the same protrusions at the leading edge of live, locomoting cells during wound-healing in vitro. We have previously defined two temporal stages of the formation of protrusions: (a) initial protrusion and (b) established protrusion (Fisher et al., 1988). Actin was slightly concentrated in initial protrusions, while myosin was either totally absent or present at extremely low levels at the base of the initial protrusions. In contrast, established protrusions contained diffuse actin and actin microspikes, as well as myosin in both diffuse and structured forms. Actin and myosin were also localized along concave transverse fibers near the base of initial and established protrusions. The dynamics of myosin penetration into a relatively stable, established protrusion was demonstrated by recording sequential images over time. Myosin was shown to be absent from an initial protrusion, but diffuse and punctate myosin was detected in the same protrusion within 1-2 min. Fluorescence photobleaching recovery indicated that myosin was 100% immobile in the region behind the leading edge containing transverse fibers, in comparison to the 21% immobile fraction detected in the perinuclear region. Possible explanations of the delayed penetration of myosin into established protrusions and the implications on the mechanism of protrusion are discussed.
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Gorbsky, G. J., and G. G. Borisy. "Microtubules of the kinetochore fiber turn over in metaphase but not in anaphase." Journal of Cell Biology 109, no. 2 (August 1, 1989): 653–62. http://dx.doi.org/10.1083/jcb.109.2.653.

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In previous work we injected mitotic cells with fluorescent tubulin and photobleached them to mark domains on the spindle microtubules. We concluded that chromosomes move poleward along kinetochore fiber microtubules that remain stationary with respect to the pole while depolymerizing at the kinetochore. In those experiments, bleached zones in anaphase spindles showed some recovery of fluorescence with time. We wished to determine the nature of this recovery. Was it due to turnover of kinetochore fiber microtubules or of nonkinetochore microtubules or both? We also wished to investigate the question of turnover of kinetochore microtubules in metaphase. We microinjected cells with x-rhodamine tubulin (x-rh tubulin) and photobleached spindles in anaphase and metaphase. At various times after photobleaching, cells were detergent lysed in a cold buffer containing 80 microM calcium, conditions that led to the disassembly of almost all nonkinetochore microtubules. Quantitative analysis with a charge coupled device image sensor revealed that the bleached zones in anaphase cells showed no fluorescence recovery, suggesting that these kinetochore fiber microtubules do not turn over. Thus, the partial fluorescence recovery seen in our earlier anaphase experiments was likely due to turnover of nonkinetochore microtubules. In contrast fluorescence in metaphase cells recovered to approximately 70% the control level within 7 min suggesting that many, but perhaps not all, kinetochore fiber microtubules of metaphase cells do turn over. Analysis of the movements of metaphase bleached zones suggested that a slow poleward translocation of kinetochore microtubules occurred. However, within the variation of the data (0.12 +/- 0.24 micron/min), it could not be determined whether the apparent movement was real or artifactual.
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Kamioka, Hiroshi, Yoshihito Ishihara, Hans Ris, Sakhr A. Murshid, Yasuyo Sugawara, Teruko Takano-Yamamoto, and Soo-Siang Lim. "Primary Cultures of Chick Osteocytes Retain Functional Gap Junctions between Osteocytes and between Osteocytes and Osteoblasts." Microscopy and Microanalysis 13, no. 2 (February 15, 2007): 108–17. http://dx.doi.org/10.1017/s143192760707016x.

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The inaccessibility of osteocytes due to their embedment in the calcified bone matrix in vivo has precluded direct demonstration that osteocytes use gap junctions as a means of intercellular communication. In this article, we report successfully isolating primary cultures of osteocytes from chick calvaria, and, using anti-connexin 43 immunocytochemistry, demonstrate gap junction distribution to be comparable to that found in vivo. Next, we demonstrate the functionality of the gap junctions by (1) dye coupling studies that showed the spread of microinjected Lucifer Yellow from osteoblast to osteocyte and between adjacent osteocytes and (2) analysis of fluorescence replacement after photobleaching (FRAP), in which photobleaching of cells loaded with a membrane-permeable dye resulted in rapid recovery of fluorescence into the photobleached osteocyte, within 5 min postbleaching. This FRAP effect did not occur when cells were treated with a gap junction blocker (18α-glycyrrhetinic acid), but replacement of fluorescence into the photobleached cell resumed when it was removed. These studies demonstrate that gap junctions are responsible for intercellular communication between adjacent osteocytes and between osteoblasts and osteocytes. This role is consistent with the ability of osteocytes to respond to and transmit signals over long distances while embedded in a calcified matrix.
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Kindermann, Stefan, and Štěpán Papáček. "On Data Space Selection and Data Processing for Parameter Identification in a Reaction-Diffusion Model Based on FRAP Experiments." Abstract and Applied Analysis 2015 (2015): 1–17. http://dx.doi.org/10.1155/2015/859849.

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Fluorescence recovery after photobleaching (FRAP) is a widely used measurement technique to determine the mobility of fluorescent molecules within living cells. While the experimental setup and protocol for FRAP experiments are usually fixed, data (pre)processing represents an important issue. The aim of this paper is twofold. First, we formulate and solve the problem ofrelevantFRAP data selection. The theoretical findings are illustrated by the comparison of the results of parameter identification when the full data set was used and the case when theirrelevant data set(data with negligible impact on the confidence interval of the estimated parameters) was removed from the data space. Second, we analyze and compare two approaches of FRAP data processing. Our proposition, surprisingly for the FRAP community, claims that the data set represented by the FRAP recovery curves in form of a time series (integrated data approachcommonly used by the FRAP community) leads to a larger confidence interval compared to thefull(spatiotemporal)data approach.
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Waterman-Storer, C. M., and W. C. Salmon. "Fluorescent Speckle Microscopy in Studies of Cytoskeletal Dynamics During Cell Motility." Microscopy and Microanalysis 7, S2 (August 2001): 6–7. http://dx.doi.org/10.1017/s1431927600026106.

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We have discovered a new method, Fluorescent Speckle Microscopy (FSM), for analyzing the dynamic movement and turnover of macromolecular protein assemblies, such as the cytoskeleton, in living cells (Waterman-Storer et al., 1998). FSM compliments or replaces the techniques of fluorescence recovery after photobleaching or photoactivation of fluorescence for measuring protein dynamics in vivo. For FSM, cells are microinjected with a very low fraction of fluorescently labeled subunits that co-assemble with unlabeled subunits to give a structure with a fluorescent speckled appearance in diffraction-limited wide-field or confocal digital fluorescence images. At low fractions of fluorescent subunits relative to unlabeled subunits, fluorescent speckles vary randomly in intensity according to the number of fluorescent subunits within a diffraction-limited region. in time-lapse FSM image series, movement of the fluorescent speckle pattern indicates translocation of structures, while changes in speckle intensity indicate subunit turnover. We have used FSM to study microtubule and actin behavior in interphase and mitotic cells. We use kymograph analysis to quantitate the movement of speckled structures (Fig 1) and are currently developing analysis procedures to quantify subunit turnover in structures.We have applied these methods to the study of microtubule and actin cytoskeletal dynamics in migrating vertebrate cells in culture. Interactions between the microtubule and actin cytoskeletons underlie fundamental cellular processes such as cytokinesis and cell locomotion, but are poorly understood.
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Riquelme, Meritxell, Salomon Bartnicki-García, Juan Manuel González-Prieto, Eddy Sánchez-León, Jorge A. Verdín-Ramos, Alejandro Beltrán-Aguilar, and Michael Freitag. "Spitzenkörper Localization and Intracellular Traffic of Green Fluorescent Protein-Labeled CHS-3 and CHS-6 Chitin Synthases in Living Hyphae of Neurospora crassa." Eukaryotic Cell 6, no. 10 (July 20, 2007): 1853–64. http://dx.doi.org/10.1128/ec.00088-07.

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ABSTRACT The subcellular location and traffic of two selected chitin synthases (CHS) from Neurospora crassa, CHS-3 and CHS-6, labeled with green fluorescent protein (GFP), were studied by high-resolution confocal laser scanning microscopy. While we found some differences in the overall distribution patterns and appearances of CHS-3-GFP and CHS-6-GFP, most features were similar and were observed consistently. At the hyphal apex, fluorescence congregated into a conspicuous single body corresponding to the location of the Spitzenkörper (Spk). In distal regions (beyond 40 μm from the apex), CHS-GFP revealed a network of large endomembranous compartments that was predominantly comprised of irregular tubular shapes, while some compartments were distinctly spherical. In the distal subapex (20 to 40 μm from the apex), fluorescence was observed in globular bodies that appeared to disintegrate into vesicles as they advanced forward until reaching the proximal subapex (5 to 20 μm from the apex). CHS-GFP was also conspicuously found delineating developing septa. Analysis of fluorescence recovery after photobleaching suggested that the fluorescence of the Spk originated from the advancing population of microvesicles (chitosomes) in the subapex. The inability of brefeldin A to interfere with the traffic of CHS-containing microvesicles and the lack of colocalization of CHS-GFP with the endoplasmic reticulum (ER)-Golgi body fluorescent dyes lend support to the idea that CHS proteins are delivered to the cell surface via an alternative route distinct from the classical ER-Golgi body secretory pathway.
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Salas, P. J., D. E. Vega-Salas, J. Hochman, E. Rodriguez-Boulan, and M. Edidin. "Selective anchoring in the specific plasma membrane domain: a role in epithelial cell polarity." Journal of Cell Biology 107, no. 6 (December 1, 1988): 2363–76. http://dx.doi.org/10.1083/jcb.107.6.2363.

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We have studied the role of restrictions to lateral mobility in the segregation of proteins to apical and basolateral domains of MDCK epithelial cells. Radioimmunoassay and semiquantitative video analysis of immunofluorescence on frozen sections showed that one apical and three basolateral glycoproteins, defined by monoclonal antibodies and binding of beta-2-microglobulin, were incompletely extracted with 0.5% Triton X-100 in a buffer that preserves the cortical cytoskeleton (Fey, E. G., K. M. Wan, and S. Penman. 1984. J. Cell Biol. 98:1973-1984; Nelson, W. T. and P. J. Veshnock. 1986. J. Cell Biol. 103:1751-1766). The marker proteins were preferentially extracted from the "incorrect" domain (i.e., the apical domain for a basolateral marker), indicating that the cytoskeletal anchoring was most effective on the "correct" domain. The two basolateral markers were unpolarized and almost completely extractable in cells prevented from establishing cell-cell contacts by incubation in low Ca++ medium, while an apical marker was only extracted from the basal surface under the same conditions. Procedures were developed to apply fluorescent probes to either the apical or the basolateral surface of live cells grown on native collagen gels. Fluorescence recovery after photobleaching of predominantly basolateral antigens showed a large percent of cells (28-52%) with no recoverable fluorescence on the basal domain but normal fluorescence recovery on the apical surface of most cells (92-100%). Diffusion coefficients in cells with normal fluorescence recovery were in the order of 1.1 x 10(-9) cm2/s in the apical domain and 0.6-0.9 x 10(-9) cm2/s in the basal surface, but the difference was not significant. The data from both techniques indicate (a) the existence of mobile and immobile protein fractions in both plasma membrane domains, and (b) that linkage to a domain specific submembrane cytoskeleton plays an important role in the maintenance of epithelial cell surface polarity.
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Wu, Yin, Hisaya Kawate, Keizo Ohnaka, Hajime Nawata, and Ryoichi Takayanagi. "Nuclear Compartmentalization of N-CoR and Its Interactions with Steroid Receptors." Molecular and Cellular Biology 26, no. 17 (September 1, 2006): 6633–55. http://dx.doi.org/10.1128/mcb.01534-05.

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ABSTRACT The repression mechanisms by the nuclear receptor corepressor (N-CoR) of steroid hormone receptor (SHR)-mediated transactivation were examined. Yellow fluorescent protein (YFP)-N-CoR was distributed as intranuclear discrete dots, while coexpression of androgen receptor (AR), glucocorticoid receptor α, and estrogen receptor α ligand-dependently triggered redistribution of YFP-N-CoR. In fluorescence recovery after photobleaching analysis, mobility of the N-CoR was reduced by 5α-dihydrotestosterone (DHT)-bound AR. The middle region of N-CoR mostly contributed to the interaction with agonist-bound SHRs and the suppression of their transactivation function. N-CoR impaired the DHT-induced N-C interaction of AR, and the impaired interaction was dose-dependently recovered by coexpression of SRC-1 and CBP. N-CoR also impaired the intranuclear complete (distinct) focus formation of SHRs. Coexpression of SRC-1 or CBP released YFP-N-CoR or endogenous N-CoR from incomplete foci and simultaneously recovered complete foci of AR-green fluorescent protein. These results indicate that the relative ratio of coactivators and corepressors determines the conformational equilibrium between transcriptionally active and inactive SHRs in the presence of agonists. The intranuclear foci formed by agonist-bound SHRs were completely destroyed by actinomycin D and α-amanitin, indicating that the focus formation does not precede the transcriptional activation. The focus formation may reflect the accumulation of SHR/coactivator complexes released from the transcriptionally active sites and thus be a mirror of transcriptionally active complex formation.
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Dissertations / Theses on the topic "Fluorescence Recovery while photobleaching"

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Gousseva, Veronika. "A quantitative fluorescence recovery after photobleaching analysis of receptor-protein interactions on vesicles /." Thesis, McGill University, 2006. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=101130.

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In a living cell, signals are usually transmitted from outside to the intracellular machinery through receptors on the cell membrane. Once bound to their ligand (e.g. a hormone or another protein), receptors become activated and trigger chemical reactions inside the cell. To stop the response, receptors are often removed from the cell membrane (internalization). A protein called arrestin binds to the receptor and targets it to a vesicle, which detaches from the membrane and fuses with other vesicles to form an endosome, a sphere-like structure containing many receptors. After internalization, arrestin must dissociate from the receptor before the latter returns to the membrane or is degraded. By using fluorescence recovery after photobleaching data and a reaction-diffusion model, we estimate the in vivo affinity of arrestin for bradykinin type 2 receptor. We use the Hough transform to automate fluorescence quantification and a Bayesian approach to fit the model to the data.
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Rodriguez-Enriquez, Ricardo. "Analysis of Bcl-2 family protein interactions in live cells by fluorescence recovery after photobleaching." Thesis, University of Manchester, 2014. https://www.research.manchester.ac.uk/portal/en/theses/analysis-of-bcl2-family-protein-interactions-in-live-cells-by-fluorescence-recovery-after-photobleaching(aa5eb271-6e43-48f3-940d-f63763ea4629).html.

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The Bcl-2 family of proteins strictly regulates the intrinsic pathway of apoptosis. Direct physical interactions between Bcl-2 proteins regulate mitochondrial outerpermeabilisation (MOMP), which occurs in response to various cell stresses andapoptotic stimuli. How changes in Bcl-2 protein activity regulate apoptosiscommitment is still unclear, especially with regard to how they interact with eachother within the context of the mitochondrial membrane. Recent studies haveshown that Bcl-2 proteins exist in a dynamic equilibrium between the mitochondriaand the cytosol. In this thesis, by using FRAP, I have measured changes in Bcl-XLand Mcl-1 dynamics in single cells. Surprisingly, individual cells within a populationshow widely differing Bcl-XL and Mcl-1 dynamics. There is a corelation betweenBcl-XL and Mcl-1 dynamics with BH3-only protein expression. Anti-apoptotic andpro-apoptotic Bcl-2 proteins stabilise each other on the OMM. Together, theseresults indicate that cells constantly fine tune mitochondrial priming and thatanalysing anti-apoptotic Bcl-2 proteins by FRAP allows this to be measured at asingle cell level in real time before MOMP.
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Gaffield, Michael A. "FRAP measurements of synaptic vesicle mobility in motor nerve terminals /." Connect to abstract via ProQuest. Full text is not available online, 2007.

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Thesis (Ph.D. in Neuroscience) -- University of Colorado Denver, 2007.
Typescript. Includes bibliographical references (leaves 84-93). Free to UCD affiliates. Online version available via ProQuest Digital Dissertations;
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Travascio, Francesco. "Modeling Molecular Transport and Binding Interactions in Intervertebral Disc." Scholarly Repository, 2009. http://scholarlyrepository.miami.edu/oa_dissertations/322.

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Low back pain represents a significant concern in the United States, with 70% of individuals experiencing symptoms at some point in their lifetime. Although the specific cause of low back pain remains unclear, symptoms have been strongly associated with degeneration of the intervertebral disc. Insufficient nutritional supply to the disc is believed to be a major mechanism for tissue degeneration. Understanding nutrients' transport in intervertebral disc is crucial to elucidate the mechanisms of disc degeneration, and to develop strategies for tissue repair (in vivo), and tissue engineering (in vitro). Transport in intervertebral disc is complex and involves a series of electromechanical, chemical and biological coupled events. Despite of the large amount of studies performed in the past, transport phenomena in the disc are still poorly understood. This is partly due to the limited number of available experimental techniques for investigating transport properties, and the paucity of theoretical or numerical methods for systematically predicting the mechanisms of solute transport in intervertebral disc. In this dissertation, a theoretical and experimental approach was taken in order to investigate the mechanisms of solute transport and binding interactions in intervertebral disc. New imaging techniques were developed for the experimental determination of diffusive and binding parameters in biological tissues. The techniques are based on the principle of fluorescence recovery after photobleaching, and allow the determination of the anisotropic diffusion tensor, and the rates of binding and unbinding of a solute to the extracellular matrix of a biological tissue. When applied to the characterization of transport properties of intervertebral disc, these methods allowed the establishment of a relationship between solute anisotropic and inhomogeneous diffusivity and the unique morphology of human lumbar annulus fibrosus. A mixture theory for charged hydrated soft tissues was presented as a framework for theoretical investigations on solute transport and binding interactions in cartilaginous tissues. Based on this theoretical framework and on experimental observations, a finite element model was developed to predict solute diffusive-convective-reactive transport in cartilaginous tissues. The numerical model was applied to simulate the effect of mechanical loading on solute transport and binding interactions in cartilage explants and intervertebral disc.
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Innhausen, u. Knyphausen Adrian zu [Verfasser], and Ralph [Akademischer Betreuer] Rupp. "A novel method for Fluorescence Recovery after Photobleaching (FRAP) analysis of chromatin proteins in pluripotent embryonic cells of the South African clawed frog X. laevis / Adrian zu Innhausen u. Knyphausen ; Betreuer: Ralph Rupp." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2020. http://d-nb.info/1221960563/34.

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Knust, Elisabeth, João Firmino, and Jean-Yves Tinevez. "Crumbs Affects Protein Dynamics In Anterior Regions Of The Developing Drosophila Embryo." Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2016. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-191623.

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Maintenance of apico-basal polarity is essential for epithelial integrity and requires particular reinforcement during tissue morphogenesis, when cells are reorganised, undergo shape changes and remodel their junctions. It is well established that epithelial integrity during morphogenetic processes depends on the dynamic exchange of adherens junction components, but our knowledge on the dynamics of other proteins and their dynamics during these processes is still limited. The early Drosophila embryo is an ideal system to study membrane dynamics during morphogenesis. Here, morphogenetic activities differ along the anterior-posterior axis, with the extending germband showing a high degree of epithelial remodelling. We developed a Fluorescence Recovery After Photobleaching (FRAP) assay with a higher temporal resolution, which allowed the distinction between a fast and a slow component of recovery of membrane proteins during the germband extension stage. We show for the first time that the recovery kinetics of a general membrane marker, SpiderGFP, differs in the anterior and posterior parts of the embryo, which correlates well with the different morphogenetic activities of the respective embryonic regions. Interestingly, absence of crumbs, a polarity regulator essential for epithelial integrity in the Drosophila embryo, decreases the fast component of SpiderGFP and of the apical marker Stranded at Second-Venus specifically in the anterior region. We suggest that the defects in kinetics observed in crumbs mutant embryos are the first signs of tissue instability in this region, explaining the earlier breakdown of the head epidermis in comparison to that of the trunk, and that diffusion in the plasma membrane is affected by the absence of Crumbs.
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Chennevière, Alexis. "Dynamique de chaînes de polymère greffés et glissement aux interfaces." Thesis, Paris 11, 2014. http://www.theses.fr/2014PA112404/document.

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Dans de nombreux cas, le développement de surfaces aux propriétés adhésives spécifiques fait appel à l’utilisation « d’interfaces décorées ». Ces interfaces sont composées d’un substrat solide sur lequel des chaînes de polymère sont plus ou moins bien ancrées. Ces chaînes se couplent mécaniquement au matériau environnant et contrôlent la transmission des contraintes de friction et d'adhésion aux interfaces. Ce couplage dépend en particulier de la pénétration des chaînes de surface dans la matrice et de leur dynamique. Dans cette thèse, les systèmes que nous avons étudiés sont constitués d’une couche de chaînes de polymère dont une extrémité est liée de manière covalente à un substrat solide. Ces brosses de polymère, constituent un système modèle pour des interfaces décorées. Notre objectif a été d’étudier la conformation et la dynamique de ces chaînes greffées lorsque ces dernières sont soumises à différents types de sollicitations afin de comprendre les mécanismes moléculaires régissant les propriétés d’adhésion et de friction de ce type d’interface. Dans un premier volet, nous avons étudié par réflectivité de neutrons la cinétique de cicatrisation d'une interface composée initialement de chaînes greffées recroquevillées sur un substrat et en contact avec un fondu. Lorsque le système est amené à une température supérieure à la température de transition vitreuse, les chaînes de polymère retrouvent une mobilité non nulle permettant ainsi la pénétration des chaînes greffées dans le fondu de polymère. La réflectivité de neutrons nous a permis d'une part de sonder à l'échelle moléculaire la cinétique de cicatrisation de ce type d’interface et d'autre part de la quantifier. L'influence des paramètres moléculaires sur cette cinétique de cicatrisation a pu être observée, ce qui nous a permis de proposer un modèle en loi d'échelle permettant d'apporter une interprétation physique au phénomène étudié. La deuxième partie de ce travail de thèse a consisté en l'élaboration d'un dispositif expérimental permettant de cisailler un système brosse/fondu au-dessus de la température de la température transition vitreuse et de geler la conformation des chaînes greffées dans leur configuration cisaillée. L'inversion des spectres de réflectivité neutrons associés a permis de mettre en évidence l'influence du cisaillement sur le degré d'interpénétration entre la brosse et le fondu qui régit la transmission des contraintes de friction sur ce type d'interface. De plus, nous avons pu mesurer la cinétique de relaxation de chaînes greffées, cisaillées au préalable, et la comparer aux expériences d’interdigitation simple. Cette comparaison a permis de divulguer l’importance du type de sollicitation sur la cinétique de relaxation d’une interface brosse / fondu.Nous avons également observé que la cinétique de relaxation et la conformation de chaînes greffées peuvent être altérées lorsque ces dernières sont confinées dans un film d'épaisseur comparable au rayon de giration des chaînes. Une étude systématique par réflectivité de neutrons a permis de mettre en évidence une accélération de la cinétique de relaxation du système en dessous d'une épaisseur critique qui pourrait être interprétée en termes de déplacement de la température de transition vitreuse. Dans un deuxième temps, nous avons étudié le glissement de solutions de polymères sur une surface greffée. La fraction volumique en chaînes libres dans la solution est un paramètre supplémentaire aux trois cas précédents qui contrôle ici le degré d'interpénétration entre chaînes libres et chaînes greffées. Une première approche théorique a permis de dissocier différents régimes de glissements à la paroi en fonction de fraction volumique. Nous avons entrepris une première série d'expériences de vélocimétrie laser après photolyse afin de mesurer le glissement à la paroi de solutions de polymère et de confronter les résultats expérimentaux avec notre approche théorique
In many cases, the development of surfaces with specific adhesive properties involves the use of "decorated interfaces." These interfaces consist of a solid substrate on which polymer chains are more or less well anchored. These chains are mechanically coupled to the surrounding material and control the transmission of friction and adhesion stresses at the interfaces. This coupling depends particularly on the penetration of the surface chains within the matrix and on their own dynamics. In this thesis, the systems we investigated are composed of a layer of polymer chains whose end is covalently linked to a solid substrate. These, so called, polymer brushes, provide a model system for decorated interfaces. Our objective was to study the conformation and dynamics of these grafted chains when they are subjected to different types of stress in order to understand the molecular mechanisms governing the adhesion and friction properties of this type of interface.In the first part, we investigated the healing kinetics of an interface composed initially of grafted chains collapsed on a substrate and in contact with a molten by using neutron reflectivity. When the system is brought above the glass transition temperature, the polymer chains mobility is high enough to allow the penetration of the grafted chains within the polymer melt. Neutrons reflectivity allowed us to probe at the molecular scale and to quantify the healing kinetics of this type of interface. The influence of molecular parameters on this healing kinetics was observed, which allowed us to propose a scaling law model to give a physical interpretation to the phenomenon studied.The second part of this thesis consisted in the development of an experimental setup which is able to shear a brush / melt interface above the glass transition temperature and to freeze the conformation of chains grafted in their sheared conformation. The inversion of the associated neutron reflectivity spectra made it possible to demonstrate the influence of shear on the degree of interpenetration between the brush and the melt which governs the transmission of stresses. In addition, we measured the kinetics of relaxation of grafted chains previously sheared and we compared it to the interdigitation experiments. This comparison highlighted the influence of the kind of solicitation on the relaxation kinetics of a brush/melt interface.We also observed that the relaxation kinetics and the conformation of the grafted chains may be altered when they are confined in a film which thickness is comparable to the radius of gyration of the chains. A systematic study using neutron reflectivity was conducted and highlighted an acceleration of the relaxation kinetics of the system below a critical thickness which could be interpreted in terms of a shift in the glass transition temperature.Secondly, we studied the slip of polymer solutions onto a grafted surface. The volume fraction of free chains in solution is an additional parameter which controls the degree of interpenetration between free chains and grafted chains. A first theoretical approach showed that different slip regimes can occur as a function of volume fraction. We have undertaken a first series of experiments using laser velocimetry after photobleaching to measure the surface velocity of flowing polymer solutions and to compare the experimental results to our theoretical approach
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Knust, Elisabeth, João Firmino, and Jean-Yves Tinevez. "Crumbs Affects Protein Dynamics In Anterior Regions Of The Developing Drosophila Embryo." Public Library of Science, 2013. https://tud.qucosa.de/id/qucosa%3A29137.

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Maintenance of apico-basal polarity is essential for epithelial integrity and requires particular reinforcement during tissue morphogenesis, when cells are reorganised, undergo shape changes and remodel their junctions. It is well established that epithelial integrity during morphogenetic processes depends on the dynamic exchange of adherens junction components, but our knowledge on the dynamics of other proteins and their dynamics during these processes is still limited. The early Drosophila embryo is an ideal system to study membrane dynamics during morphogenesis. Here, morphogenetic activities differ along the anterior-posterior axis, with the extending germband showing a high degree of epithelial remodelling. We developed a Fluorescence Recovery After Photobleaching (FRAP) assay with a higher temporal resolution, which allowed the distinction between a fast and a slow component of recovery of membrane proteins during the germband extension stage. We show for the first time that the recovery kinetics of a general membrane marker, SpiderGFP, differs in the anterior and posterior parts of the embryo, which correlates well with the different morphogenetic activities of the respective embryonic regions. Interestingly, absence of crumbs, a polarity regulator essential for epithelial integrity in the Drosophila embryo, decreases the fast component of SpiderGFP and of the apical marker Stranded at Second-Venus specifically in the anterior region. We suggest that the defects in kinetics observed in crumbs mutant embryos are the first signs of tissue instability in this region, explaining the earlier breakdown of the head epidermis in comparison to that of the trunk, and that diffusion in the plasma membrane is affected by the absence of Crumbs.
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Inthavong, Walailuk. "Elaboration et caractérisation de nanoparticules de protéines." Thesis, Le Mans, 2018. http://www.theses.fr/2018LEMA1014/document.

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Des solutions d'isolat de protéine de lactosérum (WPI) et d'isolat de protéine de soja (SPI) ont été chauffées à différentes concentrations en protéines conduisant à la formation d'agrégats fractals polydisperses de taille moyenne variable. Lastructure des solutions a été analysée par diffusion de la lumière en fonction de la concentration en protéine. La compressibilité osmotique et la longueur de corrélation dynamique diminuent quand la concentration augmente deviennent indépendantes de la taille initiale des agrégats pour les suspensions denses. Pour une taille d'agrégat donnée, la viscosité augmente initialement exponentiellement avec la concentration croissante puis diverge. Plus lesagrégats sont grands, plus l’augmentation de la viscosité apparaît à des concentrations faibles. La dépendance avec la concentration de la viscosité des solutions d'agrégats fractals est beaucoup plus forte que celle de microgels. Le comportement de mélanges de différents types d’agrégats (fractals/fractals ; fractals/microgels et WPI/SPI) a étéétudié principalement par rhéologie.Le recouvrement de fluorescence après photoblanchiment (FRAP) a été utilisé pour étudier la diffusion de chaînes de dextran marquées par des fluorophores dans des solutions d’agrégats et des gels de WPI. Une diffusion brownienne estobservée dans des suspensions d’agrégats et des gels faibles formés juste au-delà de Cg avec un coefficient de diffusion (D) qui diminue avec l'augmentation de la concentration mais, avec une dépendance plus faible que celle de la viscosité (). A des concentrations plus élevées, des gels densément réticulés sont formés, ce qui induit une forte diminution de la mobilité des chaînes de dextran. Pour ces systèmes, la recouvrance de la fluorescence est logarithmique avec le temps,suggérant une distribution exponentielle des coefficients de diffusion. La diffusion des chaînes de dextran a également été étudiée en fonction de la concentration en protéines pour les suspensions de trois types d'agrégats de WPI (petits et grands fractals et microgels)
Polydisperse fractal aggregates of varying average sizes were formed when solutions of whey protein isolate and soy protein isolate were heated at different protein concentrations and at neutral pH. The structure of these fractals aggregates solutions was analyzed by light scattering as a function of protein concentration. In dense suspension, the osmotic compressibility and the correlation length decreases with increasing concentration and become independent of the initial aggregate size. In this concentration regime, the aggregates are strongly interpenetrated and can be visualized as a set of "blobs". For a fixed aggregate size, the viscosity initially increases exponentially with increasing concentration and then diverges at the gel point. Larger fractal aggregates show a more important increase of the viscosity with increasing concentration than smaller aggregates, because they are less dense. The increase of the viscosity was much stronger for large fractal aggregates than for homogeneous microgels (microgels were formed by heating the WPI solution in present of CaCl2) of the same size.Dynamic light scattering, rheology and FRAP measurements were performed to investigate mixtures of different type of aggregates of WPI (fractals/fractals, fractals/microgels) and fractals of mixtures of WPI and SPI. Flow measurements were used to characterise the rheological properties of the aggregate suspension whereas Fluorescence recovery after Photobleaching (FRAP) was used to determine the self diffusion of fluorophore-labelled dextrans chains in mixtures over a wide range of concentrations. The results were compared to the concentration dependence of zero shear viscosity, gel stiffness, osmotic compressibility and correlation length. Brownian diffusion of the dextran chains was observed in aggregate suspensions and weak gels formed just above the gel point with a diffusion coefficient that decreased with increasing concentration, but the dependence was weaker than that of the viscosity. At higher concentrations, densely crosslinked gels were formed, which induced a sharp decrease in the mobility of the dextran chains. For these systems, the recovery of fluorescence was logarithmic over time, suggesting an exponential distribution of diffusion coefficients
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Irrechukwu, Onyi Nonye. "Role of matrix composition and age in solute diffusion within articular cartilage." Diss., Atlanta, Ga. : Georgia Institute of Technology, 2007. http://hdl.handle.net/1853/19699.

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Thesis (Ph.D)--Biomedical Engineering, Georgia Institute of Technology, 2008.
Committee Chair: Levenston, Marc; Committee Member: Garcia, Andres; Committee Member: Koros, William; Committee Member: Sambanis, Athanassios; Committee Member: Temenoff, Johnna; Committee Member: Vidakovic, Brani.
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Books on the topic "Fluorescence Recovery while photobleaching"

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Rietdorf, Jens. Microscopy Techniques (Advances in Biochemical Engineering / Biotechnology). Springer, 2005.

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Book chapters on the topic "Fluorescence Recovery while photobleaching"

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Russo, P. S., J. Qiu, N. Edwin, Y. W. Choi, G. J. Doucet, and D. Sohn. "Fluorescence Photobleaching Recovery." In Soft Matter Characterization, 605–36. Dordrecht: Springer Netherlands, 2008. http://dx.doi.org/10.1007/978-1-4020-4465-6_10.

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Deschout, Hendrik, and Kevin Braeckmans. "Fluorescence Recovery After Photobleaching." In Encyclopedia of Biophysics, 814–15. Berlin, Heidelberg: Springer Berlin Heidelberg, 2013. http://dx.doi.org/10.1007/978-3-642-16712-6_820.

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Carisey, Alex, Matthew Stroud, Ricky Tsang, and Christoph Ballestrem. "Fluorescence Recovery After Photobleaching." In Methods in Molecular Biology, 387–402. Totowa, NJ: Humana Press, 2011. http://dx.doi.org/10.1007/978-1-61779-207-6_26.

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Verkman, Alan S., Lakshmanan Vetrivel, and Peter Haggie. "Diffusion Measurements by Fluorescence Recovery After Photobleaching." In Methods in Cellular Imaging, 112–27. New York, NY: Springer New York, 2001. http://dx.doi.org/10.1007/978-1-4614-7513-2_7.

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Saito, Takumi, Daiki Matsunaga, and Shinji Deguchi. "Long-Term Fluorescence Recovery After Photobleaching (FRAP)." In Methods in Molecular Biology, 311–22. New York, NY: Springer US, 2023. http://dx.doi.org/10.1007/978-1-0716-2851-5_21.

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Kenworthy, Anne K. "Fluorescence Recovery After Photobleaching Studies of Lipid Rafts." In Methods in Molecular Biology, 179–92. Totowa, NJ: Humana Press, 2007. http://dx.doi.org/10.1007/978-1-59745-513-8_13.

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Houtsmuller, Adriaan B. "Fluorescence Recovery after Photobleaching: Application to Nuclear Proteins." In Microscopy Techniques, 177–99. Berlin, Heidelberg: Springer Berlin Heidelberg, 2005. http://dx.doi.org/10.1007/b102214.

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Wehrle-Haller, Bernhard. "Analysis of Integrin Dynamics by Fluorescence Recovery After Photobleaching." In Adhesion Protein Protocols, 173–201. Totowa, NJ: Humana Press, 2007. http://dx.doi.org/10.1007/978-1-59745-353-0_13.

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Silver, Kristen, and Rene E. Harrison. "Measuring Immune Receptor Mobility by Fluorescence Recovery After Photobleaching." In Methods in Molecular Biology, 155–67. Totowa, NJ: Humana Press, 2011. http://dx.doi.org/10.1007/978-1-61779-139-0_11.

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Carnell, Michael, Alex Macmillan, and Renee Whan. "Fluorescence Recovery After Photobleaching (FRAP): Acquisition, Analysis, and Applications." In Methods in Molecular Biology, 255–71. New York, NY: Springer New York, 2014. http://dx.doi.org/10.1007/978-1-4939-1752-5_18.

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Conference papers on the topic "Fluorescence Recovery while photobleaching"

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Li, Minghe, Aleksandr Razumtcev, Dustin M. Harmon, and Garth J. Simpson. "Fast diffusion characterization by fluorescence recovery while photobleaching (FRWP)." In Advanced Chemical Microscopy for Life Science and Translational Medicine 2023, edited by Garth J. Simpson, Ji-Xin Cheng, and Wei Min. SPIE, 2023. http://dx.doi.org/10.1117/12.2649098.

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Albro, Michael B., Vikram Rajan, Clark T. Hung, and Gerard A. Ateshian. "Fickian Behavior and Concentration-Dependence of the Diffusion of Dextran in Agarose." In ASME 2007 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2007. http://dx.doi.org/10.1115/sbc2007-176646.

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Various studies have attempted to quantify the effects of loading on nutrient transport in cartilage and other soft tissues. The application of a dynamic mechanical stimulus has been shown to significantly enhance the mechanical properties of chondrocyte-seeded agarose [1]. While the mechanism for this enhancement is still not completely understood, dynamic loading has been shown theoretically [2] as well as experimentally [3] to increase the uptake of large molecules. Since dextran is available in a wide range of molecular weights and can be conjugated with fluorphores, it has become a popular model system for studying solute transport in statically loaded and free swelling gels and tissues [4, 5]. To better characterize this model system, this study uses fluorescence recovery after photobleaching (FRAP) to investigate the Fickian behavior of linear dextran macromolecules as well as the dependence of its diffusivity on concentration.
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Jeonghoon Lee, Donghee Lee, Myoung-Ock Cho, and Jung Kyung Kim. "Toward reducing uncertainty in Fluorescence Recovery After Photobleaching." In 2013 35th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC). IEEE, 2013. http://dx.doi.org/10.1109/embc.2013.6610536.

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Khandai, Santripti, Ronald A. Siegel, and Sidhartha S. Jena. "Probing the microstructure of hydrogels using fluorescence recovery after photobleaching." In SOLID STATE PHYSICS: PROCEEDINGS OF THE 57TH DAE SOLID STATE PHYSICS SYMPOSIUM 2012. AIP, 2013. http://dx.doi.org/10.1063/1.4790947.

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Shvec, Peter, J. Miklovicova, L. Hunakova, and B. Chorvath. "Translational dynamics of immune system components by fluorescence recovery after photobleaching." In Moscow - DL tentative, edited by Sergei A. Akhmanov and Marina Y. Poroshina. SPIE, 1991. http://dx.doi.org/10.1117/12.57345.

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Gupta, Sharad, Bhawna Bhawna, Asima Pradhan, S. Swain, and Asha Agarwal. "Fluorescence photobleaching and recovery of human breast tissues and tissue phantoms." In International Symposium on Biomedical Optics, edited by Robert R. Alfano. SPIE, 2002. http://dx.doi.org/10.1117/12.465262.

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Cao, Ziyi, Dustin Harmon, Jiayue Rong, Andreas Geiger, and Garth J. Simpson. "Fourier-transform Fluorescence Recovery after Photobleaching (FT-FRAP) diffusion imaging analysis." In Advanced Chemical Microscopy for Life Science and Translational Medicine 2022, edited by Garth J. Simpson, Ji-Xin Cheng, and Wei Min. SPIE, 2022. http://dx.doi.org/10.1117/12.2607631.

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BIRMINGHAM, J. J. "PHASE-FRAP: A NEW FREQUENCY-DOMAIN VARIANT OF FLUORESCENCE RECOVERY AFTER PHOTOBLEACHING." In Proceedings of the Fifth Royal Society–Unilever Indo-UK Forum in Materials Science and Engineering. A CO-PUBLICATION OF IMPERIAL COLLEGE PRESS AND THE ROYAL SOCIETY, 2000. http://dx.doi.org/10.1142/9781848160163_0007.

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Teijeiro Gonzalez, Yurema, Klaus Suhling, Andrew Beavil, Rebecca Beavil, James Levitt, Maddy Parsons, Elena Ortiz-Zapater, et al. "Fluorescence Recovery After Photobleaching (FRAP) with simultaneous Fluorescence Lifetime and time-resolved Fluorescence Anisotropy Imaging (FLIM and tr-FAIM)." In Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XXVI, edited by Thomas G. Brown and Tony Wilson. SPIE, 2019. http://dx.doi.org/10.1117/12.2508692.

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Tylcz, Jean-Baptiste, Muriel Abbaci, Thierry Bastogne, Walter Blondel, Dominique Dumas, and Muriel Barberi-Heyob. "System identification of the fluorescence recovery after photobleaching in gap junctional intracellular communications." In 2013 IEEE 52nd Annual Conference on Decision and Control (CDC). IEEE, 2013. http://dx.doi.org/10.1109/cdc.2013.6761029.

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