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Artículos de revistas sobre el tema "Fluorescent recovery after photobleaching (FRAP)"

Autor: Grafiati
Publicado: 25 de enero de 2025
Última modificación: 31 de julio de 2025

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1

Cadar, Adrian G., Tromondae K. Feaster, Kevin R. Bersell, et al. "Real-time visualization of titin dynamics reveals extensive reversible photobleaching in human induced pluripotent stem cell-derived cardiomyocytes." American Journal of Physiology-Cell Physiology 318, no. 1 (2020): C163—C173. http://dx.doi.org/10.1152/ajpcell.00107.2019.

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Fluorescence recovery after photobleaching (FRAP) has been useful in delineating cardiac myofilament biology, and innovations in fluorophore chemistry have expanded the array of microscopic assays used. However, one assumption in FRAP is the irreversible photobleaching of fluorescent proteins after laser excitation. Here we demonstrate reversible photobleaching regarding the photoconvertible fluorescent protein mEos3.2. We used CRISPR/Cas9 genome editing in human induced pluripotent stem cells (hiPSCs) to knock-in mEos3.2 into the COOH terminus of titin to visualize sarcomeric titin incorporat
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2

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 (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 studie
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3

Srikantha, Nishanthan, Yurema Teijeiro-Gonzalez, Andrew Simpson, et al. "Determining vitreous viscosity using fluorescence recovery after photobleaching." PLOS ONE 17, no. 2 (2022): e0261925. http://dx.doi.org/10.1371/journal.pone.0261925.

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Purpose Vitreous humor is a complex biofluid whose composition determines its structure and function. Vitreous viscosity will affect the delivery, distribution, and half-life of intraocular drugs, and key physiological molecules. The central pig vitreous is thought to closely match human vitreous viscosity. Diffusion is inversely related to viscosity, and diffusion is of fundamental importance for all biochemical reactions. Fluorescence Recovery After Photobleaching (FRAP) may provide a novel means of measuring intravitreal diffusion that could be applied to drugs and physiological macromolecu
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4

Braga, José, Joana M. P. Desterro, and Maria Carmo-Fonseca. "Intracellular Macromolecular Mobility Measured by Fluorescence Recovery after Photobleaching with Confocal Laser Scanning Microscopes." Molecular Biology of the Cell 15, no. 10 (2004): 4749–60. http://dx.doi.org/10.1091/mbc.e04-06-0496.

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Fluorescence recovery after photobleaching (FRAP) is a widely used tool for estimating mobility parameters of fluorescently tagged molecules in cells. Despite the widespread use of confocal laser scanning microscopes (CLSMs) to perform photobleaching experiments, quantitative data analysis has been limited by lack of appropriate practical models. Here, we present a new approximate FRAP model for use on any standard CLSM. The main novelty of the method is that it takes into account diffusion of highly mobile molecules during the bleach phase. In fact, we show that by the time the first postblea
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5

Orlova, Darya, Eva Bartova, Valeri Maltsev, and Kozubek Stanislav. "A non-fitting method employing a spatial sine window transform for inhomogeneous effective diffusion measurements by FRAP." Biophysical Journal 100, no. 2 (2011): 507–16. https://doi.org/10.1016/j.bpj.2010.11.080.

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Determining averaged effective diffusion constants from experimental measurements of fluorescent proteins in an inhomogeneous medium in the presence of ligand-receptor interactions poses problems of analytical tractability. Here, we introduced a nonfitting method to evaluate the averaged effective diffusion coefficient of a region of interest (which may include a whole nucleus) by mathematical processing of the entire cellular two-dimensional spatial pattern of recovered fluorescence. Spatially and temporally resolved measurements of protein transport inside cells were obtained using the fluor
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6

Lorén, Niklas, Joel Hagman, Jenny K. Jonasson, et al. "Fluorescence recovery after photobleaching in material and life sciences: putting theory into practice." Quarterly Reviews of Biophysics 48, no. 3 (2015): 323–87. http://dx.doi.org/10.1017/s0033583515000013.

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AbstractFluorescence recovery after photobleaching (FRAP) is a versatile tool for determining diffusion and interaction/binding properties in biological and material sciences. An understanding of the mechanisms controlling the diffusion requires a deep understanding of structure–interaction–diffusion relationships. In cell biology, for instance, this applies to the movement of proteins and lipids in the plasma membrane, cytoplasm and nucleus. In industrial applications related to pharmaceutics, foods, textiles, hygiene products and cosmetics, the diffusion of solutes and solvent molecules cont
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7

Golebiewska, Urszula, Jason G. Kay, Thomas Masters, et al. "Evidence for a fence that impedes the diffusion of phosphatidylinositol 4,5-bisphosphate out of the forming phagosomes of macrophages." Molecular Biology of the Cell 22, no. 18 (2011): 3498–507. http://dx.doi.org/10.1091/mbc.e11-02-0114.

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To account for the many functions of phosphatidylinositol 4,5-bisphosphate (PIP2), several investigators have proposed that there are separate pools of PIP2 in the plasma membrane. Recent experiments show the surface concentration of PIP2 is indeed enhanced in regions where phagocytosis, exocytosis, and cell division occurs. Kinases that produce PIP2 are also concentrated in these regions. However, how is the PIP2 produced by these kinases prevented from diffusing rapidly away? First, proteins could act as “fences” around the perimeter of these regions. Second, some factor could markedly decre
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8

Verma, Sanjay K., Pratibha Kumari, Shagufi Naz Ansari, Mohd Ovais Ansari, Dondinath Deori, and Shaikh M. Mobin. "A novel mesoionic carbene based highly fluorescent Pd(ii) complex as an endoplasmic reticulum tracker in live cells." Dalton Transactions 47, no. 44 (2018): 15646–50. http://dx.doi.org/10.1039/c8dt02778a.

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9

Wagner, Stefan, Simion Chiosea, Maria Ivshina, and Jeffrey A. Nickerson. "In vitro FRAP reveals the ATP-dependent nuclear mobilization of the exon junction complex protein SRm160." Journal of Cell Biology 164, no. 6 (2004): 843–50. http://dx.doi.org/10.1083/jcb.200307002.

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We present a new in vitro system for characterizing the binding and mobility of enhanced green fluorescent protein (EGFP)–labeled nuclear proteins by fluorescence recovery after photobleaching in digitonin-permeabilized cells. This assay reveals that SRm160, a splicing coactivator and component of the exon junction complex (EJC) involved in RNA export, has an adenosine triphosphate (ATP)–dependent mobility. Endogenous SRm160, lacking the EGFP moiety, could also be released from sites at splicing speckled domains by an ATP-dependent mechanism. A second EJC protein, RNPS1, also has an ATP-depend
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10

Wadsworth, P., and E. D. Salmon. "Analysis of the treadmilling model during metaphase of mitosis using fluorescence redistribution after photobleaching." Journal of Cell Biology 102, no. 3 (1986): 1032–38. http://dx.doi.org/10.1083/jcb.102.3.1032.

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One recent hypothesis for the mechanism of chromosome movement during mitosis predicts that a continual, uniform, poleward flow or "treadmilling" of microtubules occurs within the half-spindle between the chromosomes and the poles during mitosis (Margolis, R. L., and L. Wilson, 1981, Nature (Lond.), 293:705-711). We have tested this treadmilling hypothesis using fluorescent analog cytochemistry and measurements of fluorescence redistribution after photobleaching to examine microtubule behavior during metaphase of mitosis. Mitotic BSC 1 mammalian tissue culture cells or newt lung epithelial cel
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11

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 im
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12

Cai, Ning, Alvin Chi-Keung Lai, Kin Liao, Peter R. Corridon, David J. Graves, and Vincent Chan. "Recent Advances in Fluorescence Recovery after Photobleaching for Decoupling Transport and Kinetics of Biomacromolecules in Cellular Physiology." Polymers 14, no. 9 (2022): 1913. http://dx.doi.org/10.3390/polym14091913.

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Among the new molecular tools available to scientists and engineers, some of the most useful include fluorescently tagged biomolecules. Tools, such as green fluorescence protein (GFP), have been applied to perform semi-quantitative studies on biological signal transduction and cellular structural dynamics involved in the physiology of healthy and disease states. Such studies focus on drug pharmacokinetics, receptor-mediated endocytosis, nuclear mechanobiology, viral infections, and cancer metastasis. In 1976, fluorescence recovery after photobleaching (FRAP), which involves the monitoring of f
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13

Waharte, François, Karine Steenkeste, Romain Briandet, and Marie-Pierre Fontaine-Aupart. "Diffusion Measurements inside Biofilms by Image-Based Fluorescence Recovery after Photobleaching (FRAP) Analysis with a Commercial Confocal Laser Scanning Microscope." Applied and Environmental Microbiology 76, no. 17 (2010): 5860–69. http://dx.doi.org/10.1128/aem.00754-10.

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ABSTRACT Research about the reactional and structural dynamics of biofilms at the molecular level has made great strides, owing to efficient fluorescence imaging methods in terms of spatial resolution and fast acquisition time but also to noninvasive conditions of observation consistent with in situ biofilm studies. In addition to conventional fluorescence intensity imaging, the fluorescence recovery after photobleaching (FRAP) module can now be routinely implemented on commercial confocal laser scanning microscopes (CLSMs). This method allows measuring of local diffusion coefficients in biofi
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14

Waters, Jennifer C. "Accuracy and precision in quantitative fluorescence microscopy." Journal of Cell Biology 185, no. 7 (2009): 1135–48. http://dx.doi.org/10.1083/jcb.200903097.

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The light microscope has long been used to document the localization of fluorescent molecules in cell biology research. With advances in digital cameras and the discovery and development of genetically encoded fluorophores, there has been a huge increase in the use of fluorescence microscopy to quantify spatial and temporal measurements of fluorescent molecules in biological specimens. Whether simply comparing the relative intensities of two fluorescent specimens, or using advanced techniques like Förster resonance energy transfer (FRET) or fluorescence recovery after photobleaching (FRAP), qu
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15

Dallon, J. C., Cécile Leduc, Christopher P. Grant, Emily J. Evans, Sandrine Etienne-Manneville, and Stéphanie Portet. "Using Fluorescence Recovery After Photobleaching data to uncover filament dynamics." PLOS Computational Biology 18, no. 9 (2022): e1010573. http://dx.doi.org/10.1371/journal.pcbi.1010573.

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Fluorescence Recovery After Photobleaching (FRAP) has been extensively used to understand molecular dynamics in cells. This technique when applied to soluble, globular molecules driven by diffusion is easily interpreted and well understood. However, the classical methods of analysis cannot be applied to anisotropic structures subjected to directed transport, such as cytoskeletal filaments or elongated organelles transported along microtubule tracks. A new mathematical approach is needed to analyze FRAP data in this context and determine what information can be obtain from such experiments. To
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16

Vikstrom, K. L., S. S. Lim, R. D. Goldman, and G. G. Borisy. "Steady state dynamics of intermediate filament networks." Journal of Cell Biology 118, no. 1 (1992): 121–29. http://dx.doi.org/10.1083/jcb.118.1.121.

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We have conducted experiments to examine the dynamic exchange between subunit and polymer of vimentin intermediate filaments (IF) at steady state through the use of xrhodamine-labeled vimentin in fluorescence recovery after photobleaching (FRAP) analysis. The xrhodamine-vimentin incorporated into the endogenous vimentin IF network after microinjection into fibroblasts and could be visualized with a cooled charge-coupled device (CCD) camera and digital imaging fluorescence microscopy. Bar shaped regions were bleached in the fluorescent IF network using a beam from an argon ion laser and the cel
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17

Kure, Jakob L., Camilla B. Andersen, Thomas E. Rasmussen, B. Christoffer Lagerholm, and Eva C. Arnspang. "Defining the Diffusion in Model Membranes Using Line Fluorescence Recovery after Photobleaching." Membranes 10, no. 12 (2020): 434. http://dx.doi.org/10.3390/membranes10120434.

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In this study, we explore the use of line FRAP to detect diffusion in synthetic lipid membranes. The study of the dynamics of these membrane lipids can, however, be challenging. The diffusion in two different synthetic membranes consisting of the lipid mixtures 1:1 DOPC:DPPC and 2:2:1 DOPC:DPPC:Cholesterol was studied with line FRAP. A correlation between diffusion coefficient and temperature was found to be dependent on the morphology of the membrane. We suggest line FRAP as a promising accessible and simple technique to study diffusion in plasma membranes.
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18

Yuste, S. B., E. Abad, and K. Lindenberg. "A reaction–subdiffusion model of fluorescence recovery after photobleaching (FRAP)." Journal of Statistical Mechanics: Theory and Experiment 2014, no. 11 (2014): P11014. http://dx.doi.org/10.1088/1742-5468/2014/11/p11014.

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19

Berk, D. A., M. A. Swartz, A. J. Leu, and R. K. Jain. "Transport in lymphatic capillaries. II. Microscopic velocity measurement with fluorescence photobleaching." American Journal of Physiology-Heart and Circulatory Physiology 270, no. 1 (1996): H330—H337. http://dx.doi.org/10.1152/ajpheart.1996.270.1.h330.

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Despite its relevance to the physiology of lymph formation and propulsion, the instantaneous flow velocity in single lymphatic capillaries has not been measured to date. The method of fluorescence recovery after photobleaching (FRAP) was adapted for this purpose and used to characterize flow in the lymphatic capillaries in tail skin of anesthetized mice during a constant-pressure intradermal injection of fluorescein isothiocyanate-dextran (mol wt 2 x 10(6). The median lymph flow velocity was 4.7 microns/s, and the velocity magnitude ranged from 0 to 29 microns/s. The direction of flow was gene
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20

Day, Charles A., and Minchul Kang. "The Utility of Fluorescence Recovery after Photobleaching (FRAP) to Study the Plasma Membrane." Membranes 13, no. 5 (2023): 492. http://dx.doi.org/10.3390/membranes13050492.

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The plasma membrane of mammalian cells is involved in a wide variety of cellular processes, including, but not limited to, endocytosis and exocytosis, adhesion and migration, and signaling. The regulation of these processes requires the plasma membrane to be highly organized and dynamic. Much of the plasma membrane organization exists at temporal and spatial scales that cannot be directly observed with fluorescence microscopy. Therefore, approaches that report on the membrane’s physical parameters must often be utilized to infer membrane organization. As discussed here, diffusion measurements
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21

Rayan, Gamal, Jean-Erik Guet, Nicolas Taulier, Frederic Pincet, and Wladimir Urbach. "Recent Applications of Fluorescence Recovery after Photobleaching (FRAP) to Membrane Bio-Macromolecules." Sensors 10, no. 6 (2010): 5927–48. http://dx.doi.org/10.3390/s100605927.

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22

Barsony, J., J. Carroll, W. McKoy, et al. "Intracellular Traffic of Glucocorticoid Receptors: Studies With Green Fluorescent Protein Chimeras in Living Cells." Microscopy and Microanalysis 3, S2 (1997): 131–32. http://dx.doi.org/10.1017/s1431927600007546.

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As ligand-regulated transcription factors, glucocorticoid receptors (GR) must traffic through the cytoplasm, traverse the nuclear pores, and subsequently traffic within the the nucleus to reach their target genes. Due to technical difficulties with immunocytology, little is known about the translocation process or the intranuclear localization. The recent characterization of a chromophore, green fluorescent protein (GFP), provided a general tool to fluorescently label proteins in living cells. With the development of a transcriptionally active GFP-GR chimera, it became possible to visualize GR
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23

Worden, Austin N. "The Alphabet Soup of Microscopy: An Introduction to Advanced Imaging Techniques. Part II: What the FLIP, FLAP, FRAP, FRET, and FLIM is Going On?" Microscopy Today 32, no. 5 (2024): 53–57. http://dx.doi.org/10.1093/mictod/qaae065.

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Abstract The use of fluorescence in imaging has been a pivotal factor in advancing our scientific understanding. As microscopy continues to evolve, the terminology used to describe these techniques becomes increasingly complex, often resulting in a bewildering array of acronyms that resemble alphabet soup. Among the most prominent acronyms are those associated with advanced fluorescence microscopy techniques: Fluorescence Loss in Photobleaching (FLIP), Fluorescence Localization after Photobleaching (FLAP), Fluorescence Recovery after Photobleaching (FRAP), Förster Resonance Energy Transfer (FR
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24

Cardarelli, Francesco, Luca Tosti, Michela Serresi, Fabio Beltram, and Ranieri Bizzarri. "Fluorescent Recovery after Photobleaching (FRAP) Analysis of Nuclear Export Rates Identifies Intrinsic Features of Nucleocytoplasmic Transport." Journal of Biological Chemistry 287, no. 8 (2011): 5554–61. http://dx.doi.org/10.1074/jbc.m111.304899.

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25

Scardigli, M., C. Crocini, C. Ferrantini, et al. "Quantitative assessment of passive electrical properties of the cardiac T-tubular system by FRAP microscopy." Proceedings of the National Academy of Sciences 114, no. 22 (2017): 5737–42. http://dx.doi.org/10.1073/pnas.1702188114.

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Well-coordinated activation of all cardiomyocytes must occur on every heartbeat. At the cell level, a complex network of sarcolemmal invaginations, called the transverse-axial tubular system (TATS), propagates membrane potential changes to the cell core, ensuring synchronous and uniform excitation–contraction coupling. Although myocardial conduction of excitation has been widely described, the electrical properties of the TATS remain mostly unknown. Here, we exploit the formal analogy between diffusion and electrical conductivity to link the latter with the diffusional properties of TATS. Fluo
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26

Huang, Zhenqiu, Sabine Kaltenbrunner, Eva Šimková, David Stanĕk, Julius Lukeš, and Hassan Hashimi. "Dynamics of Mitochondrial RNA-Binding Protein Complex in Trypanosoma brucei and Its Petite Mutant under Optimized Immobilization Conditions." Eukaryotic Cell 13, no. 9 (2014): 1232–40. http://dx.doi.org/10.1128/ec.00149-14.

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ABSTRACT There are a variety of complex metabolic processes ongoing simultaneously in the single, large mitochondrion of Trypanosoma brucei . Understanding the organellar environment and dynamics of mitochondrial proteins requires quantitative measurement in vivo . In this study, we have validated a method for immobilizing both procyclic stage (PS) and bloodstream stage (BS) T. brucei brucei with a high level of cell viability over several hours and verified its suitability for undertaking fluorescence recovery after photobleaching (FRAP), with mitochondrion-targeted yellow fluorescent protein
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Favard, Cyril. "Numerical Simulation and FRAP Experiments Show That the Plasma Membrane Binding Protein PH-EFA6 Does Not Exhibit Anomalous Subdiffusion in Cells." Biomolecules 8, no. 3 (2018): 90. http://dx.doi.org/10.3390/biom8030090.

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The fluorescence recovery after photobleaching (FRAP) technique has been used for decades to measure movements of molecules in two-dimension (2D). Data obtained by FRAP experiments in cell plasma membranes are assumed to be described through a means of two parameters, a diffusion coefficient, D (as defined in a pure Brownian model) and a mobile fraction, M. Nevertheless, it has also been shown that recoveries can be nicely fit using anomalous subdiffusion. Fluorescence recovery after photobleaching (FRAP) at variable radii has been developed using the Brownian diffusion model to access geometr
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28

Yoon, Kyeong Han, Miri Yoon, Robert D. Moir, Satya Khuon, Frederick W. Flitney, and Robert D. Goldman. "Insights into the Dynamic Properties of Keratin Intermediate Filaments in Living Epithelial Cells." Journal of Cell Biology 153, no. 3 (2001): 503–16. http://dx.doi.org/10.1083/jcb.153.3.503.

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The properties of keratin intermediate filaments (IFs) have been studied after transfection with green fluorescent protein (GFP)-tagged K18 and/or K8 (type I/II IF proteins). GFP-K8 and -K18 become incorporated into tonofibrils, which are comprised of bundles of keratin IFs. These tonofibrils exhibit a remarkably wide range of motile and dynamic activities. Fluorescence recovery after photobleaching (FRAP) analyses show that they recover their fluorescence slowly with a recovery t1/2 of ∼100 min. The movements of bleach zones during recovery show that closely spaced tonofibrils (<1 μm a
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29

Hori, Masatoshi, Jeffrey D. Jones, Lee Janson, Keith Ragsdale, and Katherine Luby-Phelps. "Light – microscopic analysis of the physical properties of cytoplasm in living cells." Proceedings, annual meeting, Electron Microscopy Society of America 54 (August 11, 1996): 734–35. http://dx.doi.org/10.1017/s0424820100166130.

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We have used a variety of optical techniques to explore the intracellular constraints on diffusion, cytoplasmic compartmentalization and biomechanics of living tissue culture cells. Fluorescence ratio imaging measurements of solvent viscosity in single cells indicate that the solvent viscosity of cytoplasm does not differ detectably from bulk water. Nevertheless, data obtained by fluorescence recovery after photobleaching (FRAP) of inert tracer molecules suggest that macromolecular crowding and molecular sieving retard the long-range translational diffusion of protein-sized molecules from 4 to
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Watanabe, Yumi, Masahito Hayashi, Toshiki Yagi, and Ritsu Kamiya. "Turnover of Actin in Chlamydomonas Flagella Detected by Fluorescence Recovery After Photobleaching (FRAP)." Cell Structure and Function 29, no. 3 (2004): 67–72. http://dx.doi.org/10.1247/csf.29.67.

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Kipper, Franciele Cristina, Alessandra Sayuri Kikuchi Tamajusuku, Darlan Conterno Minussi, et al. "Analysis of NTPDase2 in the cell membrane using fluorescence recovery after photobleaching (FRAP)." Cytometry Part A 93, no. 2 (2018): 232–38. http://dx.doi.org/10.1002/cyto.a.23317.

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32

Storrie, B., R. Pepperkok, E. H. Stelzer, and T. E. Kreis. "The intracellular mobility of a viral membrane glycoprotein measured by confocal microscope fluorescence recovery after photobleaching." Journal of Cell Science 107, no. 5 (1994): 1309–19. http://dx.doi.org/10.1242/jcs.107.5.1309.

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Fluorescence recovery after photobleaching (FRAP) has been a powerful tool for characterizing the mobility of cell surface membrane proteins. However, the application of FRAP to the study of intracellular membrane proteins has been hampered by the lack of specific probes and their physical inaccessibility in the cytoplasm. We have measured the mobility of a model transmembrane protein, the temperature-sensitive vesicular stomatitis viral membrane glycoprotein (ts-O45-G), in transit from the endoplasmic reticulum (ER) to the Golgi complex. ts-O45-G accumulates in the ER at nonpermissive tempera
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33

Orekhov, Fedor K., Arthur G. Jablokov, and Andrew A. Skrynnik. "Ybridization of laser-induced spectrofluorescence analysis (lifs), matrix-assisted laser desorption / ionization mass spectrometry (maldi), fluorescence recovery after photobleaching (frap) anf fluorescence loss in photobleaching (flip) microtechnics." Journal of Biomedical Technologies, no. 2 (December 2016): 42–52. http://dx.doi.org/10.15393/j6.art.2016.3702.

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Brandon, Elizabeth, Tomasz Szul, Cecilia Alvarez, et al. "On and Off Membrane Dynamics of the Endoplasmic Reticulum–Golgi Tethering Factor p115 In Vivo." Molecular Biology of the Cell 17, no. 7 (2006): 2996–3008. http://dx.doi.org/10.1091/mbc.e05-09-0862.

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The mechanisms regulating membrane recruitment of the p115 tethering factor in vivo are unknown. Here, we describe cycling of p115 between membranes and cytosol and document the effects of Golgi matrix proteins, Rab1, and soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein (SNAP) receptors (SNAREs) on this process. Rapid membrane/cytosol exchange is shown by swift (t1/2 ∼20 s) loss of Golgi-localized p115-green fluorescent protein (GFP) after repeated photobleaching of cell periphery and rapid (t1/2 ∼13 s) fluorescence recovery after photobleaching Golgi-localized p115-GFP. p115
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Saito, Takumi, Daiki Matsunaga, and Shinji Deguchi. "Long-term molecular turnover of actin stress fibers revealed by advection-reaction analysis in fluorescence recovery after photobleaching." PLOS ONE 17, no. 11 (2022): e0276909. http://dx.doi.org/10.1371/journal.pone.0276909.

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Fluorescence recovery after photobleaching (FRAP) is a versatile technique to evaluate the intracellular molecular exchange called turnover. Mechanochemical models of FRAP typically consider the molecular diffusion and chemical reaction that simultaneously occur on a time scale of seconds to minutes. Particularly for long-term measurements, however, a mechanical advection effect can no longer be ignored, which transports the proteins in specific directions within the cells and accordingly shifts the spatial distribution of the local chemical equilibrium. Nevertheless, existing FRAP models have
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36

Engel, Stephanie, Silvia Scolari, Bastian Thaa, et al. "FLIM-FRET and FRAP reveal association of influenza virus haemagglutinin with membrane rafts." Biochemical Journal 425, no. 3 (2010): 567–73. http://dx.doi.org/10.1042/bj20091388.

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It has been supposed that the HA (haemagglutinin) of influenza virus must be recruited to membrane rafts to perform its function in membrane fusion and virus budding. In the present study, we aimed at substantiating this association in living cells by biophysical methods. To this end, we fused the cyan fluorescent protein Cer (Cerulean) to the cytoplasmic tail of HA. Upon expression in CHO (Chinese-hamster ovary) cells HA–Cer was glycosylated and transported to the plasma membrane in a similar manner to authentic HA. We measured FLIM-FRET (Förster resonance energy transfer by fluorescence life
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37

Mullineaux, Conrad W., Anja Nenninger, Nicola Ray, and Colin Robinson. "Diffusion of Green Fluorescent Protein in Three Cell Environments in Escherichia Coli." Journal of Bacteriology 188, no. 10 (2006): 3442–48. http://dx.doi.org/10.1128/jb.188.10.3442-3448.2006.

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ABSTRACT Surprisingly little is known about the physical environment inside a prokaryotic cell. Knowledge of the rates at which proteins and other cell components can diffuse is crucial for the understanding of a cell as a physical system. There have been numerous measurements of diffusion coefficients in eukaryotic cells by using fluorescence recovery after photobleaching (FRAP) and related techniques. Much less information is available about diffusion coefficients in prokaryotic cells, which differ from eukaryotic cells in a number of significant respects. We have used FRAP to observe the di
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38

Moir, Robert D., Miri Yoon, Satya Khuon, and Robert D. Goldman. "Nuclear Lamins a and B1." Journal of Cell Biology 151, no. 6 (2000): 1155–68. http://dx.doi.org/10.1083/jcb.151.6.1155.

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At the end of mitosis, the nuclear lamins assemble to form the nuclear lamina during nuclear envelope formation in daughter cells. We have fused A- and B-type nuclear lamins to the green fluorescent protein to study this process in living cells. The results reveal that the A- and B-type lamins exhibit different pathways of assembly. In the early stages of mitosis, both lamins are distributed throughout the cytoplasm in a diffusible (nonpolymerized) state, as demonstrated by fluorescence recovery after photobleaching (FRAP). During the anaphase-telophase transition, lamin B1 begins to become co
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39

Floury, J., M. N. Madec, F. Waharte, S. Jeanson, and S. Lortal. "First assessment of diffusion coefficients in model cheese by fluorescence recovery after photobleaching (FRAP)." Food Chemistry 133, no. 2 (2012): 551–56. http://dx.doi.org/10.1016/j.foodchem.2012.01.030.

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40

Wahl, Philippe, and Fouad Azizi. "Fluorescent recovery after photobleaching (FRAP) of a fluorescent transferrin internalized in the late transferrin endocytic compartment of living A431 cells: Theory." Biochimica et Biophysica Acta (BBA) - Biomembranes 1327, no. 1 (1997): 69–74. http://dx.doi.org/10.1016/s0005-2736(97)00045-x.

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41

Redman, C. A., and J. R. Kusel. "Distribution and biophysical properties of fluorescent lipids on the surface of adult Schistosoma mansoni." Parasitology 113, no. 2 (1996): 137–43. http://dx.doi.org/10.1017/s0031182000066385.

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SUMMARYThe properties of 4 fluorescent lipid compounds in the surface membrane of adult male Schistosoma mansoni worms were examined by fluorescent microscopy and fluorescent recovery after photobleaching (FRAP). The data suggest that the probes N-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-pentanoyl) sphingosine (BODIPY FL ceramide) and PKH2 pass through the outer membrane and enter structures in or below the membrane. In contrast 5-(N-octadecanoyl)aminofluorescein (AF18) and N-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-pentanoyl) sphingosylphosphocholine (BODIP
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42

Handwerger, Korie E., Christine Murphy, and Joseph G. Gall. "Steady-state dynamics of Cajal body components in the Xenopus germinal vesicle." Journal of Cell Biology 160, no. 4 (2003): 495–504. http://dx.doi.org/10.1083/jcb.200212024.

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Cajal bodies (CBs) are evolutionarily conserved nuclear organelles that contain many factors involved in the transcription and processing of RNA. It has been suggested that macromolecular complexes preassemble or undergo maturation within CBs before they function elsewhere in the nucleus. Most such models of CB function predict a continuous flow of molecules between CBs and the nucleoplasm, but there are few data that directly support this view. We used fluorescence recovery after photobleaching (FRAP) on isolated Xenopus oocyte nuclei to measure the steady-state exchange rate between the nucl
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43

Azizi, Fouad, and Philippe Wahl. "Fluorescence recovery after photobleaching (FRAP) of a fluorescent transferrin internalized in the late transferrin endocytic compartment of living A431 cells: Experiments." Biochimica et Biophysica Acta (BBA) - Biomembranes 1327, no. 1 (1997): 75–88. http://dx.doi.org/10.1016/s0005-2736(97)00046-1.

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44

Govindaraj, Kannan, Jan Hendriks, Diane S. Lidke, Marcel Karperien, and Janine N. Post. "Changes in Fluorescence Recovery After Photobleaching (FRAP) as an indicator of SOX9 transcription factor activity." Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms 1862, no. 1 (2019): 107–17. http://dx.doi.org/10.1016/j.bbagrm.2018.11.001.

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45

Dewavrin, Jean-Yves, and Michael Raghunath. "Determining the Optimal Degree of Macromolecular Crowding in Solution Using Fluorescence Recovery after Photobleaching (FRAP)." Biophysical Journal 102, no. 3 (2012): 400a. http://dx.doi.org/10.1016/j.bpj.2011.11.2184.

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46

Zhao, Runchen, Siqi Cui, Zhuoxu Ge, et al. "Hydraulic resistance induces cell phenotypic transition in confinement." Science Advances 7, no. 17 (2021): eabg4934. http://dx.doi.org/10.1126/sciadv.abg4934.

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Cells penetrating into confinement undergo mesenchymal-to-amoeboid transition. The topographical features of the microenvironment expose cells to different hydraulic resistance levels. How cells respond to hydraulic resistance is unknown. We show that the cell phenotype shifts from amoeboid to mesenchymal upon increasing resistance. By combining automated morphological tracking and wavelet analysis along with fluorescence recovery after photobleaching (FRAP), we found an oscillatory phenotypic transition that cycles from blebbing to short, medium, and long actin network formation, and back to
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47

Seksek, Olivier, Joachim Biwersi, and A. S. Verkman. "Translational Diffusion of Macromolecule-sized Solutes in Cytoplasm and Nucleus." Journal of Cell Biology 138, no. 1 (1997): 131–42. http://dx.doi.org/10.1083/jcb.138.1.131.

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Fluorescence recovery after photobleaching (FRAP) was used to quantify the translational diffusion of microinjected FITC-dextrans and Ficolls in the cytoplasm and nucleus of MDCK epithelial cells and Swiss 3T3 fibroblasts. Absolute diffusion coefficients (D) were measured using a microsecond-resolution FRAP apparatus and solution standards. In aqueous media (viscosity 1 cP), D for the FITC-dextrans decreased from 75 to 8.4 × 10−7 cm2/s with increasing dextran size (4–2,000 kD). D in cytoplasm relative to that in water (D/Do) was 0.26 ± 0.01 (MDCK) and 0.27 ± 0.01 (fibroblasts), and independent
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48

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 (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 spatiotempo
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49

Vámosi, György, Elza Friedländer-Brock, Shehu M. Ibrahim, Roland Brock, János Szöllősi, and György Vereb. "EGF Receptor Stalls upon Activation as Evidenced by Complementary Fluorescence Correlation Spectroscopy and Fluorescence Recovery after Photobleaching Measurements." International Journal of Molecular Sciences 20, no. 13 (2019): 3370. http://dx.doi.org/10.3390/ijms20133370.

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To elucidate the molecular details of the activation-associated clustering of epidermal growth factor receptors (EGFRs), the time course of the mobility and aggregation states of eGFP tagged EGFR in the membranes of Chinese hamster ovary (CHO) cells was assessed by in situ mobility assays. Fluorescence correlation spectroscopy (FCS) was used to probe molecular movements of small ensembles of molecules over short distances and time scales, and to report on the state of aggregation. The diffusion of larger ensembles of molecules over longer distances (and time scales) was investigated by fluores
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50

Hansen, Jesper S., Nils J. Færgeman, Birthe B. Kragelund, and Jens Knudsen. "Acyl-CoA-binding protein (ACBP) localizes to the endoplasmic reticulum and Golgi in a ligand-dependent manner in mammalian cells." Biochemical Journal 410, no. 3 (2008): 463–72. http://dx.doi.org/10.1042/bj20070559.

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In the present study, we microinjected fluorescently labelled liver bovine ACBP (acyl-CoA-binding protein) [FACI-50 (fluorescent acyl-CoA indicator-50)] into HeLa and BMGE (bovine mammary gland epithelial) cell lines to characterize the localization and dynamics of ACBP in living cells. Results showed that ACBP targeted to the ER (endoplasmic reticulum) and Golgi in a ligand-binding-dependent manner. A variant Y28F/K32A-FACI-50, which is unable to bind acyl-CoA, did no longer show association with the ER and became segregated from the Golgi, as analysed by intensity correlation calculations. D
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