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Fluorescence recovery after photobleaching - Wikipedia, the free encyclopedia Fluorescence recovery after photobleaching (FRAP) is a technique used in cellular imaging where a fluorochrome attached to a molecule is destroyed on purpose with an intense flash of light (by a laser). GFP is a fluorescent protein that was isolated from jellyfish (Aequorea victoria). This technique is commonly used in conjunction with green fluorescent protein (GFP), fusion proteins, where the protein that is under study is fused to a GFP. en.wikipedia.org /wiki/Fluorescence_recovery_after_photobleaching   (231 words)

Fluorescence Recovery After Photobleaching (FRAP) FRAP is based on the principal of observing the rate of recovery of fluorescence due to the movement of a fluorescent marker into an area of the membrane which contains this same marker but which has been rendered non-fluorescent via an intense photobleaching pulse of laser light. FRAP is being used to measure the lateral diffusion of various membrane or cytoplasmic constituents. FRAP has proved to be a popular means to assess the structure of artificial and biological membranes. www-cellbio.med.unc.edu /facilities/frap.htm   (159 words)

mekentosj.com Science - Fluorescence Recovery After Photobleaching In short, the fluorescence in the region of interest is measured during the complete protocol; just before, ideally also during, and of course after photobleaching. Analysis of the fluorescence recovery can be used to determine kinetic parameters of a protein, including its diffusion constant, mobile fraction, transport rate or binding/dissociation rate from other proteins. Even after up to 180 minutes the fluorescence between nucleus and cytoplasm is not equalized, indicating that the proteasome is not diffusing between the two compartments. www.mekentosj.com /science/frap   (1260 words)

Olympus FluoView Resource Center: Confocal Microscope Product Information - PAPP for FRAP Application After the photobleaching pulse, the rate and extent of fluorescence intensity recovery in the bleached region is monitored as a function of time to generate information about repopulation by fluorophores and the kinetics of recovery. In the figure presented above, fluorescence recovery after photobleaching analysis was performed on a single cell to determine the diffusion rate of GFP-labeled proteins into the dendritic spines. In typical fluorescence recovery experiments, a measurement of average intensity is made as a function of time, as illustrated in the graph positioned on the right. www.olympusfluoview.com /products/pappproducts.html   (386 words)

Flourescence Recovery After Photobleaching (FRAP) In the diagram, the percentage of fluorescence lost due to photobleaching is X and the amount of fluorescence that returned to the bleached area is Y. In practice, the percent recovery almost never reaches 100%. The lateral mobility is determined by the slope of the curve (3). Later, there is a stabilization of the amount of fluorescence recovery (4) and a flat line is obtained. www.bio.davidson.edu /courses/Molbio/FRAPx/FRAP.html   (648 words)

Olympus FluoView Resource Center: Fluorescence Photobleaching Investigations A specific area of a floating fluorescent dye on a cell membrane, an organelle (endoplasmic reticulum and Golgi apparatus) membrane, or a floating fluorescence-labeled protein on these membranes is bleached, and the loss or recovery of fluorescence is observed to examine fluidity in the lateral direction (see Figure 1). A fluorescent dye previously conjugated with the target protein molecules or lipid membrane is bleached at the area of interest in the specimen and FRAP is used to observe how the fluorescence in the bleached portion recovers through dye turnover by means of diffusion. After the area of interest has been photobleached, fluorescence is rapidly recovered by means of diffusion. www.olympusfluoview.com /applications/flipandfrap.html   (418 words)

YGM 2002 Abstract #340 We have used fluorescence recovery after photobleaching (FRAP) to measure the turnover of GFP labeled tubulin (GFP-Tub1) within the mitotic spindle. Fluorescence Recovery After Photobleaching (FRAP) to Study the Kinetochore Microtubule Interaction. However, loss of chromosome attachment, generated by conditional alleles of the core kinetochore component ndc10-2, does not alter spindle microtubule fluorescence recovery after photobleaching. www.yeastgenome.org /yeast02/abshtml/340.html   (309 words)

Synaptic Vesicle Movements Monitored by Fluorescence Recovery after Photobleaching in Nerve Terminals Stained with FM1-43 -- Henkel et al. 16 (12): 3960 -- Journal of Neuroscience One simple explanation for the lack of recovery from photobleaching in resting nerve terminals is that the laser beam damaged the terminal and arrested vesicle movements that otherwise would have led to recovery from bleaching. Overall, 25-60 min after photobleaching there was only ~18% recovery of brightness in the bleached regions, an amount that, owing to slight amounts of focus change and photobleaching during image acquisition (see Materials and Methods), may not be different from zero (i.e., completely immobilized vesicles). In top views of both resting and stimulated nerve terminals, little or no recovery from photobleaching occurred, suggesting that synaptic vesicles are virtually immobile in resting terminals and that their lateral movements are restricted sharply as they make their way to the presynaptic membrane during repetitive nerve stimulation. www.jneurosci.org /cgi/content/full/16/12/3960   (6027 words)

Myosin II dynamics and cortical flow during contractile ring formation in Dictyostelium cells -- Yumura 154 (1): 137 -- The Journal of Cell Biology The fluorescence of nonbleached regions decreased in a reverse manner of recovery in B and diffused in the endoplasm. The large fluorescent spot near the center, which was an aggregate of filaments, was frequently observed in 3ALA myosin II cells (Egelhoff et al., 1993). manner of recovery and was diffused in the endoplasm. www.jcb.org /cgi/content/full/154/1/137   (5228 words)

A Fluorescence Recovery After Photobleaching (FRAP) Technique for the Measurement of Solute Transport Across Surfactant-Laden Interfaces The technique of Fluorescence Recovery After Photobleaching (FRAP) has been applied to the measurement of interfacial transport in two-phase systems. Browne, E.P., Hatton, T.A., A Fluorescence Recovery After Photobleaching (FRAP) Technique for the Measurement of Solute Transport Across Surfactant-Laden Interfaces, 3rd Microgravity Fluid Physics Conference, NASA Lewis Research Center, Cleveland, OH, CP 3338, pp. FRAP exploits the loss of fluorescence exhibited by certain fluorophores when over- stimulated (photobleached), so that a two-phase system, originally at equilibrium, can be perturbed without disturbing the interface by strong light from an argon-ion laser and its recovery monitored by a microscope-mounted CCD camera as it relaxes to a new equilibrium. microgravity.grc.nasa.gov /fcarchive/fluids/papers/Browne/A_Fluorescence.htm   (221 words)

Fluorescence recovery after photobleaching reveals that LPS rapidly transfers from CD14 to hsp70 and hsp90 on the cell membrane -- Triantafilou et al. 114 (13): 2535 -- Journal of Cell Science Fluorescence recovery after photobleaching reveals that LPS rapidly transfers from CD14 to hsp70 and hsp90 on the cell membrane -- Triantafilou et al. FRAP curve of hsp70 OG-Fab bound to MonoMac 6 (A) and ECV-304 cells (B) and measured at 22°C. FRAP curve of hsp90 OG-Fab bound to MonoMac 6 (C) and ECV-304 cells (D) and measured at 22°C. The best fit to the experimental data is shown. FRAP curve of FITC-LPS bound to CHO-CD14 cells (A,B) and measured at 22°C (A) or at 37°C (B). jcs.biologists.org /cgi/content/full/114/13/2535   (6495 words)

Diffusional Mobility of Parvalbumin in Spiny Dendrites of Cerebellar Purkinje Neurons Quantified by Fluorescence Recovery after Photobleaching -- Schmidt et al. 84 (4): 2599 -- Biophysical Journal Diffusional Mobility of Parvalbumin in Spiny Dendrites of Cerebellar Purkinje Neurons Quantified by Fluorescence Recovery after Photobleaching Diffusional Mobility of Parvalbumin in Spiny Dendrites of Cerebellar Purkinje Neurons Quantified by Fluorescence Recovery after Photobleaching -- Schmidt et al. www.biophysj.org /cgi/content/abstract/84/4/2599   (506 words)

fluorescence recovery after photobleaching method used to study lateral movements of membrane proteins and lipids; a small area of a cell membrane is bleached by laser light and the amount of time necessary for unbleached fluorescent marker tagged proteins to diffuse back into the bleached site is a measurement of the cell membrane's fluidity. crisp.cit.nih.gov /Thesaurus/00009832.htm   (50 words)

Confocal imaging and Frap: News from Bio-Rad Cell Science Division The use of Fluorescence recovery after photobleaching (Frap) to study the movement of a nucleolar protein in HeLa cells has been described in an application note published by Bio-Rad. If a small area of the cell is photobleached and the fluorescence intensity recovers, it indicates influx of the fluorescence-tagged molecule from the surrounding non-bleached area of the cell. Frap is a non-invasive technique with numerous applications in cell biology. www.laboratorytalk.com /news/bdc/bdc109.html   (440 words)

FLUORESCENCE RECOVERY AFTER PHOTOBLEACHING definition The rate and extent of recovery are a measure of the fluidity of the membrane and the proportion of labelled molecules that are free to exchange with adjacent areas. The recovery is due to the re population of the area by unbleached molecules and diffusion of bleached molecules to other areas. If, for example: the cell surface is labelled with a fluorescent probe and an area bleached by laser illumination, then the bleached patch that starts off as a dark area will gradually recover fluorescence. www.books.md /F/dic/fluorescencerecoveryafterphotobleaching.php   (254 words)

Analysis of Fluorophore Diffusion by Continuous Distributions of Diffusion Coefficients: Application to Photobleaching Measurements of Multicomponent and Anomalous Diffusion -- Periasamy and Verkman 75 (1): 557 -- Biophysical Journal Fluorescence recovery after photobleaching (FRAP) is widely used to measure fluorophore diffusion in artificial solutions Fluorescence photobleaching recovery measurement of protein absolute diffusion coefficients. Analysis of fluorescence decay by the maximum entropy method: influence of noise and analysis parameters on the width of the distribution of lifetimes. www.biophysj.org /cgi/content/full/75/1/557   (5047 words)

Rapid assembly dynamics of the Escherichia coli FtsZ-ring demonstrated by fluorescence recovery after photobleaching -- Stricker et al. 99 (5): 3171 -- Proceedings of the National Academy of Sciences FRAP, fluorescence recovery after photobleaching; GTPase, guanosine triphosphatase; gfp, green fluorescent protein. The cell was photobleached at the arrow and the recovery of fluorescence was monitored. Half of the Z-ring of the cell on the left was laser photobleached at the arrow, and recovery of fluorescence was monitored. www.pnas.org /cgi/content/full/99/5/3171   (4020 words)

Intracellular Macromolecular Mobility Measured by Fluorescence Recovery after Photobleaching with Confocal Laser Scanning Microscopes -- Braga et al. 15 (10): 4749 -- Molecular Biology of the Cell Klonis, N., Rug, M., Harper, I., Wickham, M., Cowman, A., and Tilley, L. Fluorescence photobleaching analysis for the study of cellular dynamics. Sprague, B.L., Pego, R.L., Stavreva, D.A., and McNally, J.G. Analysis of binding reactions by fluorescence recovery after photobleaching. For the 500-kDa-dextran, the fluorescence intensity is reduced from 1 to 0.4, whereas for the 40-kDa-dextran the reduction is from 1 to 0.7. www.molbiolcell.org /cgi/content/full/15/10/4749   (6446 words)

Direct Imaging of Dehydrogenase Activity within Living Cells Using Enzyme-Dependent Fluorescence Recovery after Photobleaching (ED-FRAP) -- Combs and Balaban 80 (4): 2018 -- Biophysical Journal Mean and SEM values for the exponential rate constant (averaged over the entire cell volume) of recovery after photobleaching versus treatment for superfused cardiac myocytes. Fluorescence correlation spectroscopy and photobleaching recovery of multiple binding reactions. Note that the recovery is exponential and similar to the recovery curve in Fig. www.biophysj.org /cgi/content/full/80/4/2018   (4469 words)

Analysis of Binding Reactions by Fluorescence Recovery after Photobleaching -- Sprague et al. 86 (6): 3473 -- Biophysical Journal Recovery times for typical FRAP experiments are generally between 1 and 1000 s, indicating that reaction dominant, effective diffusion, or full model behavior are all possible. The large amount of fluorescence retained at the nuclear matrix in azide-treated cells matches the two-state full model prediction that 44% of GFP-GR in the nucleus is bound in a second reaction state. FRAP recovery data is the sum of free and bound fluorescence, www.biophysj.org /cgi/content/full/86/6/3473   (8427 words)

4. The Plasma Membrane The fluorescent lipids moved freely in the membrane of the axon, however, they were excluded from the cell body and the dendrites (Kobayashi et al., 1992). After an initial reduction of the label, any subsequent reduction must be the result of a flip-flop from the inner layer and can be used to measure of the rate of flip-flop. The return of fluorescence is then measured to give an estimate of the diffusion of the fluorophores labelled components in the bleached area which allows calculating the fraction of diffusible proteins and the diffusion coefficient. www.albany.edu /~abio304/text/4part2.html   (10415 words)

Fluorescence Resources Fluorescence probes and conjugations are used extensively to trace the whereabouts of cellular components and protein localisation, it is used in fluorescence spectroscopy and a host of more complex applications (that give rise to great abbreviations) such as Fluorescence Resonance Energy Transfer (FRET), Fluorescence Recovery After Photobleaching (FRAP) and Fluorescence Activated Cell Sorting (FACS). Fluorescence is a phenomenon that is used routinely in life science research. Fluorescence Polarisation Guide provided in PDF format a comprehensive guide by PanVera describes receptor-ligand binding, immunoassays, DNA hybridization, theory of binding data analysis and more. www.fluorescence-resource.com   (371 words)

Dynamic analyses of lymphoblast membranes exposed to alpha interferon using flow cytometry and fluorescence recovery after photobleaching. In a second approach, changes in diffusion coefficients of plasma membrane-associated macromolecules were determined by measuring the fluorescence redistribution after pulse photobleaching (FRAP): Individual plasma membrane proteins (sIgM, Leu 12 or Leu 16) were labelled with FITC conjugated goat antibodies [F(ab')2 or Fab'] or with phycoerythrin-B conjugated monoclonal mouse antibodies. Dynamic analyses of lymphoblast membranes exposed to alpha interferon using flow cytometry and fluorescence recovery after photobleaching. In one approach, changes in plasma membrane ion flux were measured by flow cytometry, using a fluorescent dye indicator of membrane potential: Cells briefly exposed (5-10 min) to a DNA-recombinant IFN-alpha 2 (100 to 800 U/ml) manifested a consistent plasma membrane hyperpolarization (-60 to -90 mV) which could be blocked by ouabain. www.pdg.cnb.uam.es /UniPub/iHOP/gp/5861473.html   (297 words)

Energy Citations Database (ECD) - Energy and Energy-Related Bibliographic Citations We report a polarized fluorescence recovery after photobleaching (pFRAP) method to measure the rotational dynamics of fluorescent colloids over a wide dynamic range. The method is based on the polarization anisotropy in the fluorescence intensity, generated by bleaching of fluorescently labeled particles with an intense pulse of linearly polarized laser light. The performance of the equipment is demonstrated for fluorescent colloidal silica spheres, dispersed in pure solvents as well as in fd-virus suspensions. www.osti.gov /energycitations/product.biblio.jsp?osti_id=20528559   (313 words)

Age-estimations of rats based on the average lateral diffusion constant of hepatocyte membrane proteins as revealed by fluorescence recovery after photobleaching. Age-estimations of rats based on the average lateral diffusion constant of hepatocyte membrane proteins as revealed by fluorescence recovery after photobleaching. A method has been developed recently for measuring the average lateral diffusion constant of the proteins (D) in the cell membrane of hepatocytes in liver smears by fluorescence recovery after photobleaching (FRAP). A peroxide-induced autofluorescence (PIAF) of the membrane proteins was used as a fluorescent label. www.arclab.org /medlineupdates/abstract_3556455.html   (180 words)

Fluorescence recovery after photobleaching rightthumb320pxPrinciple of FRAP Fluorescence recovery after photobleaching (FRAP) is a technique used in cellular imaging where a fluorochrome attached to a molecule is destroyed on purpose with an intense flash of light (by a laser) and this in a well defined area to study the repopulating of this area with peripheral molecules still fluorescent. Sorry, no screened links relevant to Fluorescence recovery after photobleaching were found: This methods allows measurement of the speed of diffusion of molecules in living cells. www.omniknow.com /common/wiki.php?in=en&term=FRAP   (272 words)

atto bioscience BD™ CARV II High speed multi-point confocal scanning, combined with high quantum efficiency CCD cameras, minimizes photobleaching and allows real-time imaging and recording at up to 100 fps. The BD™ CARV II Confocal Imager also offers automation of fluorescence recovery after photobleaching (FRAP). Automation of internal multi-position excitation, dichroic and emission filter wheels permits fast multi-dimensional imaging of up to five or more fluorescent probes in the same sample. www.atto.com /products/carv   (175 words)

Applied Precision, LLC - Life Sciences - DeltaVision RT - Quantitative Laser Module iFRAP (Instantaneous Fluorescence Recovery After Photobleaching), FRET (Fluorescent Resonance Energy Transfer), Must have a key switch to prevent unauthorized use, a laser emission indicator, a 3 to 5 second time delay after power is applied, and a mechanical shutter to turn the beam off during use. Quantitative analysis of recovery - T½ and Mobile Fraction - Graphical and numerical results www.api.com /lifescience/dv-QLM.html   (256 words)

FRAP Typical FRAP curve obtained with the institute's FRAP setup. The background image shows the measured, flourescently labelled bilayer. www.mpip-mainz.mpg.de /~naumannr/popups/FRAP.html   (18 words)

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