Fluorescence recovery after photo-bleaching

FRAP (fluorescence recovery after photo bleaching) analysis was originally developed in the mid-1970s to study the diffusion of biomolecules in living cells (Edidin et al, 1976). Flowever, due to the increased availability of GFP tags and advances in the bleaching capabilities of confocal microscopes, this... 

Fluorescein-labeled proteins are also used to measure the translational mobility of proteins and lipids by the Fluorescence Recovery After Photo-bleaching technique [54-59]. The uniformly labeled fluorescent sample is flashed with an intense light source to bleach a spot, thus producing a concentration gradient. The rate of recovery of fluorescence in that bleached area is measured and used to calculate the diffusion coefficient of the probe dye into the bleached zone. Such diffusion coefficient measurements have been used to determine the association constants of proteins in cells [60], to measure the exchange of tubulin between the cytoplasm and the microtubules [61,62], to study the polymerization-depolymerization process of actin [63-65] and to monitor the changes that occur upon cell maturation [66,67]. 

Slomiany B.L. and Slomiany A. (2002) Disruption in gastric mucin synthesis by Helicobacter pylori lipopolysaccharide involves ERK and P38 mitogen-activated protein kinase participation. Biochem Biophys Res Commun 294, 220-224 Snapp E., Altan N. and Lippincott-Schwartz J. (2003) Fluorescence Recovery After Photo-bleaching. In Current Protocols in Cell Biology. (Bonifacino, J., Dasso, M., Harford, J., Lippincott-Schwartz, J., Yamada K., Morgan K.S., eds.) Unit 21.1 John Wiley Sons, Inc., New York... 

Self-assembly Surface plasmon resonance spectroscopy Electrochemical impedance spectroscopy Neutron reflectometry Fluorescence recovery after photo bleaching (FRAP)... 

Sbalzarini, I. F., Mezzacasa,A., Helenius,A., and Koumoutsakos, P. (2005) Effects of organelle shape on fluorescence recovery after photo-bleaching. Biophys.J. 89,1482-1492. 

Silicon substrates have been covered by a closed S-layer lattice and subsequently bilayers have been deposited by the Langmuir-Blodgett technique [142-144]. The lateral diffusion of fluorescently labeled lipid molecules in both layers has been investigated by fluorescence recovery after photo-bleaching studies [145]. In comparison with hybrid lipid bilayers (lipid monolayer on alkylsilanes) and lipid bilayers on dextran, the mobility of lipids is highest in S-layer-supported bilayers. Furthermore, an S-layer cover causes an enhanced mobility of the labeled molecules in the adjacent lipid layer. In addition, the S-layer cover can also prevent the formation of cracks and other inhomogenities in the bilayer [21,145]. 

Measurement of Self-Diffusion Coefficient What is measured in DLS is D. DLS cannot measure because it does not distinguish one solute from another. It is necessary to use other more specialized techniques such as forced Rayleigh scattering (FRS), fluorescence recovery after photo-bleaching (FRAP), and pulsed-field gradient nuclear magnetic resonance (PFG-NMR). 

Jourdainne L, Lecuyer S, Arntz Y et al (2008) Dynamics of poly(L-lysine) in hyaluronic acid/poly(L-lysine) multilayer films studied by fluorescence recovery after pattern photo-bleaching. Langmuir 24 7842-7847... 

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