Mobility measurement by analysis of fluorescence photobleaching recovery kinetics
- PMID: 786399
- PMCID: PMC1334945
- DOI: 10.1016/S0006-3495(76)85755-4
Mobility measurement by analysis of fluorescence photobleaching recovery kinetics
Abstract
Fluorescence photobleaching recovery (FPR) denotes a method for measuring two-dimensional lateral mobility of fluorescent particles, for example, the motion of fluorescently labeled molecules in approximately 10 mum2 regions of a single cell surface. A small spot on the fluorescent surface is photobleached by a brief exposure to an intense focused laser beam, and the subsequent recovery of the fluorescence is monitored by the same, but attenuated, laser beam. Recovery occurs by replenishment of intact fluorophore in the bleached spot by lateral transport from the surrounding surface. We present the theoretical basis and some practical guidelines for simple, rigorous analysis of FPR experiments. Information obtainable from FPR experiments includes: (a) identification of transport process type, i.e. the admixture of random diffusion and uniform directed flow; (b) determination of the absolute mobility coefficient, i.e. the diffusion constant and/or flow velocity; and (c) the fraction of total fluorophore which is mobile. To illustrate the experimental method and to verify the theory for diffusion, we describe some model experiments on aqueous solutions of rhodamine 6G.
Similar articles
-
Measurement of molecular diffusion in solution by multiphoton fluorescence photobleaching recovery.Biophys J. 1999 Nov;77(5):2837-49. doi: 10.1016/S0006-3495(99)77115-8. Biophys J. 1999. PMID: 10545381 Free PMC article.
-
Measuring surface dynamics of biomolecules by total internal reflection fluorescence with photobleaching recovery or correlation spectroscopy.Biophys J. 1981 Mar;33(3):435-54. doi: 10.1016/S0006-3495(81)84905-3. Biophys J. 1981. PMID: 7225515 Free PMC article.
-
Measurement of lateral transport on cell surfaces.Prog Clin Biol Res. 1976;9:137-47. Prog Clin Biol Res. 1976. PMID: 1030797
-
Interpretation of fluorescence correlation spectroscopy and photobleaching recovery in terms of molecular interactions.Methods Cell Biol. 1989;30:307-32. doi: 10.1016/s0091-679x(08)60984-x. Methods Cell Biol. 1989. PMID: 2648114 Review.
-
Mobility of cell surface receptors: a re-evaluation.FEBS Lett. 1998 Jun 23;430(1-2):88-91. doi: 10.1016/s0014-5793(98)00595-x. FEBS Lett. 1998. PMID: 9678600 Review.
Cited by
-
A comparative analysis of the mobility of 45 proteins in the synaptic bouton.EMBO J. 2020 Aug 17;39(16):e104596. doi: 10.15252/embj.2020104596. Epub 2020 Jul 6. EMBO J. 2020. PMID: 32627850 Free PMC article.
-
Dissection of cell cycle-dependent dynamics of Dnmt1 by FRAP and diffusion-coupled modeling.Nucleic Acids Res. 2013 May;41(9):4860-76. doi: 10.1093/nar/gkt191. Epub 2013 Mar 27. Nucleic Acids Res. 2013. PMID: 23535145 Free PMC article.
-
Multiscale diffusion in the mitotic Drosophila melanogaster syncytial blastoderm.Proc Natl Acad Sci U S A. 2012 May 29;109(22):8588-93. doi: 10.1073/pnas.1204270109. Epub 2012 May 16. Proc Natl Acad Sci U S A. 2012. PMID: 22592793 Free PMC article.
-
Time-Resolved X-ray Observation of Intracellular Crystallized Protein in Living Animal.Int J Mol Sci. 2023 Nov 29;24(23):16914. doi: 10.3390/ijms242316914. Int J Mol Sci. 2023. PMID: 38069236 Free PMC article.
-
Hydration- and Temperature-Dependent Fluorescence Spectra of Laurdan Conformers in a DPPC Membrane.Cells. 2024 Jul 23;13(15):1232. doi: 10.3390/cells13151232. Cells. 2024. PMID: 39120265 Free PMC article.
References
Publication types
MeSH terms
LinkOut - more resources
Full Text Sources
Other Literature Sources