Characterization of lipid bilayer phases by confocal microscopy and fluorescence correlation spectroscopy Export

@article{citeulike:4080343, abstract = {We report the application of confocal imaging and fluorescence correlation spectroscopy (FCS) to characterize chemically well-defined lipid bilayer models for biomembranes. Giant unilamellar vesicles of dilauroyl phosphatidylcholine/dipalmitoyl phosphatidylcholine (DLPC/DPPC)/cholesterol were imaged by confocal fluorescence microscopy with two fluorescent probes, 1,1′-dieicosanyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI-C) and 2-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza--indacene-3-pentanoyl)-1-hexadecanoyl--glycero-3-phosphocholine (Bodipy-PC). Phase separation was visualized by differential probe partition into the coexisting phases. Three-dimensional image reconstructions of confocal -scans through giant unilamellar vesicles reveal the anisotropic morphology of coexisting phase domains on the surface of these vesicles with full two-dimensional resolution. This method demonstrates by direct visualization the exact superposition of like phase domains in apposing monolayers, thus answering a long-standing open question. Cholesterol was found to induce a marked change in the phase boundary shapes of the coexisting phase domains. To further characterize the phases, the translational diffusion coefficient, , of the DiI-C was measured by FCS. values at ∼25°C ranged from ∼ 3 × 10 cm/s in the fluid phase, to ∼ 2 × 10 cm/s in high-cholesterol-content phases, to ∼ 2 × 10 cm/s in the spatially ordered phases that coexist with fluid phases. In favorable cases, FCS could distinguish two different values of in a region of two-phase coexistence on a single vesicle.}, author = {Korlach, Jonas and Schwille, Petra and Webb, Watt W. and Feigenson, Gerald W.}, citeulike-article-id = {4080343}, citeulike-linkout-0 = {http://www.pnas.org/content/96/15/8461.abstract}, citeulike-linkout-1 = {http://www.pnas.org/content/96/15/8461.full.pdf}, citeulike-linkout-2 = {http://view.ncbi.nlm.nih.gov/pubmed/10411897}, citeulike-linkout-3 = {http://www.hubmed.org/display.cgi?uids=10411897}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, keywords = {bilayer, correlation, fcs, fluorescence, korlac99pdf, lipid, microscopy, phase}, month = {July}, number = {15}, pages = {8461--8466}, posted-at = {2009-07-03 20:28:10}, priority = {2}, title = {Characterization of lipid bilayer phases by confocal microscopy and fluorescence correlation spectroscopy}, url = {http://www.pnas.org/content/96/15/8461.abstract}, volume = {96}, year = {1999} }

TY - JOUR ID - citeulike:4080343 L3 - citeulike-article-id:4080343 N2 - We report the application of confocal imaging and fluorescence correlation spectroscopy (FCS) to characterize chemically well-defined lipid bilayer models for biomembranes. Giant unilamellar vesicles of dilauroyl phosphatidylcholine/dipalmitoyl phosphatidylcholine (DLPC/DPPC)/cholesterol were imaged by confocal fluorescence microscopy with two fluorescent probes, 1,1′-dieicosanyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiI-C) and 2-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza--indacene-3-pentanoyl)-1-hexadecanoyl--glycero-3-phosphocholine (Bodipy-PC). Phase separation was visualized by differential probe partition into the coexisting phases. Three-dimensional image reconstructions of confocal -scans through giant unilamellar vesicles reveal the anisotropic morphology of coexisting phase domains on the surface of these vesicles with full two-dimensional resolution. This method demonstrates by direct visualization the exact superposition of like phase domains in apposing monolayers, thus answering a long-standing open question. Cholesterol was found to induce a marked change in the phase boundary shapes of the coexisting phase domains. To further characterize the phases, the translational diffusion coefficient, , of the DiI-C was measured by FCS. values at ∼25°C ranged from ∼ 3 × 10 cm/s in the fluid phase, to ∼ 2 × 10 cm/s in high-cholesterol-content phases, to ∼ 2 × 10 cm/s in the spatially ordered phases that coexist with fluid phases. In favorable cases, FCS could distinguish two different values of in a region of two-phase coexistence on a single vesicle. IS - 15 JF - Proceedings of the National Academy of Sciences of the United States of America EP - 8466 TI - Characterization of lipid bilayer phases by confocal microscopy and fluorescence correlation spectroscopy VL - 96 SP - 8461 KW - bilayer KW - correlation KW - fcs KW - fluorescence KW - korlac99pdf KW - lipid KW - microscopy KW - phase AU - Korlach, Jonas AU - Schwille, Petra AU - Webb, Watt AU - Feigenson, Gerald PY - 1999/07/20/ UR - http://www.pnas.org/content/96/15/8461.abstract ER -