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Unilamellar Vesicles (unilamellar + vesicle)
Kinds of Unilamellar Vesicles Selected AbstractsAnalysis of several fluorescent detector molecules for protein microarray useLUMINESCENCE: THE JOURNAL OF BIOLOGICAL AND CHEMICAL LUMINESCENCE, Issue 1 2003Rick Wiese Abstract The utility of several streptavidin-linked fluorescent detector molecules was evaluated on two protein microarray platforms. Tested detector molecules included: Alexa Fluor 546; R-phycoerythrin (RPE), orange fluospheres; Cy3-containing liposomes (Large Unilamellar Vesicles, LUV) labelled with Cy3; and an RPE,antibody complex. The two array architectures tested consisted of an array of murine Fc,biotin and an array of murine IgG (the murine IgG array was probed with a biotinylated rabbit anti-murine IgG). These platforms allowed for the direct comparison of detector utility by detector recognition of array-bound biotin. All of the fluorescent detectors examined demonstrated utility on each of the array platforms. For the Fc,biotin array, detector signal intensity (background adjusted) was as follows: RPE,antibody complex,>,fluospheres,>,RPE,>,liposomes,>,Alexa 546: for the IgG array: RPE/antibody complex,>,RPE,>,fluospheres,>,Alexa546,>,liposomes. The RPE,antibody complex fluoresced 67% and 150% more intensely than the next closest detector molecule for the Fc,biotin and the murine IgG arrays, respectively. A marked increase in background fluorescence (as compared to RPE alone) did not accompany the increase in signal intensity gained through RPE,antibody complex use (a true increase in signal:noise ratio). These results suggest that the RPE,antibody complex is superior to other molecules for fluorescent detection of analytes on protein microarrays. Copyright © 2002 John Wiley & Sons, Ltd. [source] Microfluidic Formation of Monodisperse, Cell-Sized, and Unilamellar Vesicles,ANGEWANDTE CHEMIE, Issue 35 2009Sadao Ota Sanft dem Strom hinab: Mit einer Mikrofluidiktechnik werden in einem kontinuierlichen Fluidstrom monodisperse unilamellare Phospholipidvesikel aus einer einzigen Doppelschicht erzeugt (siehe Bild). Da die Vesikel robust sind und effizient eine Vielzahl an Molekülen in hohen Konzentrationen einkapseln, eignen sie sich als Transportvehikel und als Modelle für Zellsysteme. [source] Compositional effects on electrophoretic and chromatographic figures of merit in electrokinetic chromatography with cetyltrimethylammonium bromide/sodium octyl sulfate vesicles as the pseudostationary phase.ELECTROPHORESIS, Issue 5 2008Part 1: Effect of the phase ratio Abstract The effect of the phase ratio on the electrophoretic and chromatographic properties of unilamellar vesicles comprised of cetyltrimethylammonium bromide (CTAB) and sodium octyl sulfate (SOS) was investigated in EKC. The surfactant concentration of the vesicles was 0.9, 1.2, 1.5, and 1.8% w/v, with a mole ratio of 1:3.66 (CTAB/SOS). Results were compared to those obtained using SDS micelles at concentrations of 1.0% (w/v, 35,mM) and 1.5% (52,mM). The CTAB/SOS vesicles (0.9,1.8% w/v) provided a significantly larger elution range (5.7,,,tves/t0,,,8.7) and greater hydrophobic (methylene) selectivity (2.8,,,,CH2,,,3.1) than SDS micelles (3.1,,,tmc/t0,,,3.3; ,CH2,=,2.2). Whereas the larger elution range can be attributed to the 25% reduction in EOF due to the interaction of unaggregated CTAB cations and the negatively charged capillary wall, the higher methylene selectivity is likely due to the lower concentration of water expected in the CTAB/SOS vesicle bilayer compared to the Palisades layer of SDS micelles. For a given phase ratio, CTAB/SOS vesicles are somewhat less retentive than SDS micelles, although retention factors comparable to those observed in 1.0,1.5% SDS can be obtained with 1.5,1.8% CTAB/SOS. A linear relationship was observed between phase ratio and retention factor, confirming the validity of the phase ratio model for these vesicles. Unique polar group selectivities and positional isomer shape selectivities were obtained with CTAB/SOS vesicles, with both types of selectivities being nearly independent of the phase ratio. For four sets of positional isomers, the elution order was always para < ortho < meta. Finally, the thermodynamics of solute retention was qualitatively similar to that reported for other surfactant aggregates (micelles and microemulsions); the enthalpic contribution to retention was consistently favorable for all compounds, whereas the entropic contribution was favorable only to hydrophobic solutes. [source] The assembly factor P17 from bacteriophage PRD1 interacts with positively charged lipid membranesFEBS JOURNAL, Issue 20 2000Juha M. Holopainen The interactions of the assembly factor P17 of bacteriophage PRD1 with liposomes were investigated by static light scattering, fluorescence spectroscopy, and differential scanning calorimetry. Our data show that P17 binds to positively charged large unilamellar vesicles composed of the zwitterionic 1-palmitoyl-2-oleoyl-phosphatidylcholine and sphingosine, whereas only a weak interaction is evident for 1-palmitoyl-2-oleoyl-phosphatidylcholine vesicles. P17 does not bind to negatively charged membranes composed of 1-palmitoyl-2-oleoyl-phosphatidylglycerol and 1-palmitoyl-2-oleoyl-phosphatidylcholine. Our differential scanning calorimetry results reveal that P17 slightly perturbs the phase behaviour of neutral phosphatidylcholine and negatively charged multilamellar vesicles. In contrast, the phase transition temperature of positively charged dimyristoylphosphatidylcholine/sphingosine multilamellar vesicles (molar ratio 9 : 1, respectively) is increased by approximately 2.4 °C and the half width of the enthalpy peak broadened from 1.9 to 5.6 °C in the presence of P17 (protein : lipid molar ratio 1 : 47). Moreover, the enthalpy peak is asymmetrical, suggesting that lipid phase separation is induced by P17. Based on the far-UV CD spectra, the ,-helicity of P17 increases upon binding to positively charged micelles composed of Triton X-100 and sphingosine. We propose that P17 can interact with positively charged lipid membranes and that this binding induces a structural change on P17 to a more tightly packed and ordered structure. [source] Plant fructans stabilize phosphatidylcholine liposomes during freeze-dryingFEBS JOURNAL, Issue 2 2000Dirk K. Hincha Fructans have been implicated as protective agents in the drought and freezing tolerance of many plant species. A direct proof of their ability to stabilize biological structures under stress conditions, however, is still lacking. Here we show that inulins (linear fructose polymers) isolated from chicory roots and dahlia tubers stabilize egg phosphatidylcholine large unilamellar vesicles during freeze-drying, while another polysaccharide, hydroxyethyl starch, was completely ineffective. Liposome stability was assessed after rehydration by measuring retention of the soluble fluorescent dye carboxyfluorescein and bilayer fusion. Inulin was an especially effective stabilizer in combination with glucose. Analysis by HPLC showed that the commercial inulin preparations used in our study contained no low molecular mass sugars that could be responsible for the observed stabilizing effect of the fructans. Fourier transform infrared spectroscopy showed a reduction of the gel to liquid-crystalline phase transition temperature of dry egg PtdCho by more than 20 °C in the presence of inulin. A direct interaction of inulin with the phospholipid in the dry state was also indicated by dramatic differences in the phosphate asymmetric stretch region of the infrared spectrum between samples with and without the polysaccharide. [source] Method of separated form factors for polydisperse vesiclesJOURNAL OF APPLIED CRYSTALLOGRAPHY, Issue 3 2006Jeremy Pencer Use of the Schulz or Gamma distribution in the description of particle sizes facilitates calculation of analytic polydisperse form factors using Laplace transforms, [f(u)]. Here, the Laplace transform approach is combined with the separated form factor (SFF) approximation [Kiselev et al. (2002). Appl. Phys. A, 74, S1654,S1656] to obtain expressions for form factors, P(q), for polydisperse spherical vesicles with various forms of membrane scattering length density (SLD) profile. The SFF approximation is tested against exact form factors that have been numerically integrated over the size distribution, and is shown to represent the vesicle form factor accurately for typical vesicle sizes and membrane thicknesses. Finally, various model SLD profiles are used with the SFF approximation to fit experimental small-angle neutron scattering (SANS) curves from extruded unilamellar vesicles. [source] Spatial fluorescence cross-correlation spectroscopy by means of a spatial light modulatorJOURNAL OF BIOPHOTONICS, Issue 5 2008Yoann Blancquaert Abstract Spatial fluorescence cross-correlation spectroscopy is a rarely investigated version of fluorescence correlation spectroscopy, in which the fluorescence signals from different observation volumes are cross-correlated. In the reported experiments, two observation volumes, typically shifted by a few ,m, are produced, with a spatial light modulator and two adjustable pinholes. We illustrated the feasibility and potentiality of this technique by: i) measuring molecular flows, in the range 0.2,1.5 ,m/ms, of solutions seeded with fluorescent nanobeads or rhodamine molecules (simulating active transport phenomenons); ii) investigating the permeability of the phospholipidic membrane of giant unilamellar vesicles versus hydrophilic or hydrophobic molecules (in that case the laser spots were set on both sides of the membrane). Theoretical descriptions are proposed together with a discussion about fluorescence-correlation-spectroscopy-based, alternative methods. (© 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] Detection of single lipid bilayers with coherent anti-Stokes Raman scattering (CARS) microscopyJOURNAL OF RAMAN SPECTROSCOPY, Issue 9 2003Eric O. Potma Abstract We investigated vibrational imaging of phospholipid bilayers with CARS microscopy. Single lipid membranes of supported bilayers, giant unilamellar vesicles and intact erythrocyte membrane are detected with the strong resonant signal of the C,H stretching vibration. In addition, it is shown that the CARS signal field of the lipids near the glass/water interface is amplified through mixing with the back-reflected non-resonant CARS field of the glass coverslip. Furthermore, interference effects between two separate bilayers are observed, allowing intermembrane distances to be determined beyond the diffraction-limited resolution of the microscope. Copyright © 2003 John Wiley & Sons, Ltd. [source] Giant Vesicles: Preparations and ApplicationsCHEMBIOCHEM, Issue 7 2010Peter Walde Prof. Dr. Abstract There is considerable interest in preparing cell-sized giant unilamellar vesicles from natural or nonnatural amphiphiles because a giant vesicle membrane resembles the self-closed lipid matrix of the plasma membrane of all biological cells. Currently, giant vesicles are applied to investigate certain aspects of biomembranes. Examples include lateral lipid heterogeneities, membrane budding and fission, activities of reconstituted membrane proteins, or membrane permeabilization caused by added chemical compounds. One of the challenging applications of giant vesicles include gene expressions inside the vesicles with the ultimate goal of constructing a dynamic artificial cell-like system that is endowed with all those essential features of living cells that distinguish them from the nonliving form of matter. Although this goal still seems to be far away and currently difficult to reach, it is expected that progress in this and other fields of giant vesicle research strongly depend on whether reliable methods for the reproducible preparation of giant vesicles are available. The key concepts of currently known methods for preparing giant unilamellar vesicles are summarized, and advantages and disadvantages of the main methods are compared and critically discussed. [source] 4-Fluorophenylglycine as a Label for 19F NMR Structure Analysis of Membrane-Associated PeptidesCHEMBIOCHEM, Issue 11 2003Sergii Afonin Abstract The non-natural amino acid 4-fluorophenylglycine (4F-Phg) was incorporated into several representative membrane-associated peptides for dual purpose. The 19F-substituted ring is directly attached to the peptide backbone, so it not only provides a well-defined label for highly sensitive 19F NMR studies but, in addition, the D and L enantiomers of the stiff side chain may serve as reporter groups on the transient peptide conformation during the biological function. Besides peptide synthesis, which is accompanied by racemisation of 4F-Phg, we also describe separation of the epimers by HPLC and removal of trifluoroacetic acid. As a first example, 18 different analogues of the fusogenic peptide "B18" were prepared and tested for induction of vesicle fusion; the results confirmed that hydrophobic sites tolerated 4F-Phg labelling. Similar fusion activities within each pair of epimers suggest that the peptide is less structured in the fusogenic transition state than in the helical ground state. In a second example, five doubly labelled analogues of the antimicrobial peptide gramicidin S were compared by using bacterial growth inhibition assays. This cyclic ,-sheet peptide could accommodate both L and D substituents on its hydrophobic face. As a third example, we tested six analogues of the antimicrobial peptide PGLa. The presence of d- 4F-Phg reduced the biological activity of the peptide by interfering with its amphiphilic ,-helical fold. Finally, to illustrate the numerous uses of l- 4F-Phg in 19F NMR spectroscopy, we characterised the interaction of labelled PGLa with uncharged and negatively charged membranes. Observing the signal of the free peptide in an aqueous suspension of unilamellar vesicles, we found a linear saturation behaviour that was dominated by electrostatic attraction of the cationic PGLa. Once the peptide is bound to the membrane, however, solid-state 19F NMR spectroscopy of macroscopically oriented samples revealed that the charge density has virtually no further influence on the structure, alignment or mobility of the peptide. [source] Design of a pH-sensitive pore-forming peptide with improved performanceCHEMICAL BIOLOGY & DRUG DESIGN, Issue 1 2004D.H. Haas Abstract:, GALA is a 30 residue synthetic peptide designed to interact with membranes in a pH-sensitive manner, with potential applications for intracellular drug and gene delivery. Upon reduction of the pH from neutral to acidic, GALA switches from random coil to , -helix, inserts into lipid bilayers, and forms oligomeric pores of defined size. Its simple sequence and well-characterized behavior make the peptide an excellent starting point to explore the effects of sequence on structure, pH sensitivity, and membrane affinity. We describe synthesis and characterization of two derivatives of GALA, termed GALAdel3E and YALA. GALAdel3E has a deletion of three centrally located glutamate residues from GALA, while YALA replaces one glutamate residue with the unusual amino acid 3,5-diiodotyrosine. Both derived peptides retain pH sensitivity, showing no ability to cause leakage of an encapsulated dye from unilamellar vesicles at pH 7.4 but substantial activity at pH 5. Unlike GALA, neither peptide undergoes a conformational change upon reduction of the pH, remaining , -helical throughout. Interestingly, the pH at which the peptides activate is shifted, with GALA becoming active at pH ,5.7, GALAdel3E at pH ,6.2, and YALA at pH ,6.7. Furthermore, the peptides GALAdel3E and YALA show improved activity compared with GALA for cholesterol-containing membranes, with YALA retaining the greatest activity. Improved activity in the presence of cholesterol and onset of activity in the critical range between pH 6 and 7 may make these peptides useful in applications requiring intracellular delivery of macromolecules, such as gene delivery or anti-cancer treatments. [source] Probing Biomembrane Dynamics by Dual-Wavelength Reflection Interference Contrast MicroscopyCHEMPHYSCHEM, Issue 16 2009Cornelia Monzel Abstract We present an improved analysis of reflection interference contrast microscopy (RICM) images, recorded to investigate model membrane systems that mimic cell adhesion. The model systems were giant unilamellar vesicles (GUV) adhering via specific ligand,receptor interactions to supported lipid bilayers (SLB) or to patterns of receptors. Conventional RICM and dual-wavelength RICM (DW,RICM) were applied to measure absolute optical distances between the biomembranes and planar substrates. We developed algorithms for a straightforward implementation of an automated, time-resolved reconstruction of the membrane conformations from RICM/DW,RICM images, taking into account all the interfaces in the system and blurring of the data due to camera noise. Finally, we demonstrate the validity and usefulness of this new approach by analyzing the topography and fluctuations of a bound membrane in the steady state and its dynamic adaptation to osmotic pressure changes. These measurements clearly show that macroscopic membrane flow through tightly adhered area is possible in our system. [source] | |||||||||||||