Vesicle Adsorption and Lipid Bilayer Formation on Glass Studied by Atomic Force Microscopy

Abstract

The adsorption of phosphatidylcholine (PC) vesicles (30, 50, and 100 nm nominal diameters) and of dye-labeled PC vesicles (labeled with 6% Texas Red fluorophore (TR) and encapsulated carboxy fluorescein (CF)) to glass surfaces was studied by contact mode atomic force microscopy in aqueous buffer. These studies were performed in part to unravel details of the previously observed isolated rupture of dye-labeled PC vesicles on glass (Johnson, J. M.; Ha, T.; Chu, S.; Boxer, S. G. Biophys. J. 2002, 83, 3371-3379), specifically to differentiate partial rupture, that is, pore formation and leakage of entrapped dye, from full rupture to form bilayer disks. In addition, the adhesion potential of PC vesicles on glass was calculated based upon the adhesion-driven flattening of adsorbed vesicles and a newly developed theoretical model. The vesicles were found to flatten considerably upon adsorption to glass (width-to-height ratio of approximately 5), which leads to an estimate for the adhesion potential and for the critical rupture radius of 1.5 x 10(-4) J/m2 and 250 nm, respectively. Independent of vesicle size and loading with dye molecules, the adsorption of intact vesicles was observed at all concentrations below a threshold concentration, above which the formation of smooth lipid bilayers occurred. In conjunction with previous work (Johnson, J. M.; Ha, T.; Chu, S.; Boxer, S. G. Biophys. J. 2002, 83, 3371-3379), these data show that 6% TR 20 mM CF vesicles adsorb to the surface intact but undergo partial rupture in which they exchange content with the external buffer.

Quantification of nano-scale intermembrane contact areas by using fluorescence resonance energy transfer

“…2B Inset and reveals that small vesicles on average deform only slightly, whereas larger vesicles adopt a more hemispherical shape on the substrate. Severe deformations of vesicles have previously been observed for SUVs immobilized on unprotected glass surfaces having high adhesion potential (29). The equilibrium shape of vesicles on adhesive substrates is determined by the gain in adhesion energy W A at the cost of bending and stretching the bilayer.…”

Formation of an intact liposome layer adsorbed on oxidized gold confirmed by three complementary techniques: QCM‐D, AFM and confocal fluorescence microscopy

“…[9] The height of the liposomes is much smaller than the nominal diameter and also smaller than the thickness of the adsorbed layer estimated with QCM-D. In fact, according to many reports in the literature, [44,45] the height is usually underestimated because the high surface density of the liposomes prevents the tip from accessing the substrate. Furthermore, the liposomes appear to be located in different planes, which may be caused by rearrangements of the liposomes during imaging, causing a reduction in image resolution.…”

Coarse-grained modeling of interactions of lipid bilayers with supports

“…7,8 Even interactions with whole cells have been studied. 9,10 Characterization of supported lipid bilayers has been accomplished with numerous methods including atomic force microscopy, 11 x-ray and neutron scattering and reflectometry, [12][13][14][15] nuclear magnetic resonance, 16 quartz crystal microbalance, 17 surface plasmon resonance, [18][19][20][21] ellipsometry, 6 electrical impedance spectroscopy, 15 and many versions of fluorescence microscopy such as fluorescence resonance energy transfer, 22 fluorescence recovery after photobleaching, 23 fluorescence interference contrast, 24,25 fluorescence correlation spectroscopy, 26 and total internal reflection fluorescence. 27 Phospholipid bilayers adsorb to hydrophilic surfaces to create a supported bilayer leaving a layer of water up to 5 Å thick-in addition to the hydration shell of the head groups-between the substrate and the head groups of the lipids in the proximal ͑closest to the solid͒ leaflet.…”