BALM: Watching the Formation of Tethered Bilayer Lipid Membranes with Submicron Lateral Resolution.

Tethered bilayer lipid membranes (tBLMs) are artificial membranes largely used for the in situ study of biological membranes and membrane-associated proteins. To date, the formation of these membranes was essentially monitored by surface averaging techniques like surface plasmon resonance (SPR) and quartz crystal microbalance with dissipation monitoring (QCM-D), which cannot provide both local and real-time information in a single approach. Here, we report an original application of backside absorbing layer microscopy (BALM), a novel white-light wide-field optical microscopy, to study tBLMs. Thanks to the combination of sensitivity and resolution, BALM not only allowed the real-time quantitative monitoring of tBLM formation but also enabled the high-resolution visualization of local fluxes and matter exchanges taking place at each step of the process. Quantitative BALM measurements of the final layer thickness, reproduced in parallel with SPR, were consistent with the achievement of a continuous lipid bilayer. This finding was confirmed by BALM imaging, which additionally revealed the heterogeneity of the bilayer during its formation. While established real-time techniques, like SPR or QCM-D, view the surface as homogeneous, BALM showed the presence of surface patterns appearing in the first step of the tBLM formation process and governing subsequent matter adsorption or desorption steps. Finally, matter fluxes persisting even after rinsing at the end of the tBLM formation demonstrated the lasting presence of dispersed vesicular pockets with laterally fluctuating positions over the final single and continuous lipid bilayer. These new mechanistic insights into the tBLM formation process demonstrate the great potential of BALM in the study of complex biological systems.

Surface characterization and efficiency of a matrix-free and flat carboxylated gold sensor chip for surface plasmon resonance (SPR).

We report the preparation and characterization of a matrix-free carboxylated surface plasmon resonance (SPR) sensor chip with high sensing efficiency by functionalizing a bare gold thin film with a self-assembled monolayer of 16-mercaptohexadecanoic acid (SAM-MHDA chip). The self assembled monolayer surface coverage of the gold layer was carefully evaluated and the SAM was characterized by infrared reflection absorption spectroscopy, X-ray photoemission spectroscopy, atomic force microscopy, X-ray reflectivity-diffraction, and SPR experiments with bovine serum albumin. We compared the SPR signal obtained on this chip made of a dense monolayer of carboxylic acid groups with commercially available carboxylated sensor chips built on the same gold substrate, a matrix-free C1 chip, and a CM5 chip with a ~100 nm dextran hydrogel matrix (GE Healthcare). Two well-studied interaction types were tested, the binding of a biotinylated antibody (immunoglobulin G) to streptavidin and an antigen-antibody interaction. For both interactions, the well characterized densely functionalized SAM-MHDA chip gave a high signal-to-noise ratio and showed a gain in the availability of immobilized ligands for their partners injected in buffer flow. It thus compared favourably with commercially available sensor chips.

Late stage spreading of stratified liquids: theory.

Spreading of a liquid crystal droplet on a solid surface resembles the collapse of a zigguratlike structure and ends up with only two molecular terraces. A model is proposed to describe the time evolution of this late stage structure. It differs from the previous de Gennes-Cazabat theory by several aspects: (1) it avoids previous approximations and gives exact solutions of the kinetic equations; (2) it covers strongly layered liquids such as smectic liquid crystals; (3) it introduces the two-dimensional Laplace pressure as an essential motor for spreading; (4) it takes into account the liquid∕gas transition in the surface layer that was consistently reported in experiments with 8CB.

Surface enhanced ellipsometric contrast (SEEC) basic theory and lambda/4 multilayered solutions.

The fundamentals of a new high contrast technique for optical microscopy, named "Surface Enhanced Ellipsometric Contrast" (SEEC), are presented. The technique is based on the association of enhancing contrast surfaces as sample stages and microscope observation between cross polarizers. The surfaces are designed to become anti-reflecting when used in these conditions. They are defined by the simple equation r(p) + r(s) = 0 between their two Fresnel coefficients. Most often, this equation can be met by covering a solid surface with a single lambda/4 layer with a well defined refractive index. A higher flexibility is obtained with multilayer stacks. Solutions with an arbitrary number of all-dielectric lambda/4 layers are derived.

Hole nucleation in thin diblock copolymer films.

This paper reports an experimental study of a homogeneous nucleation process in the direct space, using an atomic-force microscope. Nucleated patterns are holes in a thin diblock copolymer film. The formation of nuclei from the metastable majority phase is followed either by the dissolution or by the growth of the hole. It is also preceded by the unexpected presence of a bump at the free surface. From the study of the nucleus surface topography at different stages of the evolution, we infer a description of the nucleation mechanism at a molecular level. It involves the existence of ordered metastable states characterized by the presence of compressed internal layers, the formation of pores in the center of the film, and the existence of interfacial flows across these pores. The presence of compressed internal layers was directly demonstrated using Neutron Reflectivity experiments, Our scheme is also consistent with all existing experimental data. After hole nucleation, we identify two distinct growth regimes: the first one is individual and the second one is interactive.

Quantitative studies of evanescent wave intensity profiles using optical fluorescence.

The fluorescence of uniformly distributed chromophores in an organic solution has been used to probe the energy density profile of an optical evanescent wave, generated by total internal reflection at the solid-solution interface. The results obtained in the case of an aqueous fluorescein solution in contact with a highly polished silica surface are in good quantitative agreement with the expected exponential decay of the optical energy at the interface. It also justifies the use of the newly developed evanescent wave-induced fluorescence technique to study adsorption and depletion layers of polymer in solution close to solids walls. In such experiments the fluorescence intensity is assumed to be the Laplace transform of the concentration profile of the fluorescently labeled polymer chains. The present data validate the above assumption.