On the Formation of Supported Phospholipid Bilayers

Doktorsavhandling, 2004

Lipid bilayers are a central component of both the structure and function of biological membranes, like the one surrounding cells of living organisms. Since the lipid bilayer provides a host matrix for membrane proteins, which perform many of the specialized tasks in cells like energy, signal and material transduction, researchers have tried to develop artificial model membranes to study a variety of membrane protein functions. One of the most promising model systems during the last two decades have been the supported lipid bilayer, which is compatible with plenty of surface analytical tools and has many potential applications, including drug screening, medical diagnostics and non-fouling surfaces. Supported phospholipid bilayers (SPBs) are increasingly prepared through lipid vesicle fusion with surfaces. The process by which a planar membrane is formed from a spherical vesicle is still not fully understood. It provides an interesting case of molecular self-assembly, from a physics point of view, in addition to being important for preparing high quality membrane mimics for applications. In this work vesicle adsorption to several hydrophilic surfaces has been studied. However, most of the work is focused on phosphatidylcholine lipid vesicle adsorption and subsequent SPB formation on SiO2. In the course of determining the detailed kinetics of the vesicle-to-SPB transformation, new instrumentation and methods for data interpretations were developed, which are also presented in this thesis. A combination of quartz crystal microbalance with dissipation monitoring (QCM-D) and surface plasmon resonance (SPR) measurements was used to monitor the time-evolution of the vesicle and SPB abundance on the surface. These results were compared to atomic force microscope (AFM) measurements to get the microscopic distribution. It was possible to calculate the critical coverage of vesicles on the surface, required to trigger rupture. A bulk vesicle size and concentration dependent desorption of lipids to complete the SPB was also quantified. Single-technique, QCM-D, measurements of the influence of surface chemistry, osmotic pressure, lipid concentration, vesicle size and temperature were also performed. It was possible to find factors facilitating vesicle rupture and high-quality SPB formation (e.g. increased temperature and hypotonic vesicles), as well as factors inhibiting vesicle rupture and causing defects in the final SPB (e.g. low temperature and high concentrations). Combined with the QCM-D/SPR results, these measurements also made it possible to refine our understanding of the rupture mechanism. The presented results strongly indicate that SPB-formation is dominated by SPB-edge induced rupture of neighboring vesicles, where the interaction of vesicles in the solution with the edges seems to play a crucial role. Monte Carlo computer simulations were performed to find a mechanistic picture that could explain the observed temperature dependence of the SPB formation. A good correspondence between theoretical and experimental results was found, both qualitatively and quantitatively. Furthermore, in an experiment, where the outer monolayer of the vesicles was selectively tagged, it was shown that the opening and adsorption of the vesicle membrane follows a pathway leaving the outer monolayer primarily facing the substrate after SPB formation.