Abstract
Under physiological environments, the dynamically physical and chemical process within the cell membrane surfaces serves as indispensable parts of biology signaling pathways. Supported lipid bilayer (SLB), demonstrated as a model of cell membranes, has been widely expanded to prospective applications in bioanalytical chemistry or biotechnology. In addition, combination of SLB and nanoparticles provides a novel design for developing highly applicable functional biomaterials. In the first section of this dissertation, the formation of SLB and their potential biofunctionality against protein adsorption were investigated by dual polarization interferometry (DPI) and capillary electrophoresis (CE). DPI studies on different formulations of double-chained, zwitterionic phospholipidlipids, allow the process of bilayer formation to be followed in situ and in real time. Furthermore the anti-protein adsorption effect provided by the various formulated SLBs was examined by DPI. In addition, the SLB coatings of the same lipid formulations were subsequently employed in CE experiments as a pseudo-stationary phase for demonstrating more efficient separation of alkaline protein standard mixtures. SLB-assisted CE was found to be capable of separating 4 alkaline proteins. This study demonstrates the applicability of DPI to monitor the process of SLB formation; and our findings, obtained by both DPI and CE, confirm that the presence of the SLB reduced drastically the problematic interactions between cationic, alkaline proteins and the negatively charged silica capillary wall, leading to better recovery and efficient separation of the proteins under investigation. In the second section, we prepared the biomimetic silica microspheres, lipid raft-presenting silica microspheres, from binary phase of lipid mixtures. The formation of lipid raft on resulted silica spheres led to produce nearly 3-time affinity capacity toward lysozyme than non-raft presenting lipid structure, where charged lipids distribute homogeneously on the membrane. The desorption of adsorbed lysozyme was simply achieved by modulating temperature; it was also found in this study that the catalytical activity of adsorbed lysozyme was significantly preserved. Furthermore, DPI was employed to investigate the affinity of lysozyme toward lipid rafts fabricated on the surface of silica chip. Based on the real-time sensorgram acquired by DPI, it ws revealed that (i) lipid raft was capable of enhancing the affinity toward lysozyme; (ii) the cooperative formation of lipid raft was observed. The biomimetic silica microspheres hold the potential of being integrated with other biotechnological tools, for purifying membrane protein or for exploring the mechanism of lipid rafting. At last, a facile approach in synthesizing mono-dispersed, lipid-capped gold nanoparticles was demonstrated. Such the nanomaterial, fully coated with lipid bilayer and capable of resisting aggregation in high-salt and detergent-containing solutions, was able to be engaged in the development of a novel sensor for monitoring ligand-receptor interaction on the lipid membrane. Furthermore, such sensor offers the feasibility in studying the interaction kinetics between glycolipid and lectin on lipid membrane as well. To prove of the concept, GM1/PNA pair was chosenas model example, and the lipid-capped gold nanoparticle allowed evaluation of apparent dissociation constant (KD). Results show that 3-D configuration of the gold nanoparticles enhanced the multivalent binding effect of glycolipid GM1 to PNA futher comparing to that of the planner format. The addition of cholesterol led to the enhanced affinity of GM1 toward PNA by several folds, which should be attributed to lipid raft formation. The results obtained by atomic force microscopy (AFM) also confirmed the positive correlation between the cholesterol concentration and the size of lipid raft.