| Description |
The importance of lipid bilayers to the structure and function of cellular membranes coupled with their inherent complexity has driven the development of analytical techniques capable of high-throughput investigation of these surfaces. This work describes a continuous flow microspotter (CFM) that was modified to create micropatterned lipid bilayer arrays (MLBAs). This dissertation is divided into four main parts, with the first chapter focusing on the characterization of MLBAs using fluorescence microscopy to ensure bilayer formation and integrity. The individually addressable nature of the CFM was also demonstrated using a multi-ligand array containing ganglioside GM1, dinitrophenyl (DNP) and biotin. A multiple protein-ligand assay was performed using the ligand array to detect three different fluorescently labeled proteins (cholera toxin b (CTb), anti-DNP antibody and NeutrAvidin) from solution simultaneously. The second part of this dissertation concentrates on creating stable MLBAs using a polymerizable lipid, poly(bis-SorbPC) in order to generate a more robust biosensing platform. The poly(lipid) arrays were compared directly to the MLBAs prepared without the polymerizable lipids using fluorescence microscopy to demonstrate their superior stability. A multiple protein-ligand assay was also performed to demonstrate the utility of these arrays and their potential application as a sensor substrate. iv Next, the MLBAs were used to investigate the impact of fifteen different lipid components on small molecule-membrane binding. The lipophilic dye merocyanine 540 (MC540) was used as a model small molecule and its binding was monitored by fluorescence microscopy. These studies demonstrate the potential of using MLBAs to investigate drug membrane interactions while preserving time and cost-effectiveness. Finally, sum-frequency vibrational imaging (SFVI) was developed to provide a surface specific noninvasive, analytical technique capable of monitoring lipid structure and dynamics in a high-throughput manner. The vibrational sensitivity of SFVI was investigated with an asymmetric lipid bilayer patterned by ultraviolet (UV) radiation lithographically. The phase behavior of three different binary mixtures in a MLBA was successfully investigated using SFVI. The SFVI setup had the sensitivity, resolution and field of view required for exploring lipid bilayer properties in an array format. This dissertation presents a new approach for assembling lipid bilayer arrays in combination with a powerful analytical technique to allow exploration of the physical properties of lipid membranes in a high-throughput and noninvasive manner. |
