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Title: Development of liquid crystal-based system for biomolecule and nanomaterial characterization
Keywords: liquid crystals, phospholipids, phospholipases, gold nanoparticles, cell membranes, protein sensor
Issue Date: 3-Aug-2009
Citation: DENY HARTONO (2009-08-03). Development of liquid crystal-based system for biomolecule and nanomaterial characterization. ScholarBank@NUS Repository.
Abstract: Liquid crystal (LC)-based system is a promising platform for sensing due to the unique properties of LCs. It can potentially be used for real-time and label-free detection with high sensitivity and without complex instrumentation. The research work described in this thesis explores the use of thermotropic LCs for probing and imaging molecular-scale interactions occur at an aqueous-LC interface. The research exploration is organized into two categories.

The first category focuses on the biomolecule sensing. A novel air-supported LC-based system that permits real-time and label-free interfacial examination with high-throughput speed and small sample quantity was first designed and developed. Using this system, the enzymatic hydrolysis of phospholipid monolayer self-assembled at aqueous-LC interface by various phospholipases (PLA2, PLC, PLD) were characterized. During these enzymatic events, orientational transitions of LCs were triggered and the corresponding optical signals reflecting the spatial and temporal distribution of phospholipids were generated. The mechanisms of phospholipase-induced LC orientational changes were also investigated. Finally, introducing phospholipase inhibitors together with the respective phospholipases inhibited the enzymatic activities and resulted in no measurable optical response of LCs.

The air-supported LC system was next used to identify phospholipase-like toxins. Beta-bungarotoxin exhibits Ca2+-dependent phospholipase A2 activity whereas alpha-bungarotoxin and myotoxin II do not exhibit any phospholipase activity. The LC sensor selectively identified beta-bungarotoxin when it hydrolyzed a phospholipid monolayer self-assembled at aqueous-LC interface and triggered orientational responses of LCs. The sensor was also very sensitive and required less than 5 pg of beta-bungarotoxin for the detection. When phospholipase A2 inhibitors were introduced together with beta-bungarotoxin, no orientational response of LCs could be observed. In addition, the regeneration of the sensor could be done without affecting the sensing performance.

After demonstrating the feasibility of studying enzymatic activities, we further employed the air-supported LC-based system to self-assemble nitrilotriacetic acid-terminated amphiphiles loaded with Ni2+ ions at the aqueous-LC interface. This LC surface was capable for immobilizing histidine-tagged proteins in a well-defined orientation via complex formation between Ni2+ and histidine. Using histidine-tagged ubiquitin as a model protein to decorate LC surface, orientational transitions of LCs was observed by exposing the surface to antibody target to induce specific protein-protein binding events. The resultant sharp LC optical switching from dark to bright can readily be observed under polarized lighting. This work demonstrates that the air-supported LC system provides a facile platform for biomolecule characterization including for studying enzymatic reaction and inhibition, toxin identification inhibitor screening as well as specific protein-protein binding events.

The second category focuses on the nanomaterial characterization. Protein-coated gold nanoparticles were found to disrupt cell membrane model system consisting of either phospholipid or mixed phospholipid/cholesterol monolayers self-assembled at aqueous-LC interface. The monolayer disruption was found to depend strongly on the type of protein (albumin, neutravidin and fibrinogen) adsorbing onto nanoparticle surfaces. Hydrophobic interaction was found to play a major role in the disruption. Furthermore, mixed phospholipid/cholesterol monolayers with higher cholesterol contents were more susceptible to the disruption by protein-coated AuNPs. Results obtained from this study may offer new understanding in the potential nanotoxicity pathway, where the biophysical interaction between nanomaterials and cell membrane is an important step.
Appears in Collections:Ph.D Theses (Open)

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