Dissertation
Measuring Membrane Mobility: A Comparison of Three Techniques Including Instrument and Method Development
Doctor of Philosophy (PhD), Washington State University
01/2014
Handle:
https://hdl.handle.net/2376/116850
Abstract
Supported lipid membranes have and continue to be of great interest in building surrogate environments for studying biologically important systems in vitro. These biomimetic assemblies provide researchers with the ability to isolate and study a system of interest in a controllable and customizable environment. However, many of the properties of biomimetic assemblies are highly susceptible to variation due to thermal fluctuations, in particular the phase of the lipid. In this dissertation the mobility of four different membranes was studied as a function of temperature by measuring the diffusion coefficient. Three of the lipid membrane systems were homogeneous single component systems and the fourth was a complex mixture of three different lipids commonly used in studies involving cytochrome P450.
A major component of this dissertation was the design and construction of three different optical systems. Measurement of membrane mobility was studied using all three techniques. The three microscopes were a single molecule microscope in total internal reflectance mode for single particle tracking, a confocal fluorescence correlation spectroscopy microscope, and a fluorescence recovery after photobleaching microscope. Additionally, a thermal control system was designed and built to regulate the temperature of both the sample and objective for all three microscopes with a thermal accuracy of ±0.1°C. The system was designed so that both the sample and the objective could be either heated or cooled to promote system stability and had a useful range of -10 - 47°C for these experiments.
Observations of diffusion coefficients and general trends in mobility are consistent with literature values for the homogeneous systems, however, the ternary lipid blend had a high degree of variability. This was due to the formation of domains, or lipid rafts, which have varying mobility from one domain to another based on the characteristics of the lipids, the conditions used to prepare the liposomes for the bilayer, and the sample temperature.
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Details
- Title
- Measuring Membrane Mobility: A Comparison of Three Techniques Including Instrument and Method Development
- Creators
- Adam O'Neill Barden
- Contributors
- James A Brozik (Advisor)Jeffrey P Jones (Committee Member)Alexander D Li (Committee Member)ChulHee Kang (Committee Member)
- Awarding Institution
- Washington State University
- Academic Unit
- Chemistry, Department of
- Theses and Dissertations
- Doctor of Philosophy (PhD), Washington State University
- Number of pages
- 169
- Identifiers
- 99900581844401842
- Language
- English
- Resource Type
- Dissertation