Electroformation of Hydrogel –stamped Protein and Lipid Deposits for the Creation of Giant Vesicle Arrays
Open Access
- Author:
- Gaddes, Erin Richards
- Graduate Program:
- Bioengineering
- Degree:
- Master of Science
- Document Type:
- Master Thesis
- Date of Defense:
- April 09, 2013
- Committee Members:
- Sheereen Majdzarringhalamaraghy, Thesis Advisor/Co-Advisor
- Keywords:
- hydrogel stamping
giant vesicles
proteoliposomes
protein-lipid interactions
electroformation - Abstract:
- The cellular membrane is a vital barrier between the cell and its extracellular environment, providing mechanical support, allowing growth and movement, regulating transport, and organizing communication. However, ascribing specific biological functions to particular membrane compositions remains difficult because of the difficulty of precisely controlling composition or measuring function in cells. Giant vesicles made of lipids and proteins offer a simplified and controllable platform for simulating natural cell membranes and can be used to study biological processes such as vesicle budding and protein-lipid interactions. The formation of giant vesicles arrays, in turn, can permit high-throughput investigation of these membrane processes. In this thesis, a simple technique to form arrays of giant vesicles from hydrogel-stamped lipid and protein deposits is described. First, topographically-patterned agarose stamps were applied that could absorb and transfer aqueous solutions of lipids, proteins, and cell membrane fragments to an indium tin oxide substrate. Next, these patterned deposits were used to produce giant vesicles through electroformation. Through the variation of stamp feature size, the size and number of vesicles formed at each stamped location could be prescribed. The versatility of this technique was demonstrated by forming vesicle arrays from stamped proteoliposome solution, stamped membrane fragment, and in physiologically relevant solutions. Furthermore, protein-lipid interactions were studied by observing the binding of fluorescently-labeled proteins to phosphatidylserine in vesicles. Finally, the future outlook of this technique is discussed and potential future studies that take advantage of its simplicity are suggested.