Development of a Tethered Bilayer Lipid Membrane (tBLM) Lipase and Phospholipase Sensor

Mirissa Lankage, Upeksha Shashikala Randunu

Development of a Tethered Bilayer Lipid Membrane (tBLM) Lipase and Phospholipase Sensor

Mirissa Lankage, Upeksha Shashikala Randunu

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Publication Type:: Thesis
Issue Date:: 2023

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Field	Value	Language
dc.contributor.author	Mirissa Lankage, Upeksha Shashikala Randunu
dc.date.accessioned	2023-11-27T03:52:56Z
dc.date.available	2023-11-27T03:52:56Z
dc.date.issued	2023
dc.identifier.uri	http://hdl.handle.net/10453/173585
dc.description	University of Technology Sydney. Faculty of Science.	en_US.UTF-8
dc.description.abstract	Tethered bilayer lipid membrane (tBLMs) is the name for lipid bilayers that can be anchored to a conductive metal substrate. Used in conjunction with electrical impedance spectroscopy (EIS), they are a valuable tool for research into the properties of lipid bilayer interactions with proteins, peptides and toxins. This project sought to create a phospholipase biosensor that can detect Inflammatory bowel diseases (IBDs) using phospholipid tBLMs. IBDs are a group of chronic bowel conditions that include ulcerative colitis and Crohn's disease. Currently, the diagnosis of IBD conditions typically involves colonoscopies and biopsies. However, attempts have been made to diagnose IBD conditions by the levels of phospholipase A2 (PLA2)enzyme in serum and stool samples. A blind trial using 40 stool samples was tested on the DOPC tBLMs, and the presence of PLA2 activity was measured using EIS. The data suggest that tBLM arrays are capable of detecting PLA2 activity in fecal samples and that this has the potential to be developed into a point-of-care diagnostic for IBDs. Another part of this project was to investigate biomedical diagnostics for acute pancreatitis, which is a life-threatening inflammatory condition that causes severe abdominal pain in patients. Current tests for acute pancreatitis involve the use of enzyme-linked immunosorbent assays (ELISAs) of pancreatic lipase (PL) levels in blood samples which aren’t conducive to a rapid point-of-care diagnosis. This project also reports on developing an impedance sensor that uses a novel tethered membrane architecture that incorporates triglyceride triolein as a substrate to identify the presence of PL. The hydrolysis of triolein leads to membrane disruption that can then be detected using EIS. The creation of a triglyceride-tethered membrane and the ability to rapidly measure pancreatic lipase activity shows great potential for the use of this technology as a point-of-care acute pancreatitis diagnostic. This would have a particular benefit to many remote communities where there is no access to rapid pathology testing currently. These triglyceride-tethered membranes were further tested with multiple lipases that are produced commercially to demonstrate their potential to be used as a sensor in industrial research and other biomedical diagnostics. Finally, this project tested triolein tethered membranes using neutron reflectometry (NR) in order to determine their membrane thickness and water volume fraction. The NR data shows that triolein membranes typically form as a monolayer around the tethering molecules and are really a tethered monolayer lipid membrane (tMLM) architecture.	en_US.UTF-8
dc.format	Thesis (PhD)
dc.language.iso	en_US	en_US.UTF-8
dc.relation	https://opus.lib.uts.edu.au/bitstream/10453/173585/1/thesis.pdf
dc.rights	info:eu-repo/semantics/openAccess
dc.rights	The author owns the copyright in this thesis including all reproduction and reuse rights for the work. The work may not be altered without the permission of the copyright owner. Attribution is essential when quoting or paraphrasing from this thesis.
dc.rights	© 2023 Upeksha Shashikala Randunu Mirissa Lankage
dc.rights	au.edu.uts.lib/cph
dc.title	Development of a Tethered Bilayer Lipid Membrane (tBLM) Lipase and Phospholipase Sensor	en_US.UTF-8
dc.type	Thesis
utslib.copyright.status	open_access	*

Abstract:

Tethered bilayer lipid membrane (tBLMs) is the name for lipid bilayers that can be anchored to a conductive metal substrate. Used in conjunction with electrical impedance spectroscopy (EIS), they are a valuable tool for research into the properties of lipid bilayer interactions with proteins, peptides and toxins. This project sought to create a phospholipase biosensor that can detect Inflammatory bowel diseases (IBDs) using phospholipid tBLMs. IBDs are a group of chronic bowel conditions that include ulcerative colitis and Crohn's disease. Currently, the diagnosis of IBD conditions typically involves colonoscopies and biopsies. However, attempts have been made to diagnose IBD conditions by the levels of phospholipase A2 (PLA2)enzyme in serum and stool samples. A blind trial using 40 stool samples was tested on the DOPC tBLMs, and the presence of PLA2 activity was measured using EIS. The data suggest that tBLM arrays are capable of detecting PLA2 activity in fecal samples and that this has the potential to be developed into a point-of-care diagnostic for IBDs. Another part of this project was to investigate biomedical diagnostics for acute pancreatitis, which is a life-threatening inflammatory condition that causes severe abdominal pain in patients. Current tests for acute pancreatitis involve the use of enzyme-linked immunosorbent assays (ELISAs) of pancreatic lipase (PL) levels in blood samples which aren’t conducive to a rapid point-of-care diagnosis. This project also reports on developing an impedance sensor that uses a novel tethered membrane architecture that incorporates triglyceride triolein as a substrate to identify the presence of PL. The hydrolysis of triolein leads to membrane disruption that can then be detected using EIS. The creation of a triglyceride-tethered membrane and the ability to rapidly measure pancreatic lipase activity shows great potential for the use of this technology as a point-of-care acute pancreatitis diagnostic. This would have a particular benefit to many remote communities where there is no access to rapid pathology testing currently. These triglyceride-tethered membranes were further tested with multiple lipases that are produced commercially to demonstrate their potential to be used as a sensor in industrial research and other biomedical diagnostics. Finally, this project tested triolein tethered membranes using neutron reflectometry (NR) in order to determine their membrane thickness and water volume fraction. The NR data shows that triolein membranes typically form as a monolayer around the tethering molecules and are really a tethered monolayer lipid membrane (tMLM) architecture.

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