Development of a microfluidic biosensing system for real-time monitoring of acetylcholinesterase activity
File(s)
Author(s)
Smith, Georgia
Type
Thesis
Abstract
This thesis describes the development of a microfluidic biosensing system for the real-time monitoring acetylcholinesterase (AChE) activity in blood samples following exposure to organophosphorus (OP) compounds.
Two methods are developed using choline oxidase (ChOx) to sense choline, the product of acetyl-
choline (ACh) hydrolysis, as a way of measuring AChE activity. A system where ChOx is added in flow was developed, based on the adaptation of an existing microfluidic platform comprised of LabSmith programmable components. This gave a high sensitivity of 24.68 ± 0.09 nA/mM when using optimal experimental parameters. A choline biosensor was fabricated as an alternative sensing approach, by immobilising a ChOx hydrogel layer on a combined needle microelectrode surface. The choline biosensor typically gave a sensitivity of 4 nA/mM. The choline biosensor was successfully used to monitor choline concentration in a brain microdialysate stream in a porcine model of cardiac arrest and resuscitation.
The microfluidic platform was adapted to allow addition of ACh and AChE streams to create a system where either ACh or AChE calibrations can be carried out. Using the existing microfluidic analysis system coupled with the choline biosensor, a successful AChE calibration was demonstrated. Preliminary experiments were made to utilise microdialysis with the aim of sampling in complex
media such as blood. Results demonstrate the use of microdialysis to sample different AChE activities in a stationary solution. Retrodialysis was also successfully performed to deliver ACh into AChE-
containing solutions.
A microfluidic flow cell that incorporates microdialysis probe was developed, to create a flowing system in which reactions can be tightly controlled...
Two methods are developed using choline oxidase (ChOx) to sense choline, the product of acetyl-
choline (ACh) hydrolysis, as a way of measuring AChE activity. A system where ChOx is added in flow was developed, based on the adaptation of an existing microfluidic platform comprised of LabSmith programmable components. This gave a high sensitivity of 24.68 ± 0.09 nA/mM when using optimal experimental parameters. A choline biosensor was fabricated as an alternative sensing approach, by immobilising a ChOx hydrogel layer on a combined needle microelectrode surface. The choline biosensor typically gave a sensitivity of 4 nA/mM. The choline biosensor was successfully used to monitor choline concentration in a brain microdialysate stream in a porcine model of cardiac arrest and resuscitation.
The microfluidic platform was adapted to allow addition of ACh and AChE streams to create a system where either ACh or AChE calibrations can be carried out. Using the existing microfluidic analysis system coupled with the choline biosensor, a successful AChE calibration was demonstrated. Preliminary experiments were made to utilise microdialysis with the aim of sampling in complex
media such as blood. Results demonstrate the use of microdialysis to sample different AChE activities in a stationary solution. Retrodialysis was also successfully performed to deliver ACh into AChE-
containing solutions.
A microfluidic flow cell that incorporates microdialysis probe was developed, to create a flowing system in which reactions can be tightly controlled...
Version
Open Access
Date Issued
2023-04-12
Date Awarded
01/10/2023
License URL
Advisor
Boutelle, Martyn
Goodchild, Sarah
Sponsor
Engineering and Physical Sciences Research Council
Defence Science and Technology Laboratory (Great Britain)
Publisher Department
Bioengineering
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)