Zhang, Xin (2017) Application of centrifugal microfluidics and fluorescence-based detection for rapid biological analysis. PhD thesis, Dublin City University.
Abstract
The goal of this research was to develop and optimise microfluidic systems and assays
for rapid detection of selected targets such as marine toxins and human IgG. The
measurement of IgG, as the major format of therapeutic antibodies, was used as a
prototype to develop a centrifugal-based microfluidic system for effectively monitoring
biopharmaceutical production. To achieve this, a sandwich immunoassay for human
IgG detection was developed and used to study the implementation of a new
microfluidic CD-based cartridge. The centrifugal-based microfluidic CD adopted a
serial siphon technique for implementation of automated sequential delivery of the
assay reagents. Surface-confined supercritical angel fluorescence (SAF)-based detection
was designed to sensitively measure the fluorescence signal from the microfluidic CDbased immunoassay. The CD substrate was functionalized with
aminopropyltriethoxysilane (APTES) using plasma enhanced chemical vapour
deposition (PECVD) for the immobilization of analyte capture protein. The developed
prototype microfluidic system could automatically run a microfluidic assay in less than
30 min, and accurately measure industrial bioprocess samples that contained 10 mg mL1 of human IgG. Additionally, computational simulations were performed to
fundamentally understand the kinetics of immunoassays in a microfluidic system. The
effects of varying assay parameters on the capture of analytes in microfluidic-based
heterogeneous immunoassays under real-world operating conditions, was examined by
using theoretical modeling and experimental binding assay results. A marine toxin,
saxitoxins (STX), was another potential target for analysis using the developed
centrifugal-based microfluidic CD. Work was carried out to generate recombinant
antibodies (scFv) to saxitoxin and its derivatives. After successful immunization of the
host animal with different STX-conjugates, an scFv antibody phage library against antiSTX was constructed by PCR amplification of scFv genes, cloning the genes library
into phagemid vector and transforming into E. coli cells. The phages expressing high
affinity antibody gene were isolated by “bio-panning”.
Metadata
Item Type: | Thesis (PhD) |
---|---|
Date of Award: | November 2017 |
Refereed: | No |
Supervisor(s): | O'Kennedy, Richard and Murphy, Caroline |
Subjects: | Biological Sciences > Microfluidics |
DCU Faculties and Centres: | DCU Faculties and Schools > Faculty of Science and Health > School of Biotechnology |
Use License: | This item is licensed under a Creative Commons Attribution-NonCommercial-No Derivative Works 3.0 License. View License |
ID Code: | 21904 |
Deposited On: | 17 Nov 2017 14:16 by Richard O'Kennedy . Last Modified 28 Jul 2021 16:30 |
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