Cassedy, Arabelle (2021) Tailored immunoassay and molecular methods for viral and fungal pathogen detection. PhD thesis, Dublin City University.
Abstract
The work described in this thesis involves the development of assays for the detection of
two crop pathogens, potato virus Y (PVY) and Rhynchosporium commune (R. commune).
Infection of crops by these pathogens results in significant economic losses and reduced
yields. Therefore, efficient control of their spread is paramount. It was proposed in this work
to develop immunoassays and molecular techniques for PVY and R. commune to provide
novel additional detection and control capabilities for these important pathogens.
An antibody library was generated in order to isolate PVY-specific scFv. The library was
screened using a combination of ELISA, phage display, and biopanning approaches. From
this screening, scFv were isolated and their PVY-binding abilities characterised in ELISA.
Anti-PVY mAbs were structurally engineered into recombinant scAb and scFv fragments.
The performance characteristics of the parental antibodies and recombinant derivatives were
determined through a combination of immunoblotting, ELISA and SPR. Subsequently, dot
blots and SPR-based immunoassays were developed for the enhanced detection of PVY.
An isothermal nucleic acid amplification-based immunoassay for the detection of PVYrelated
nucleic acids was also generated. This method combines recombinase polymerase
amplification with lateral flow immunoassay technology. The developed assay could detect
PVY nucleic acids at high sensitivity from both DNA and RNA templates.
A recombinant scFv library was generated for the isolation of antibodies with R. commune
specificity. Additionally, anti-R. commune avian pAbs were generated and characterised.
This work led to the development of paper-based immunoassays for the detection of R.
commune-associated antigens.
Both the immunoassay and molecular diagnostic approaches offer a means of early detection
of the target pathogens such that their burden in the field can be ascertained and tailored
treatment can be implemented at early stages of disease outbreaks.
Metadata
Item Type: | Thesis (PhD) |
---|---|
Date of Award: | November 2021 |
Refereed: | No |
Supervisor(s): | O'Kennedy, Richard, Parle-McDermott, Anne and Mullins, Ewen |
Uncontrolled Keywords: | Pathogen Detection |
Subjects: | Biological Sciences > Biosensors |
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 |
Funders: | Department of Agriculture, Food and Marine |
ID Code: | 26222 |
Deposited On: | 29 Oct 2021 09:33 by Anne Parle-Mcdermott . Last Modified 01 Oct 2022 03:30 |
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