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Title: Biopanning for Salmonella antigens
Author: Khalili, Shirin Fatima
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2004
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Of the two currently available typhoid fever vaccines, one is potentially inappropriate for immuno-compromised individuals while the other could be rendered ineffective as the pathogen evolves. This work addresses these shortcomings and explores the possibility of identifying novel antigens for use in a multiple subunit vaccine. S. enterica serovar Typhimurium infection of mice is a well-established animal model of human typhoid fever. Phage display is a method of generating particles that embody a physical link between a given DNA sequence and the polypeptide it encodes. In this work, a phage display library was constructed from random fragments of Salmonella genomic DNA. Hyper-immune serum from infected mice was used for affinity selection of potential antigens from this library in a process called biopanning. Individual clones encoding potential antigens were subjected to further screening alongside a positive and negative control, and 10 were consistently positive. Sequences encoding the potential antigens were then sub-cloned into an expression vector encoding a His tag for affinity purification.  Six of the ten sub-clones could not be induced to express the fusion peptide. Poor induction of the remaining four led to multiple purification steps. The four potential antigens were then tested by Western blot and ELISA alongside the same positive control for antigenicity. It was then found that the positive control in itself was poorly antigenic and the four candidates were not antigens. Possibilities for modification of the original affinity selection process, as well as phage-displayed positive control antigens are discussed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available