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Title: Cloning and expression of antibody fragments for detection of Listeria monocytogenes in food
Author: Dubbels, Anne M.
ISNI:       0000 0001 3433 3709
Awarding Body: University of Aberdeen
Current Institution: University of Aberdeen
Date of Award: 1996
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Single chain Fv (scFv) and Fv antibody fragments derived from a monoclonal antibody recognising flagellae of Listeria species were cloned, expressed and purified from Escherichia coli. ScFv and Fv were purified by affinity chromatography on anti-flagellar and anti-hydrophil 2 affinity columns respectively. These fragments successfully compete with the parent monoclonal antibody for antigen binding but do not complete with a second monoclonal antibody which recognises a different epitope on the flagellae. The amount of antibody fragments expressed has been estimated as 0.1 mg 1-1 for scFc and 0.22 mg 1-1 for Fv. The relative affinities of the scFv, Fv and parental monoclonal antibody for binding to antigen were compared. Characterisation of ScFv by antigen binding profile showed it to have a sensitivity of the same order of magnitude as the parent monoclonal. Fv had a sensitivity one order of magnitude below that of the parent monoclonal and the scFv. ScFv was purified as a dimer and analysed by HPLC size exclusion chromatography. The variable heavy region (VH) sequence has been determined for a second anti-flagellar monoclonal antibody and this has been cloned into an E. coli expression vector. An IgM monoclonal antibody with broad range specificity to bacteria has been generated and the VH sequence determined.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Food technology & food microbiology