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Title: CHARACTERIZATION OF A LISTERIA MONOCYTOGENES SPECIES-SPECIFIC SINGLE-CHAIN PHAGE ANTIBODY

Author
item Paoli, George
item Brewster, Jeffrey

Submitted to: American Society for Microbiology Annual Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 3/1/2004
Publication Date: 5/25/2004
Citation: Paoli, G., Brewster, J.D. 2004. Characterization of a listeria monocytogenes species-specific single-chain phage antibody. American Society for Microbiology Annual Meeting.

Interpretive Summary:

Technical Abstract: There are seven species of Listeria, of which only L.monocytogenes is a human pathogen. Outbreaks of Listeriosis due to contaminated foods underscore the need for rapid and specific detection of L.monocytogenes. The development of rapid methods for the detection of L. monocytogenes in food has been hampered by the lack of polyclonal serum or monoclonal antibodies that can specifically detect the organism at the species level. Phage display has proven a useful tool for the isolation of antibody fragments with desired specificities. Although phage display has advantages over conventional polyclonal and monoclonal antibody production, it has not been widely used for the selection of reagents for the detection of food-borne pathogens. We have selected and screened phage displayed scFv fragments to isolate a phage displayed antibody that detects several strains of L. monocytogenes, and does not cross-react with any of the other five species of Listeria. Thus, this antibody fragment shows the high degree of specificity required for accurate detection of L.monocytogenes. As determined by western blot analysis, the antibody binds to polypeptides with apparent molecular masses of 65 kD and 120 kD that are both on the surface of L. monocytogenes cells and exported out of the cells. The efficacy of this antibody for the detection of L.monocytogenes was examined by ELISA using L. monocytogenes grown under a variety of conditions, including growth media commonly used for the isolation of L. monocytogenes from food. Cells at all phases of growth synthesized the surface antigen. The antigen was present on the surface of the cells grown between 20oC and 42oC but was not present on the cell surface when cells were grown at or below15oC. Cells grown in Brain-Heart Infusion and Fraser Broth expressed the surface antigen, but the antigen was not detected on cells grown in Listeria Enrichment Broth or Modified University of Vermont Media. The identity of the polypeptide antigen is being determined in order to take a more rational approach to the development of an antibody-based method for the specific detection of L.monocytogenes.