|Kerr, Paul - THE QUEEN'S UNIV.|
|Traynor, Imelda - VETERINARY SCIENCES DEPT.|
|Elliott, Christopher - VETERINARY SCIENCES DEPT|
Submitted to: Association Official Analytical Chemists Annual Intrl Meeting & Exposition
Publication Type: Abstract Only
Publication Acceptance Date: July 2, 2004
Publication Date: September 22, 2004
Citation: Brandon, D.L., Mandrell, R.E., Kerr, P.G., Traynor, I.M., Elliott, C.T. 2004. Detection of e. coli O157 by surface plasmon resonance [abstract. 118th meeting of AOAC International, St. Louis, MO. 19-23 Sept. 2004. Abstract No. 1518. Technical Abstract: The goal of this work is to develop a protocol for characterizing the ability of monoclonal antibodies to serve as 'capture' antibodies in techniques such as ELISA, surface plasmon resonance (SPR), and immunomagnetic separations. Antibodies were developed against E. coli O157, using lipopolysaccharide (LPS)-enriched fractions. Outer membrane complexes or a fimbrial-LPS mixture (Kerr et al., J. Appl. Microbiol. 90:543-549, 2001) were used as immunogens to obtain monoclonal antibodies derived from BALB/c mice. Direct assay by SPR used antibody immobilized on a dextran-gold chip (Biacore AB, Uppsala, Sweden) to 'capture' bacteria from a suspension. E. coli O157 (Strain S784, an enterohemorrhagic strain of bovine origin) were heat-killed and adjusted to a density of 1 x 107 cells/mL. The suspension was diluted 1:1 in buffered saline. Injections were performed at a flow rate of 5 mL/min for 5 min (i.e., 1.25 x 105 cells), using a Biacore 2000 optical biosensor. The limit of detection (LOD) was estimated as 4 x 104 cells/mL or 103 cells/injected sample. The LOD was similar when the analysis was done as a homologous sandwich of MAb 7A6. This methodology provided sensitivity similar to sandwich ELISA using MAb 7A6, which has a LOD of 1 x 105 E. coli (Kerr et al.). Even without potential enhancements achievable by sandwich labeling techniques, SPR could provide the basis for a one-step assay for E. coli O157. Although all the antibodies were specific for O157, not all antibodies functioned as effective capture reagents in SPR. The basis for these differences could reside in the isotype, specificity, or binding kinetics of the antibodies and is currently under study.