A whole cell BIAcore assay to evaluate P1-mediated adherence of Streptococcus mutans to human salivary agglutinin and inhibition by specific antibodies.
Researchers now recognize the utility of surface plasmon resonance technology to evaluate interactions of microbial pathogens with host components. The surface adhesin and candidate vaccine antigen P1 of Streptococcus mutans, the main causative agent of dental caries, interacts with a high molecular weight glycoprotein called salivary agglutinin, or gp340, in the salivary pellicle. We optimized a BIAcore assay to measure P1-mediated Ca(2+) dependent binding of S. mutans whole cells to this physiological ligand immobilized on a Pioneer F1 sensor chip. Regeneration conditions allowed cells to be eluted from the sensor chip permitting multiple reuse of the agglutinin-coated surface. An isogenic P1-deficient S. mutans mutant did not bind to immobilized agglutinin demonstrating specificity of the detected interaction. Glutaraldehyde-fixation of bacterial cells showed the assay measured a whole cell-ligand interaction and was not an artifact of solubilized or leached proteins. Adherence inhibition assays demonstrated varying degrees of disruption of the S. mutans-agglutinin interaction by anti-P1 monoclonal antibodies recognizing different epitopes, whereas a polyclonal reagent demonstrated more complete inhibition. This report describes an improved method to assess salivary agglutinin-mediated adherence of S. mutans in vitro under physiological-like conditions and to evaluate the effectiveness of antibodies of differing specificities to inhibit binding.
Oli
Monika W
MW
Department of Oral Biology, Health Science Center, University of Florida, Gainesville, 32610-0424, USA.
McArthur
William P
WP
Brady
L Jeannine
LJ
D000373:Agglutinins / Q000378:metabolism*; D000907:Antibodies, Bacterial / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000276:immunology / Q000378:metabolism*; D015374:Biosensing Techniques / Q000379:methods; D006801:Humans; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D012463:Saliva / Q000737:chemistry / Q000378:metabolism*; D013295:Streptococcus mutans / Q000166:cytology* / Q000502:physiology
D000373:Agglutinins; D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D008562:Membrane Glycoproteins; C055764:S-layer proteins