title: Characterization of the physiological requirements for the bactericidal effects of a monoclonal antibody to OspB of Borrelia burgdorferi by confocal microscopy.
abstract: A confocal microscopy study was undertaken to characterize the bactericidal effects of the Fab fragments of CB2, an immunoglobulin G1kappa murine monoclonal antibody, to an epitope in the carboxy region of the outer surface protein B (OspB) of Borrelia burgdorferi. Simultaneous direct labeling of both fixed and live spirochetes with fluorochrome-labeled Fab-CB2 and 11G1, and an immunoglobulin Mkappa monoclonal antibody to OspA, showed that OspA and OspB seem to colocalize in dead spirochetes but do not appear to be physically associated when the organisms are alive. A polar bleb composed of a Fab-CB2-OspB complex, followed by incorporation of 11G1-OspA, precedes the formation of a spheroplast. The spheroplasts contain both OspA and OspB and are a terminal stage in the bactericidal process induced by Fab-CB2. Outer membrane destabilization by Fab-CB2, but not cell wall or cytoplasmic membrane alterations, was demonstrated experimentally by the sequential treatment of spirochetes with Fab-CB2 and monoclonal antibodies to flagellin and DnaK. The action of Fab-CB2 is epitope specific, as another monoclonal antibody to an epitope in the amino terminus of OspB was not bactericidal. The bactericidal effect of Fab-CB2 is not dependent on the induction of spirochetal proteases but is dependent on the presence of Ca2+ and Mg2+. Supplementation of Ca2(+)- and Mg2(+)-free medium with these cations restored the bactericidal effects of Fab-CB2. The mechanism by which a Fab fragment of an antibody destroys a bacterium directly may represent a novel form of antibody-organism interaction.
authors.affiliation: Department of Pathology, State University of New York, Stony Brook 11794, USA.
authors.forename: R
authors.initials: R
authors.lastname: Escudero
authors.forename: M L
authors.initials: ML
authors.lastname: Halluska
authors.forename: P B
authors.initials: PB
authors.lastname: Backenson
authors.forename: J L
authors.initials: JL
authors.lastname: Coleman
authors.forename: J L
authors.initials: JL
authors.lastname: Benach
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology; D000942:Antigens, Bacterial; D000954:Antigens, Surface / Q000276:immunology* / Q000378:metabolism / Q000502:physiology*; D001425:Bacterial Outer Membrane Proteins / Q000276:immunology* / Q000378:metabolism / Q000502:physiology*; D001428:Bacterial Vaccines; D015748:Borrelia burgdorferi Group / Q000276:immunology* / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D002473:Cell Wall / Q000378:metabolism / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000096:biosynthesis; D004790:Enzyme Induction; D000939:Epitopes / Q000276:immunology* / Q000378:metabolism / Q000502:physiology*; D029968:Escherichia coli Proteins; D005408:Flagellin / Q000276:immunology; D019085:Fluorescent Antibody Technique, Direct; D019084:Fluorescent Antibody Technique, Indirect; D018840:HSP70 Heat-Shock Proteins / Q000276:immunology; D007140:Immunoglobulin Fab Fragments / Q000276:immunology* / Q000502:physiology*; D008074:Lipoproteins; D008274:Magnesium / Q000378:metabolism; D018613:Microscopy, Confocal
chemical_list: D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; D000939:Epitopes; D029968:Escherichia coli Proteins; D018840:HSP70 Heat-Shock Proteins; D007140:Immunoglobulin Fab Fragments; D008074:Lipoproteins; C072045:OspA protein; D005408:Flagellin; C077030:OspB protein, Borrelia burgdorferi; D010450:Endopeptidases; C063842:dnaK protein, E coli; D008274:Magnesium; D002118:Calcium