Characterization of the F41 fimbrial antigen of enterotoxigenic Escherichia coli by using monoclonal antibodies.
We produced monoclonal antibodies (MAbs) from 23 different murine hybridoma cell lines against the F41 fimbrial antigen of bovine and porcine enterotoxigenic Escherichia coli. Cell lines were created by fusing myeloma cells and spleen cells of mice that were immunized with either purified F41 or with Formalin-killed whole cells. The specificity of the MAbs to the F41 antigen was proven by enzyme-linked immunosorbent assays (ELISAs) and radioimmunoprecipitation tests. Epitope analysis with a competition ELISA revealed that the 23 MAbs recognized at least five epitopes. These results were corroborated by those of immunodiffusion tests, in which all possible combinations of two MAbs were tested against ultrasonically disintegrated F41 antigen. In a double-antibody sandwich ELISA, all peroxidase-conjugated MAbs bound to the F41 antigen of all 182 bacterial strains that were tested. Apparently, the epitopes recognized by the MAbs are highly conserved. Immunoelectron microscopy revealed that the MAbs were directed to fimbrial structures 3 to 4 nm in diameter and that the epitopes were equally distributed along the fimbriae. Consequences for the replacement of polyclonal antisera by MAbs in diagnostic tests are discussed. The results of the radioimmunoprecipitation assay suggested that F41 fimbriae are composed of a single repeating 29,000-dalton protein subunit; however, we could not exclude the possibility of the existence of minor fimbrial components.
van Zijderveld
F G
FG
Department of Bacteriology, Central Veterinary Institute, Lelystad, The Netherlands.
Westenbrink
F
F
Anakotta
J
J
Brouwers
R A
RA
van Zijderveld
A M
AM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D000918:Antibody Specificity; D000942:Antigens, Bacterial / Q000032:analysis* / Q000276:immunology; D001667:Binding, Competitive; D002460:Cell Line; D004768:Enterotoxins; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis; D004926:Escherichia coli / Q000032:analysis* / Q000276:immunology / Q000648:ultrastructure; D005260:Female; D036781:Fimbriae Proteins; D010861:Fimbriae, Bacterial / Q000276:immunology*; D006825:Hybridomas / Q000378:metabolism; D007122:Immunoelectrophoresis; D007132:Immunoglobulin Isotypes / Q000032:analysis; D051379:Mice; D008807:Mice, Inbred BALB C; D011233:Precipitin Tests
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D004768:Enterotoxins; D000939:Epitopes; D007132:Immunoglobulin Isotypes; C022924:colonization factor antigens; D036781:Fimbriae Proteins