Antibody response in salmonids against the 70 kDa serine protease of Aeromonas salmonicida studied by a monoclonal antibody-based ELISA.
An antigen-capture enzyme-linked immunosorbent assay (cELISA) based on monoclonal antibodies (mAb) was set up and evaluated for selective detection of salmonid antibody responses to the antigen P1, which is a weakly immunogenic exoprotease of typical Aeromonas salmonicida. This new assay permits a specific determination of anti-protease-antibodies, without antigen purification. Serum antibodies induced by the strongly immunogenic lipopolysaccharide could reliably be discriminated from anti-P1-antibodies. Antibody titres of 45 experimental antisera recorded by cELISA were moderately correlated with titres determined by routinely used indirect ELISA (iELISA) by detecting partially different antibody populations (r=0.753). Substitutions of immunoreactants and confirmatory immunoblotting strongly suggest that the mAb-based assay selectively recognises antibodies directed to epitopes of native protease. A conjugate of inhibited protease and cationized bovine serum albumin (cBSA) was found to engender a significant anti-protease-response in three salmonid species (P<0.05), whereas the unconjugated antigen and Apoject 1-Fural were proved to be ineffective. Recorded specific antibody titres were as high as 1:381,400, indicating a considerable enhanced immunogenicity of cBSA-conjugated P1 and high assay sensitivity. The established cELISA offers a promising approach to further improvement of monitoring fish humoral immune response to surface accessible epitopes of the immunosuppressive exoprotease, P1, and to scrutinize its protective significance.
Wagner
U
U
Institute for Zoology, University of Leipzig, Talstrasse 33, 04103, Leipzig, Germany. wagner.uli@t-online.de
Hädge
D
D
Gudmundsdóttir
B K
BK
Nold
K
K
Drössler
K
K
D000333:Aeromonas / Q000276:immunology*; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood*; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001426:Bacterial Proteins; D001428:Bacterial Vaccines / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D008970:Molecular Weight; D017686:Oncorhynchus mykiss; D012697:Serine Endopeptidases / Q000276:immunology*; D014337:Trout; D014611:Vaccination
D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D001428:Bacterial Vaccines; D012697:Serine Endopeptidases; C078810:aspA protein, Aeromonas salmonicida