Enhancement by a serum factor of the immunoaccessibility of an enterobacterial porin protein domain.
Monoclonal antibody (MAb) generated against the domain Po I on the outer membrane (OM) porin (Po) protein of an Escherichia coli 055 strain showed weak binding by the OM in ELISA. Human serum and sera from different animal species enhanced the MAb binding in ELISA when the antibody was incubated together with serum or the OM was pretreated with serum. Human serum also enhanced the MAb binding when coats were prepared by using OMs from various cross-reacting bacteria. The ability of human serum to amplify the MAb binding by OMs was similar to that of lysostaphin. Amplification by serum was not observed when MAbs against three other enterobacterial OM proteins were tested. The amplifying serum factor was destroyed by heating (100 degrees C) and by mercaptoethanol. It appeared in fractions which corresponded to an apparent molecular weight of 75,000-80,000 after gel permeation, and, after ion-exchange chromatography, in fractions which contained a protein of 60 kD when analysed by SDS-PAGE. These data support the notion that the serum-induced enhancement of the anti-Po I MAb binding was due to a previously described serum amidase (N-acetylmuramyl-L-alanine amidase) which has peptidoglycan-degrading activity. The effects of the amplifying serum factor may influence the antibody levels which are measured when OMs from Gram-negative bacteria are used as antigen in a serodiagnostic test.
Henriksen
A Z
AZ
Department of Microbiology, Faculty of Medicine, University of Trondheim, Norway.
Maeland
J A
JA
D000911:Antibodies, Monoclonal / Q000276:immunology; D000915:Antibody Affinity / Q000187:drug effects / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis / Q000276:immunology* / Q000378:metabolism; D001798:Blood Proteins / Q000494:pharmacology*; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004754:Enterobacter / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D015151:Immunoblotting; D018272:Porins
D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins; D001798:Blood Proteins; D018272:Porins