Production and characterization of monoclonal antibodies against human Cu/Zn superoxide dismutase and the establishment of a super-rapid enzyme-linked immunosorbent assay (SURALISA).
Murine monoclonal IgG1 antibodies directed against four different epitopes of human Cu/Zn superoxide dismutase (SOD) were produced by immunization with recombinant Cu/Zn SOD. The antibodies reacted well with the recombinant protein and Cu/Zn SOD purified from human erythrocytes, with binding constants ranging from 8.8 X 10(9) to 2.2 X 10(10) l/mol. When mixed, these antibodies completely prevented the binding of rabbit and sheep polyclonal antibodies raised against erythrocyte Cu/Zn SOD. Whereas one antibody was directed against a common homology region of bovine and human Cu/Zn SOD, all the other antibodies reacted exclusively or preferentially with human Cu/Zn SOD. Only one epitope on the human Cu/Zn SOD molecule was accessible at two different sites as demonstrated in a homologous two-site assay with one and the same antibody used as both capture and indicator antibody. In the indirect two-site assay with unlabelled monoclonal antibodies, and additive effect with a steeper dose-response curve was obtained by mixing antibodies against different epitopes. A super-rapid one-step two-site enzyme immunoassay (overall duration 20 min) was established with antibodies against two different epitopes. Its detection limit was 0.5 micrograms SOD/l.
Porstmann
T
T
Department of Medical Immunology, Medical School (Charité), Humboldt University, Berlin, G.D.R.
Wietschke
R
R
Grunow
R
R
Jahn
S
S
Porstmann
B
B
Schmechta
H
H
Pergande
M
M
Von Baehr
R
R
D000911:Antibodies, Monoclonal / Q000276:immunology; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis; D006801:Humans; D013482:Superoxide Dismutase / Q000032:analysis* / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D013482:Superoxide Dismutase