Epitope mapping of human thyrotropin.
Epitope mapping of hTSH was carried out using 19 monoclonal antibodies prepared with hTSH or its beta-subunit as antigen. The affinity constants of the monoclonal antibodies ranged from 9.6 X 10(7) to 5.7 X 10(9) mol/l for hTSH. The binding activities of monoclonal antibodies were maintained or in some cases rather enhanced after removal of the sugar moiety of the subunits of hTSH, and completely diminished after reduction of intramolecular S-S bonds in the subunits of hTSH. Ten monoclonal antibodies recognized the epitopes on hTSH (alpha:beta subunit combined form) and on free alpha-subunit form. Eight other antibodies recognized the epitopes on free/or combined form of beta-subunit, all of which did not recognize any other human glycoprotein hormones. The monoclonal antibodies directed against the alpha-subunit could bind also other human glycoprotein hormones to a varying extent. On the basis of results from competitive binding studies, the antibodies directed against alpha-subunit and those against beta-subunit were each classified into five subgroups recognizing different antigenic determinants. The remaining one antibody recognized an epitope expressed only by hTSH and not by the free subunits. In addition, a positive cooperativity on the binding of hTSH was observed between monoclonal antibodies directed towards a particular epitope on the alpha-subunit and those towards a epitope on the beta-subunit. From these data, two-dimensional map of epitopes on hTSH was constructed. The epitopes on each subunit were found to form a cluster with complicated overlapping, suggesting a highly conformational structure.
Endo
Y
Y
Department of Laboratory Medicine, Osaka University Medical School, Japan.
Miyai
K
K
Iijima
Y
Y
Nakajima
T
T
Eda
Y
Y
Fujita
H
H
Unoki
M
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001667:Binding, Competitive; D002459:Cell Fusion; D000939:Epitopes / Q000276:immunology*; D006801:Humans; D051379:Mice; D013154:Spleen / Q000166:cytology; D013972:Thyrotropin / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D013972:Thyrotropin