Immunological characterization of rat kininogens with monoclonal antibodies to T-kininogen. Distinction between the different domains of T-kininogen and the multiple rat kininogens.
A panel of 16 monoclonal antibodies (mAb) were produced against rat T-kininogen to characterize this family of proteins. These mAbs bound 125I-T-kininogen by radioimmunoassay as well as reacting strongly with immobilized T-kininogen in an enzyme-linked immunosorbent assay (ELISA). The reactivity of these antibodies with proteolytic fragments of T-kininogen demonstrated the recognition of several different epitopes. One antibody was specific for the domain 1 of the heavy chain and/or the light chain, twelve antibodies were specific for domain 2 and three antibodies were specific for domain 3. All monoclonal antibodies recognized the two forms of T-kininogen encoded by the two different T-kininogen genes, TI and TII kininogen, except antibody TK 16-3.1 which uniquely reacted with TII kininogen. Two antibodies recognizing domain 2 cross-reacted with the high-molecular-mass kininogen (H-kininogen), whereas all the other monoclonal antibodies were specific to T-kininogen and did not recognize the heavy chain of H-kininogen. None of the antibodies tested altered the thiol protease inhibitory activity of T-kininogen, its partial proteolysis by rat mast cell chymase or the hydrolysis of H-kininogen by rat urinary kallikrein. The use of these antibodies in the development of sensitive ELISA to measure T-kininogen levels in plasma, urine, liver microsomes and hepatocytes is described. Two different forms of T-kininogen were distinguished by these monoclonal antibodies in Western blotting using rat plasma. The localization of T-kininogen was defined using these monoclonal antibodies by immunohistochemistry in rat liver hepatocytes and rat kidney.
Lesage
S
S
INSERM U36, Paris, France.
Bouhnik
J
J
Richoux
J P
JP
Baussant
T
T
Gauthier
F
F
Eager
K
K
Corvol
P
P
Alhenc-Gelas
F
F
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015153:Blotting, Western; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006868:Hydrolysis; D007150:Immunohistochemistry; D007668:Kidney / Q000378:metabolism; D007704:Kininogens / Q000276:immunology / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008297:Male; D011863:Radioimmunoassay; D051381:Rats; D011919:Rats, Inbred Strains
D000911:Antibodies, Monoclonal; D007704:Kininogens