The comparison of the ability of monoclonal antibodies directed to different proteins (human IgG, human myoglobin and HRP) and bispecific antibodies derived thereof to bind antigens immobilized on a surface of a solid phase.
BACKGROUND
Bindings of mouse monoclonal antibodies (mAbs) and affinity purified bispecific antibodies (bAbs), derived thereof, to antigens adsorbed on immunoplates have been compared, using ELISA and RIA methods.


METHODS
The analysed panel of antibodies included mAbs specific to human myoglobin (Mb), human IgG (hIgG) and horseradish peroxidase (HRP) and biologically produced bAbs with double specificity to Mb and HRP, and to hIgG and HRP.


RESULTS
The degree of difference between different mAbs and corresponding bAbs varied markedly from antibody to antibody, depending on whether the parental mAbs could bind immobilized antigens bivalently. The observed equilibrium binding constant (K(obs)) for anti-HRP mAbs was 21-38 times higher that of anti-HRP site of bAbs (anti-hIgG/HRP or anti-Mb/HRP, respectively), due to bivalent binding of mAbs. Anti-Mb mAbs also bound bivalently with immobilized Mb. On the contrary, anti-hIgG mAbs bound monovalently with immobilized hIgG in the same conditions. The avidity of anti-Mb/HRP bAbs increased, if both antigens were simultaneously adsorbed on a solid phase.


CONCLUSIONS
The obtained data indicate that the use of bAbs in heterogeneous immunoassays instead of traditional mAb-enzyme conjugates hardly can provide the significant gain in assay performance if parental mAbs bind bivalently.
Dmitriev
D A
DA
Division of Chemical Enzymology, Department of Chemistry, Moscow State University, Moscow, Russia. dmitr@rcmh.msk.ru
Massino
Y S
YS
Segal
O L
OL
Smirnova
M B
MB
Kolyaskina
G I
GI
Pavlova
E V
EV
Osipov
A P
AP
Egorov
A M
AM
Dmitriev
A D
AD
D000327:Adsorption; D000818:Animals; D018033:Antibodies, Bispecific / Q000276:immunology / Q000378:metabolism*; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000378:metabolism*; D000915:Antibody Affinity / Q000276:immunology; D000937:Antigen-Antibody Reactions / Q000276:immunology*; D000941:Antigens / Q000276:immunology / Q000378:metabolism; D001666:Binding Sites, Antibody / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D006735:Horseradish Peroxidase / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D006825:Hybridomas; D055767:Immobilized Proteins; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D009211:Myoglobin / Q000276:immunology / Q000378:metabolism*; D011863:Radioimmunoassay
D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D000941:Antigens; D055767:Immobilized Proteins; D007074:Immunoglobulin G; D009211:Myoglobin; D006735:Horseradish Peroxidase