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 pur ified bispecific antibodies (bAbs), derived thereof, to antigens adsorbed. on immunoplates have been compared, using ELISA and RIA methods. Methods: T he analysed panel of antibodies included mAbs specific to human myoglobin ( Mb), human IgG (hIgG) and horseradish peroxidase (HRP) and biologically pro duced bAbs with double specificity to Mb and HRP, and to hIgG and HRP, Resu lts: The degree of difference between different mAbs and corresponding bAbs varied markedly from antibody to antibody, depending on whether the parent al mAbs could bind immobilized antigens bivalently. The observed equilibriu m 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 immobili zed Mb. On the contrary, anti-hIgG mAbs bound monovalently with immobilized hIgG in the same conditions. The acidity 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 immunoassa ys instead of traditional mAb-enzyme conjugates hardly can provide the sign ificant gain in assay performance if parental mAbs bind bivalently. (C) 200 1 Elsevier Science B.V. All rights reserved.