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Clinical Chemistry 39: 879-883, 1993;
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Clinical Chemistry, Vol 39, 879-883, Copyright © 1993 by American Association for Clinical Chemistry

Role of antibody valency in hapten-heterologous immunoassays

U Piran, D Silbert-Shostek and EH Barlow
Ciba Corning Diagnostics Corp., Walpole, MA 02032.

We studied the effects of hapten heterology on immunoassays of triiodothyronine (T3), digoxin, and cortisol, in a format involving labeled monoclonal antibodies and immobilized, protein-conjugated ligands. Replacing the homologous conjugated ligands T3, digoxin, and cortisol with their respective analogs diiodothyronine, digitoxin, and corticosterone led in each case to a decrease in the midpoint of displacement (ED50) for the same zero-dose signal. The mechanism of this phenomenon was studied by converting the bivalent anti-T3 to a monovalent whole antibody (bispecific monoclonal anti-T3 x anti-glucose- 6-phosphate dehydrogenase) by cell fusion. The monovalent antibody was effective as a tracer in the homologous T3 assay, but generated a very low zero-dose signal with the heterologous solid phase, thus precluding sensitivity enhancement. On the basis of these results and additional kinetic and double-labeling experiments, we propose that the use of hapten heterology relies on bivalent binding of the antibody to the solid phase to compensate for a lower intrinsic affinity. This binding mechanism leads to lower assay concentrations of the ternary complex analyte-labeled antibody-immobilized hapten, thereby providing enhanced sensitivity.


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Copyright © 1993 by the American Association for Clinical Chemistry.