Equine anti-hapten antibody-V the non-precipitability of bivalent antibody

Abstract

The basis for the non-precipitability of the bivalent γG(T) anti- p-azophenyl- β-lactoside (Lac) antibody, in its interaction with human serum albumin multiply conjugated with Lac groups (Lac-HSA), has been sought by examination of complex formation both by γG(T) and γG anti-Lac antibody with Lac-HSA and Lac dye. Sedimentation analysis of mixtures of antibody and Lac-HSA has been carried out both in antibody and antigen excess. These experiments have provided an estimate of the molar composition of the complexes formed. Further clarification has been provided by measurement of the binding of Lac dye γG and γG(T) antibody and the inhibition of such binding by Lac-HSA. The capability of a molecule of anti-Lac antibody to bind simultaneously one molecule of Lac-HSA and one of Lac dye or two molecules of Lac-HSA has been evaluated by the use of an immunoadsorbent consisting of Lac-HSA covalently linked to cellulose. The capacity of the immunoadsorbent, layered on a filter pad, to bind antibody specifically was initially ascertained with radiolabeled antibody. Such immobilized antibody, bound to the Lac-HSA of the immunoadsorbent, was examined for its capacity to bind from solution radiolabeled Lac-HSA or Lac dye. The results of the several experimental approaches can be unified and interpreted by the notion that the γG(T) anti-Lac antibody in its interaction with Lac-HSA exhibits monogamous bivalency. That is, such antibody finds it energetically advantageous to occupy both of its combining regions with Lac groups present on a single protein molecule. This preference accounts for the non-precipitability of this antibody and distinguishes it from the γG anti-Lac antibody. The latter, however, in addition to its precipitability also shows considerable tendency to similar bivalent interaction.