Serologic Demonstration of Dual Specificity of Rabbit Bivalent Hybrid Antibody

Abstract

Treatment of rabbit 7S antibody with pepsin results in the removal of an inactive fragment, leaving a bivalent residue with an average molecular weight of 106,000 and a sedimentation coefficient of approximately 5S (1, 2). Subsequent reduction of one labile disulfide bond splits this residual molecule into two univalent, non-precipitating fragments which migrate as a single 3.5S peak (3) in the ultracentrifuge. These fragments closely resemble the univalent fractions of a papain digest (4) in a number of their properties.The univalent 3.5S fragments obtained by peptic digestion and reduction of a specifically purified rabbit antibody can be recombined by oxidation, in yields up to 75 per cent, to give 5S antibody which again possesses precipitating activity (5). When a mixture of purified rabbit antibodies of two different specificities is reduced and reoxidized, the recombined product contains a considerable amount of bivalent 5S "hybrid antibody" (6, 7). 1 The recombination of fragments of two different specificities appears to be essentially random, as shown by quantitative analysis of the hapten-binding capacity of hybrid antiovalbumin-antibenzoate antibody before and after treatment with an adsorbent specific for the antiovalbumin (7).Previous reports from these and other laboratories demonstrated that the 5S derivatives of several 7S rabbit antibodies retain most of the original agglutinating activity on a molar basis (8, 9); the 3.5S derivatives lack agglutinating properties but combine with the homologous antigen, as shown by inhibition of hemagglutination (9, 10) or by a modified antiglobulin reaction (8).The present communication extends these hemagglutination studies to 5S antibodies obtained by reoxidation of the 3.5S univalent fragments.

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Some improvement in tissue preparation and colloidal‐gold immunolabeling for electron microscopy

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