A Quantitative in Vitro Study of the Chromatographic Distribution and Immunoglobulin Characteristics of Human Blocking Antibody

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Abstract
Summary: This report describes the immunoglobulin and chromatographic characteristics of human blocking antibody. Studies were carried out on a pool of serum from 20 allergic donors who had been immunized with ragweed antigen E as a form of therapy for ragweed hay fever. The serum and serum fractions were subjected to chromatography on DEAE-Sephadex, CM-cellulose, DEAE-cellulose and Sephadex G 200. Blocking activity was assayed by an in vitro technique: the inhibition of antigen E induced histamine release from isolated human leukocytes. This technique is more than 10 times as precise as the usual skin assay. Immunoglobulins were determined qualitatively by immunodiffusion against specific antisera and quantitatively by the Preer technique. Recovery of blocking activity was about 70%. In each fractionation procedure blocking antibody and IgG eluted together. Purified IgA had less than 1% the activity of IgG preparations and IgM antibody contributed less than 1% to the activity of the whole serum. Absorption of the serum with anti-IgA and treatment with 2-mercaptoethanol did not decrease its blocking titer, while absorption with anti-IgG caused a decrement of more than 97%. Circulating reaginic antibody appeared to account for less than 1% of the total serum activity. It was concluded that blocking activity is essentially a function of IgG molecules.

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