The Effect of an Ethanol-Extractable Thyroxine Binding Substance in Plasma on the Measurement of Thyroxine by Competitive Binding

Abstract

A T₄ binding substance was consistently extracted from normal serum by the standard extraction method used in the T₄-by-competitive-protein-binding-assay. In the usual assay this substance was present in the aqueous phase in the unknowns but not in the standards. It spuriously increased ¹²⁵I-T₄ in the aqueous phase of the unknowns causing an underestimation of T₄ of 12% when assayed in barbital buffer pH 8.6 using resin for separation of free and bound ¹²⁵I-T₄, and 25% in Tris buffer pH 7.4 using Sephadex. The substance also bound ¹²⁵I-T₄ when added to the standard curve system, and when dried serum extracts were resuspended in buffer-tracer only. Centrifugation of an aliquot of the aqueous phase gave a deposit having much more ¹²⁵I-T₄ per mg than the aqueous phase, the ¹²⁵I-T₄ concentration of which was significantly reduced. This procedure enabled the corrected T₄ figures to be calculated. This substance may be significant cause of the lower T₄ values measured by competitive binding as compared with PBI.