Available systems for evaluation of degree of unsaturation of thyroxine-binding globulin are hampered at the bound-free separation step. In current assays, inorganic sorbants or ion-exchange resins are used to separate free 125I-labeled triiodothyronine from that bound to thyroxine-binding globulin. The techniques are laborious, time consuming, and not readily adaptable to total automation. In the assay we describe, triiodothyronine antibody, immobilized on the walls of polypropylene test tubes, is used as the bound-free separating agent in the evaluation of degree of unsaturation of thyroxine-binding globulin. The assay is simplified to four steps; washing, centrifugation, and use of columns are eliminated; and the procedure is readily automatable with existing pipetting equipment. Correlation with existing methods is excellent.

Filter paper discs saturated with dried blood can be used in the Immunotube solid-phase thyroxine radioimmunoassay. This assay utilizes polypropylene tubes to which antibody to thyroxine is covalently bound. The filter paper standards and samples are placed in the tubes, followed by an assay buffer that contains I125-labeled thyroxine and compounds to displace thyroxine from its binding proteins. After incubation, bound and free thyroxine are separated by aspirating or decanting the disc and buffer from the tube. The test can be used with 0.32 cm (1/8 inch) or 0.64 cm(1/4 inch) discs, and gives quantitative results that correlate well with those for serum samples. The intra-assay coefficient of variation is less than 10%. The assay may readily be mechanized with existing disc-punching equipment, and results of its use in mass screening programs are described.