The required components for assessing the binding capacity of human serum are enzymeT3 conjugate, thyroxine, binding protein (P), and immobilized thyroxine antibody (Ab). Upon mixing the enzymeconjugate and thyroxine with the specimen, a binding reaction results between the patient’s binding proteins and the added thyroxine but not with the enzyme conjugate. The added thyroxine (T4) not consumed in reaction 1 then competes with the enzymeT3 conjugate for a limited number of insolubulized binding sites. After equilibrium is attained, the antibodybound fraction is separated from unbound enzymeantigen by decantation or aspiration. The enzyme activity in the antibodybound fraction is directly proportional to the binding capacity of the specimen. Thus, in hypothyroidism, the binding proteins are relatively unsaturated (due to the low level of thyroid hormones) resulting in higher consumption of the added thyroxine than a euthyroid specimen. This leads to higher binding of the enzymetriiodothyronine conjugate caused by the reduced concentration of the available thyroxine. In hyperthyroidism, the reverse is true. The binding proteins are relatively saturated with thyroxine (due to the high level of thyroid hormone) resulting in lower consumption of the added thyroxine. The remaining thyroxine is relatively much higher than an euthyroid specimen resulting in lower enzymeantigen antibody binding due to the increased competition of the thyroxine for the limited antibody sites.