Binding of Thyroxine by Serum Proteins Evaluated by Equilibrum Dialysis and Electrophoretic Techniques. Alterations in Nonthyroidal Illness.

Considerable evidence suggests that the concentration of free thyroxine, circulating unbound to serum proteins, is more closely relattd to a subject's thyroidal status than is the concentration of total thyroxine (1). Because of the intense binding of thyroxine by the serum proteins, the concentration of free thyroxine is very small and not measurable by conventional chemical methods. Robbins and Rall (2), on the basis of electrophoretic data and the binding characteristics of bovine serum albumin, estimated that the concentration of free thyroxine in normal subjects was in the order of 6 x 10-1" M. By equilibrium dialysis of whole serum, Sterling and Hegedus (3) arrived at a figure approximately twice this. Other techniques for assessing the relative concentrations of free thyroxine in various clinical and physiological states include measurement of the fractional rate of dialysis of I'1"-labeled thyroxine (T4-I131 ) across a semipermeable membrane from one serum compartment to another (4, 5), and simultaneous determination of red-cell uptake of I131-labeled triiodothyronine (T3-"131) and serum protein-bound iodine (PBI) (6). In the present study, we used two independent methods to provide a relative measure of the fraction of thyroxine bound to serum proteins and the concentration of free thyroxine in serum. One method is based on the principle of equilibrium dialysis of diluted serum in an aqueous buffer at pH1 7.4; the other involves determination of critical ratios by filter-paper electrophoresis of whole