Simultaneous measurements of free thyroxine and triiodothyronine fractions in human serum by means of equilibrium dialysis (author's transl)

Abstract

Simultaneous measurements of serum free thyroxine (T4) and triiodothyronine (T3) fractions were studied using a modification of equilibrium dialysis described by Sterling and Brenner. To 1.2 ml of the serum to be assayed, 131I-T4 and 125I-T3 were added in a concentration of 2 mug/dl and 25 ng/dl, respectively, both of which were preliminarily dialysed according to Schussler and Plager. Half ml of the serum with tracers added was dialysed against 9 ml of phosphate buffer (ionic strength 0.15, pH 7.4) for 18 hours at 37 degrees C and 0.1 ml was reserved from the rest in a counting tube (in duplicate). After the completion of dialysis, the dialysate was mixed with 1 ml of pool serum and the contaminating inorganic iodide (in the form of 131I or 125I) was eliminated by adsorption on anion exchange resin. The radioactivity of 3 ml of the dialysate and 0.1 ml of the preserved serum (with the tracers added) was counted and the free (or dialysable) fractions were expressed as a ratio of the count of the former divided by that of the latter adjusted to an equal volume by calculation. The amount of either T4 or T3 added as tracers had no influence on free T4 or T3 fraction unless either of them was added to a concentration of 10 mug/dl. When 125I-T3 of low specific activity (50 muCi/ug) was used as a tracer, free T3 fraction measured simultaneously with free T4 fraction tended to be higher than that measured with a single tracer. When 125I-T3 of higher specific activity (300 muCi/mug) was employed, free T3 fraction obtained with two methods did not differ significantly. Using serum T4 and T3 concentrations measured by competitive protein binding analysis and radioimmunoassay, respectively, the free T4 and T3 concentrations were estimated with sera of normal, hyperthyroid, hypothyroid and uncomplicated pregnant subjects.