Radioimmunoassay of thyroxine and triiodothyronine in urine using extraction and separation on Sephadex® columns

Abstract

Radioimmunological methods for the determination of thyroxine (T4) and triiodothyronine (T3) in urine have been developed. The methods are based on trapping of T4 and T3 from unextracted urine, followed by separation between free and antibody bound hormone on the same Sephadex® column. The T4 method has been compared to a competitive protein binding (CPB) assay using ethyl acetate extraction. The methods are evaluated in sixty-seven euthyroid controls, twenty-four hyperthyroid and seven hypothyroid patients. In the T4 RIA detection limit was 3.9 pg, intra-assay coefficient of variation (cv) was 5.2% and inter-assay cv was 6.9%. In the T3 RIA detection limit was 7.2 pg, intra-assay cv 3.9% and inter-assay cv 10.8%. Recovery of added amounts of hormones and serial dilutions gave satisfactory results. The CPB assay was found unreliable with unspecific and false high values. In euthyroid controls 24 h urinary T4 excretion as measured by RIA was 1.8±0.5 nmol, and urinary T3 excretion was 0.7±0.3 nmol. T4 and T3 excretion was greatly elevated in hyperthyroid patients and decreased in hypothyroidism.

Key Words:

• radioimmunoassay
• radioligand assay
• thyroxine
• urine
• triiodothyronine
• urine