Improved ultrafiltration method for simultaneous measurement of free thyroxin and free triiodothyronine in serum.

Abstract

We describe a new ultrafiltration method for measuring concentrations of free thyroxin (FT4) and free triiodothyronine (FT3). Serum containing [131I]T4 and [125I]T3 is diluted 10-fold in phosphate buffer (500 mmol/L, pH 7.4 at 37 degrees C), then passed through a YMT membrane in an Amicon MPS-1 centrifugal ultrafiltration device. Radioactive iodide and protein-bound thyronines are separated from FT4 and FT3 in the ultrafiltrate by use of a Sephadex G-25 microcolumn. In normal controls, the fraction of T4 that passes into the ultrafiltrate is 0.021 +/- 0.006% (mean +/- SD), FT4 is 19.6 +/- 6.5 ng/L, the fraction of T3 that passes into the ultrafiltrate is 0.18 +/- 0.5%, and the FT3 is 2.03 +/- 0.50 ng/L. Intra-assay precision (CV) is 5.4% for FT4 and 4.2% for FT3; the respective interassay CVs are 10.1% and 8.0%. In various groups of patients with thyroid disease and other conditions that influence serum T4 and T3 concentrations, FT4 by ultrafiltration correlated with FT4 measured by equilibrium dialysis (r = 0.84, p less than 0.001) and FT3 by ultrafiltration correlated with FT3 measured by the Diagnostic Products RIA kit (r = 0.87, p less than 0.001). The accuracy, reproducibility, speed, and simplicity of the ultrafiltration method compared favorably with the more cumbersome method of equilibrium dialysis.