A Sensitive Radioimmunoassay for T3 and T4 Determinations in Plasma and Tissues

Abstract

This chapter details protocols for determining plasma thyroid hormone (TH) levels and tissue TH content by competitive radioimmunoassays (RIAs). These protocols include: an initial test of the chromatographic performance, isotopic labeling to produce high activity ¹²⁵I-T3 and ¹²⁵I-T4, free iodide estimation of the labeled products, purification of tracers from iodide by paper electrophoresis, extraction of THs from plasma and tissue samples, and the RIA procedures. The RIA involves the competition between radioactive labeled and unlabeled hormones for specific antibody binding, and due to its high sensitivity is capable of detecting a minimum of 2.5 pg of T4 and 0.4 pg of T3.Drs. María Jesús Obregon and Gabriella Morreale de Escobar improved these protocols in the 1970s, enhancing sensitivity and accuracy. Their improvements enabled the detection of TH content in tissues, providing crucial insights into maternal THs role in fetal development and the importance of iodine intake during gestation. Their research also facilitated the early detection of congenital hypothyroidism, preventing neurological impairments in newborns. Internationally and in Spain, their contributions are widely acknowledged, leading to substantial public health impacts, including the implementation of nationwide neonatal screening programs. Despite progress, thyroid diseases remain prevalent, underscoring the need for continued research into thyroid physiology and associated disorders, employing highly sensitive techniques like the RIA outlined herein that we continue using in our research.

Keywords: Chromatography; Hypothyroidism; Iodide; Radioactivity; Radioimmunoassay; T3; T4; Thyroid hormones; Thyroxine; Triiodothyronine.