Free thyroid hormone: Methods and standardization

• Published in: Clinica chimica acta 2025-01, Vol.565, p.119944, Article 119944
• Main Authors: Ma, Zijia, Liu, Zhenni, Deng, Yuhang, Bai, Xuanchang, Zhou, Weiyan, Zhang, Chuanbao
• Format: Article
• Language: English
• Subjects: Chromatography, Liquid - standards; FT3; FT4; Humans; Immunoassay; Immunoassay - standards; Interference Factors; LC-MS/MS; Reference Standards; Standardization; Tandem Mass Spectrometry - standards; Thyroid Function Tests - standards; Thyroid Hormones - analysis; Thyroid Hormones - blood; Thyroid Hormones - standards; Thyroxine - analysis; Thyroxine - blood; Triiodothyronine - analysis; Triiodothyronine - blood
• ISSN: 0009-8981; 1873-3492; 1873-3492
• DOI: 10.1016/j.cca.2024.119944

•Free thyroid hormone can enter directly into cells to act, so its detection is important in diagnosing thyroid disease.•Equilibrium dialysis or ultrafiltration facilitates the precise separation of the free state of the hormone.•We discuss the current status of free thyroid hormones immunoassay detection and advances in LC-MS/MS detection.•LC-MS/MS combined with equilibrium dialysis or ultrafiltration is expected to achieve high-throughput detection. Free thyroid hormone (FTH) serves as the preferred indicator for the clinical assessment of thyroid function, mainly encompassing free thyroxine and free triiodothyronine. The immunoassay commonly employed in the clinical setting exhibits certain unresolvable deficiencies. The results of over 5,500 clinical laboratories for FTH from China in 2024 demonstrated that the outcomes of immunoassay were not comparable, with robust CVs calculated in accordance with ISO 13528 ranging from 13.82% to 21.42%. Establishing reference methods is an important tool to achieve accurate and comparable results of free hormones. Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) holds a distinct advantage in the precise detection of small molecules, and two reference methods for free thyroxine based on LC-MS/MS are included in the JCTLM list. This article conducts a comprehensive review of the detection methods and standardization of FTH. It presents the metabolism of thyroid hormones, the significance of detection, the techniques, and application examples of free thyroid hormone assays, and deliberates on the current status, prospects, and recommendations for the standardization of FTH assays. Immunoassay and LC-MS/MS, as significant techniques for FTH detection, are predominantly emphasized in the case references. Ultrafiltration and equilibrium dialysis, which are utilized to separate FTH, are also addressed. This article aims to discuss the status quo of FTH detection and clarify the advantages of LC-MS/MS in FTH detection, propose that LC-MS/MS can be utilized as an auxiliary validation method or alternative method in clinical applications, and offer suggestions for the standardization of testing results.