Clin Chem Lab Med. 2025 Dec 18. doi: 10.1515/cclm-2025-0874. Online ahead of print.
ABSTRACT
OBJECTIVES: Free triiodothyronine (FT3) and free thyroxine (FT4) are important diagnostic markers for assessing thyroid function. However, their accurate quantification remains challenging due to low serum concentrations. Significant variability exists among current assay methods for measuring FT3 and FT4. This study aims to establish a candidate Reference Measurement Procedure (cRMP) for simultaneous quantification of serum FT3 and FT4 based on isotope-dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) technology.
METHODS: A convenient and reusable equilibrium dialysis (ED) device was utilized to separate free thyroid hormones from their protein-bound counterparts in serum. Key dialysis parameters, including temperature, pH, membrane type, and duration, were optimized to ensure consistent and reliable performance. The dialysate containing FT3 and FT4 was directly quantified by ID-LC-MS/MS. The method underwent systematic validation, comparative analysis with existing assays, and a comprehensive uncertainty assessment.
RESULTS: The developed cRMP demonstrated limits of quantification (LoQ) of 1.54 pmol/L for FT3 and 3.22 pmol/L for FT4, and with an imprecision of less than 3 %. No interference from endogenous analogs was observed, and the method showed good consistency in interlaboratory comparison. In contrast, chemiluminescent immunoassay results exhibited poor agreement with and the cRMP.
CONCLUSIONS: This study developed a highly precise, accurate, specific, and sensitive ID-LC-MS/MS-based cRMP for the simultaneous measurement of FT3 and FT4 in human serum. This method provides a reliable tool for standardizing routine thyroid function tests.
PMID:41405951 | DOI:10.1515/cclm-2025-0874