Z-Project: Analytical procedures for the determination of the thyroid hormone status in biological samples collected in the framework of the priority programme SPP 1629

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Thyroid hormones (TH) and their metabolites (THM) such as thyronamines (TAM), thyroacetic acids (TA) and several conjugated forms circulate in the blood in free and protein-bound form. During the first SPP funding period two immunoassays (IA) for 3,5-T2 and 3-T1AM in human serum have been optimized, validated and applied for data acquisition in cooperation with SPP 1629 partners. A liquid-liquid extraction (LLE)-based liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for detection of THM profiles in cell culture supernatants has been fully validated and further advanced for cell and tissue extracts using a modified pre-analytical approach involving solid phase extraction (SPE). Optimization and validation of LC-MS/MS methods including pre-analytical sample workup for human and mouse serum is still ongoing. Several experimental limitations in recovery and process efficiency caused by various matrix interferences lead to ion suppression in MS. Radioimmunoassays (RIA) for analysis of rT3 in human serum as well as T4 and T3 in mouse serum have been successfully applied for data generation together with SPP partners. During the second funding period the following aims will be pursued: (1) application of IA to define age and sex-specific reference range(s) for 3T1AM in human serum of children and adults, (2 characterization of development-, gender-, age-, and disease-specific THM profiles in human serum for (personalized) diagnostics and monitoring of therapeutic interventions, (3) completion of validation of THM LC-MS/MS analytics for human and mouse serum once quality control parameters requested by authorities have been accomplished, (4) optimization of pre-analytic TAM extraction from serum-containing matrices. The Z-project as a central platform will provide validated IA and MS-based methods to SPP partners for analytical tasks and quantitative analysis of THM profiles in clinical and experimental samples.