Endocrine Abstracts

Introduction: The measurement of free thyroxine (FT4) and free 3,3′,5-triiodothyronine (FT3) in serum by equilibrium dialysis immunoassay methods may suffer from a lack of specificity, therefore measurement by LC/MS/MS has the potential to increase the accuracy of the results. Here we present a sensitive method, employing a simple ultrafiltration sample preparation.

Description of methods: 400 uL of serum were processed for 90 minutes using a 3kDa centrifugal filter, and 50uL of the ultrafiltrate was directly injected onto an AB SCIEX Triple Quad® 6500 LC/MS/MS system.

Results: The method presented here achieved a Limit of Quantitation (LOQ) of <0.5 pg/mL for both FT3 and FT4, and demonstrated excellent linearity, accuracy and precision over the concentration range from 0.5 pg/mL to 100 pg/mL. A method comparison was performed, using a cohort of samples that had been previously analyzed by immunoassay, and an excellent correlation was obtained.

Conclusion: We have demonstrated a sensitive method for the analysis of FT3 and FT4 in serum by LC/MS/MS, which takes advantage of the simple and rapid sample preparation afforded by ultrafiltration. The method provides good sensitivity, accuracy and precision, enabling researchers to investigate FT3 and FT4 levels in serum across the expected range of concentrations. Unlike earlier MS-based methods, this method does not require a large sample injection volume, since only 50 uL of the final sample is required.