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A direct comparison of liquid chromatography-mass spectrometry with clinical routine testing immunoassay methods for the detection and quantification of thyroid hormones in blood serum

Authors
  • Bowerbank, Samantha L.
  • Carlin, Michelle G.
  • Dean, John R.
Type
Published Article
Journal
Analytical and Bioanalytical Chemistry
Publisher
Springer-Verlag
Publication Date
May 11, 2019
Volume
411
Issue
13
Pages
2839–2853
Identifiers
DOI: 10.1007/s00216-019-01724-2
Source
Springer Nature
Keywords
License
Green

Abstract

A new and improved method was developed for the determination and quantification of four “free” thyroid hormones (i.e. 3,5-diiodothyronine (T2), 3,3′,5-triiodothyronine (T3), 3,3′,5′-triiodothyrone (rT3) and 3,5,3′,5′-tetraiodothyronine (T4)) in human serum by low- and high-resolution liquid chromatography-mass spectrometry (LC-MS). Several sample preparation strategies were investigated to obtain matrix-independent results. These strategies included solid phase extraction and matrix dilution. The developed analytical methods were then directly compared, in a blind study using patient-derived human blood serum samples, to the current clinical routine testing methods, i.e. electrochemiluminescence immunoassay and enzyme-linked immunosorbent assay. Chromatographic separation was achieved on a pentafluorophenyl (F5) column with an isocratic method of 30% aqueous phase, 70% organic phase where mobile phase A is 0.1% formic acid in water (pH 4) and mobile phase B is 0.1% formic acid in methanol (pH 4) (v/v). The high-resolution LC-MS was able to give a significant improvement in sensitivity with limits of quantification of 0.002 to 0.008 pmol/L for all four “free” thyroid hormones, as well as reduced sample preparation, making this the preferred method. However, the increase in capital cost may be beyond the capabilities of some laboratories. The LC-MS methods allow for the analysis of “free” thyroid hormones to be carried out in a significantly reduced analysis time. Clinical sample analysis showed that there was no statistical difference between the results obtained by ECLIA/ELISA and both LC-MS methods. Graphical abstract

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