A new method for measuring thyroid hormones using nano-LC-MS/MS

Abstract

This paper describes a novel mass spectrometry based analytical method for analyzing thyroid hormones (THs). Thyroid hormones play a critical role in the regulation of many biological processes such as growth, metabolism and development. Several analytical methods using liquid chromatography-mass spectrometry (LC-MS) and tandem mass spectrometry (LC-MS/MS) have previously been developed to measure THs, especially in humans. For biomedical and toxicological research using small animal models, and in ecophysiological research using wild species where sample volume is limiting, sensitive methods are needed. In this study, we developed a nano-LC-MS/MS method enabling quantification of low concentrations of two key THs, thyroxine (T4) and 3,3',5-triiodothyronine (T3). The method was tested with egg yolk samples. We used a low flow rate (300 nl/min) to obtain maximal sensitivity of the method. The limit of quantitation was 10.6 amol for T4 and 17.9 amol for T3. The method shows good linearity (r > 0.99), repeatability and reproducibility (CVs <10%). We also reanalyzed yolk samples with radioimmunoassay for a comparison of the newly developed and previously used methods. Finally, we applied the methodology to measure hormones in egg yolk extracts in multiple avian species, and report interesting variation in maternal TH deposition. The newly developed nano-LC-MS/MS method is thus suitable for measuring THs in low concentrations and across species.

Testing for context-dependent effects of prenatal thyroid hormones on offspring survival and physiology: an experimental temperature manipulation

“…Plasma T3 and T4 levels were analyzed using nano-LC–MS/MS following previously published methods 39 and are expressed as ng/ml. Two randomly picked samples per nest were selected due to logistical constraints.…”

Effects of parental exposure to glyphosate-based herbicides on embryonic development and oxidative status: a long-term experiment in a bird model

“…Eggs were thawed and the yolk and shell were separated and weighed (accuracy 1 mg). T3 and T4 were measured from yolk following previously published methods 46 and were expressed as pg/mg yolk.…”

Testing for context-dependent effects of maternal thyroid hormones on offspring survival and physiology: an experimental approach manipulating temperature in a wild bird species

“…Plasma T3 and T4 levels were analyzed using nano-LC-MS/MS following previously published methods (36) and are expressed as ng/ml. Two randomly picked samples per nest were selected due to logistical constraints.…”