Salivary cortisol measurement in horses: immunoassay or LC-MS/MS?

Sauer, F. J.; Gerber, V.; Frei, S.; Bruckmaier, R. M.; Grössl, M. (2020). Salivary cortisol measurement in horses: immunoassay or LC-MS/MS? Domestic animal endocrinology, 72, p. 106445. Elsevier 10.1016/j.domaniend.2020.106445

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The aim of the present study was to measure salivary cortisol concentrations of horses before and after hypothalamic-pituitary-adrenal (HPA) axis stimulation by means of liquid chromatography-tandem-mass spectrometry (LC-MS/MS) and an immunoassay (cELISA) for method comparison. Nine clinically healthy horses participated in the study. An ACTH stimulation test was performed. Saliva samples were collected before (T0) and 60 (T60) min after intravenous injection of 1 μg/kg BW synthetic ACTH1-24. LC-MS/MS was assessed for the determination of equine salivary cortisol. The results of these measurements were then compared to the results obtained by a cELISA, which has previously been validated for use in horses. The Pearson correlation coefficient was calculated and showed no correlation at T0 (r = -0.2452; P = 0.5249) and significantly correlated results at T60 (r = 0.8334; P = 0.0053). Bland-Altman-Plots of T60 revealed that immunoassay measurements led to higher outcome values than LC-MS/MS. On average, immunoassay results were 2.3 times higher. Poor agreement between both methods at T0 is potentially a consequence of inaccuracy in the very low measuring range of the immunoassay, and to a smaller extent, structurally similar cross-reacting agents and matrix effects, which might bias the results. Overestimation of immunoassay results at T60 might be due to different standardization of both methods, non-avoidable matrix effects on the antigen-antibody interaction in the ELISA, and possibly cross-reactions of other steroids. While immunoassay measurements of equine salivary cortisol yielded higher but reasonably correlated results for elevated cortisol concentrations after stimulation of the HPA axis, LC-MS/MS provided more accurate results, particularly for baseline cortisol concentrations close to the limit of detection of the ELISA.

Item Type:

Journal Article (Original Article)


05 Veterinary Medicine > Research Foci > Veterinary Public Health / Herd Health Management
04 Faculty of Medicine > Department of Dermatology, Urology, Rheumatology, Nephrology, Osteoporosis (DURN) > Clinic of Nephrology and Hypertension
05 Veterinary Medicine > Department of Clinical Veterinary Medicine (DKV) > Equine Clinic
05 Veterinary Medicine > Department of Clinical Veterinary Medicine (DKV)
05 Veterinary Medicine > Department of Clinical Research and Veterinary Public Health (DCR-VPH) > Veterinary Physiology
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Unit Childrens Hospital > Forschungsgruppe Nephrologie / Hypertonie

UniBE Contributor:

Sauer, Fay Joanne; Gerber, Vinzenz; Bruckmaier, Rupert and Grössl, Michael


600 Technology > 610 Medicine & health
600 Technology > 630 Agriculture








Ursula Therese Horst

Date Deposited:

15 May 2020 15:46

Last Modified:

28 Apr 2021 11:44

Publisher DOI:


PubMed ID:


Uncontrolled Keywords:

ACTH stimulation test Horse Immunoassay Liquid chromatography-tandem-mass spectrometry Salivary cortisol




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