Validation of an LC–MS/MS Method for Urinary Cortisol in Dogs and Reference Interval, with an Exploratory Comparison to Immunoassay
Year: 2025
Type: article
Abstract: Cortisol measurement in dogs commonly supports the diagnosis of hypercortisolism or hypoadrenocorticism. Unlike serum cortisol, urine cortisol is less affected by daily fluctuations, and collection is non-invasive, making the urine cortisol-to-creatinine ratio (UCCR) a useful diagnostic tool. Immunoassays are commonly used but lack specificity for free, biologically active cortisol. Liquid chromatography–tandem mass spectrometry (LC-MS/MS) is recommended to overcome this limitation. This study aimed to validate an LC-MS/MS method for urinary cortisol measurement, establish a reference interval for UCCR in dogs, and compare the results with the commercial immunoassay system Immulite® 2000 Xpi (Siemens, Cary, NC, USA). A comprehensive method validation was performed. Across different concentrations, the intra- and inter-assay coefficients of variation (CV) ranged from 2.39% to 9.45%, accuracy ranged from 87.7% to 105.5%, and bias varied from −12.3% to 5.53%, which were all within acceptable limits. The method demonstrated excellent linearity (r2 ≥ 0.98), and stability under various conditions was satisfactory (RSD: −12.4% to 13.6%). Interference from endogenous and exogenous corticosteroids was negligible. The established reference interval for UCCR was 0.21–2.84 × 10−6. The exploratory comparison with the immunoassay revealed systematic and proportional differences between methods. The LC-MS/MS method demonstrated excellent analytical performance in terms of accuracy and specificity, and showed suitability for clinical application.
Source: Animals
DOI: 10.3390/ani15182682
Language: English
Subfield: Small Animals
Field: Veterinary
Domain: Health Sciences
Open Access status: gold
Estimated APC paid: $2,896
OpenAlex ID: W4414182132