The aim was to validate a commercial enzyme immunoassay for the determination of salivary cortisol in donkeys. Seven stallions were included. Saliva samples were collected at 8:30 AM on thirteen not consecutive days by a cotton-based swab (Salivette, Germany). The swab was grasped with a clamp, inserted at the angle of the lips into the mouth of the donkey and placed gently under the tongue for 1 minute and afterwards returned into a polypropylene tube. Tubes were then centrifuged for 10 min at 700 g and the obtained saliva was frozen at 20°C until analysis. A commercial enzyme immunoassay without extraction (Demeditec Diagnostics, Kiel-Wellsee, Germany) was used for cortisol saliva determination. The assay was validated for donkey saliva by measuring recovery of cortisol standard, serial dilution curve parallelism, intra-assay and inter-assay coefficient of variation and limit of detection. Cortisol mean levels and standard deviation were calculated for each day of sampling. One-way ANOVA for repeated measures and Tukey’s test were performed. The ELISA method was found to be sensitive and reproducible for cortisol concentration determination in donkey saliva. Recovery of cortisol standard to donkey saliva was 107.9% and serial dilution of saliva samples with assay buffer resulted in changes in optical density parallel to the standard curve. The intra-assay coefficient of variation was 10.7%, the inter-assay variation was 8.0% and the minimal detectable concentration was 0.01 ng/mL. No statistical significant differences were found among cortisol levels in each animal on the 13 different days of collection.

DETERMINATION OF SALIVARY CORTISOL IN DONKEY STALLIONS

BONELLI, FRANCESCA;ROTA, ALESSANDRA;BARAGLI, PAOLO;PANZANI, DUCCIO;CAMILLO, FRANCESCO;GATTA, DOMENICO;MEUCCI, VALENTINA;SGORBINI, MICAELA
2017-01-01

Abstract

The aim was to validate a commercial enzyme immunoassay for the determination of salivary cortisol in donkeys. Seven stallions were included. Saliva samples were collected at 8:30 AM on thirteen not consecutive days by a cotton-based swab (Salivette, Germany). The swab was grasped with a clamp, inserted at the angle of the lips into the mouth of the donkey and placed gently under the tongue for 1 minute and afterwards returned into a polypropylene tube. Tubes were then centrifuged for 10 min at 700 g and the obtained saliva was frozen at 20°C until analysis. A commercial enzyme immunoassay without extraction (Demeditec Diagnostics, Kiel-Wellsee, Germany) was used for cortisol saliva determination. The assay was validated for donkey saliva by measuring recovery of cortisol standard, serial dilution curve parallelism, intra-assay and inter-assay coefficient of variation and limit of detection. Cortisol mean levels and standard deviation were calculated for each day of sampling. One-way ANOVA for repeated measures and Tukey’s test were performed. The ELISA method was found to be sensitive and reproducible for cortisol concentration determination in donkey saliva. Recovery of cortisol standard to donkey saliva was 107.9% and serial dilution of saliva samples with assay buffer resulted in changes in optical density parallel to the standard curve. The intra-assay coefficient of variation was 10.7%, the inter-assay variation was 8.0% and the minimal detectable concentration was 0.01 ng/mL. No statistical significant differences were found among cortisol levels in each animal on the 13 different days of collection.
2017
https://onlinelibrary.wiley.com/doi/10.1111/jvim.14649
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11568/836534
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