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Measurement of chromogranin A in porcine saliva: validation of a time-resolved immunofluorometric assay and evaluation of its application as a marker of acute stress

Published online by Cambridge University Press:  09 November 2012

D. Escribano ,
L. Soler ,
A. M. Gutiérrez ,
S. Martínez-Subiela  and
J. J. Cerón
D. Escribano
Affiliation:
Department of Animal Medicine and Surgery, Regional Campus of International Excellence ‘Campus Mare Nostrum’, University of Murcia, 30100 Espinardo, Murcia, Spain
L. Soler
Affiliation:
Division of Livestock-Nutrition-Quality, Biosystems Department, Biosciences Engineering Faculty, Katholieke Universiteit Leuven, 3001 Heverlee, Belgium
A. M. Gutiérrez
Affiliation:
Department of Animal Medicine and Surgery, Regional Campus of International Excellence ‘Campus Mare Nostrum’, University of Murcia, 30100 Espinardo, Murcia, Spain
S. Martínez-Subiela
Affiliation:
Department of Animal Medicine and Surgery, Regional Campus of International Excellence ‘Campus Mare Nostrum’, University of Murcia, 30100 Espinardo, Murcia, Spain
J. J. Cerón*
Affiliation:
Department of Animal Medicine and Surgery, Regional Campus of International Excellence ‘Campus Mare Nostrum’, University of Murcia, 30100 Espinardo, Murcia, Spain
*
†E-mail: jjceron@um.es
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Abstract

The objective of this study was to develop and validate a time-resolved immunofluorometric assay (TR-IFMA) for porcine salivary chromogranin A (CgA) measurements, using a species-specific antibody, and evaluate its behaviour in an acute stress model. Polyclonal antibodies were produced in rabbits immunized with a synthetic porcine fragment of CgA359−379 and used to develop a sandwich TR-IFMA. This TR-IFMA was analytically validated and showed intra- and inter-assay coefficients of variation of 6.23% and 5.82%, respectively, an analytical limit of detection of 4.27 × 10−3 μg/ml and a limit of quantification of 24.5 × 10−3 μg/ml. The assay also demonstrated a high level of accuracy, as determined by linearity under dilution (r = 0.975) and recovery tests. When a model of experimental acute stress, in which animals were immobilized for 3 min with a nose snare (stressor stimulus), was applied, a significant increase (P < 0.05) in CgA levels in saliva was detected at 15 min post-stressor stimulus. These results indicate that the assay developed in this study could measure CgA in porcine saliva in a reliable way and that the concentrations of CgA in saliva samples of pigs increase after an acute stress situation.

Keywords

Chromogranin A (CgA)TR-IFMAporcine salivastressvalidation
Type
Behaviour, welfare and health
Information
animal , Volume 7 , Issue 4 , April 2013 , pp. 640 - 647
Copyright
Copyright © The Animal Consortium 2012

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