title: 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.
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.
authors.affiliation: Department of Animal Medicine and Surgery, Regional Campus of International Excellence Campus Mare Nostrum, University of Murcia, 30100 Espinardo, Murcia, Spain.
authors.forename: D
authors.initials: D
authors.lastname: Escribano
authors.forename: L
authors.initials: L
authors.lastname: Soler
authors.forename: A M
authors.initials: AM
authors.lastname: Gutiérrez
authors.forename: S
authors.initials: S
authors.lastname: Martínez-Subiela
authors.forename: J J
authors.initials: JJ
authors.lastname: Cerón
mesh_terms: D000818:Animals; D000906:Antibodies / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D053379:Chromogranin A / Q000378:metabolism*; D015200:Fluoroimmunoassay / Q000379:methods* / Q000662:veterinary; D008297:Male; D012463:Saliva / Q000378:metabolism*; D012680:Sensitivity and Specificity; D013312:Stress, Physiological; D034421:Sus scrofa / Q000502:physiology*
chemical_list: D000906:Antibodies; D015415:Biomarkers; D053379:Chromogranin A