Influence of Collection and Storage Conditions on Adrenocorticotropic Hormone (ACTH) Measurements in Bottlenose Dolphins (Tursiops truncatus)

Abstract

Identifying the effects of stressors on marine mammal health is critical for species conservation and collection. Adrenocorticotropic Hormone (ACTH) is a key indicator of peracute stress; however, handling methods may affect results. Thus, the aim of this study was to examine the effects of collection methods and storage temperature on the stability of bottlenose dolphin (Tursiops truncatus) ACTH measurements. Blood was collected from five managed-care bottlenose dolphins in four vacutainers ™ in the following sequence: EDTA, sodium heparin, serum separator, and 2nd EDTA. The specimens were centrifuged, processed immediately, aliquoted into plasma and serum, and divided into four equal volumes. These aliquots were immediately stored at either 4º or -80º C for 24 h prior to the ACTH assay. ACTH concentrations were compared between the collection tube types and the storage temperatures. There was a significant difference in ACTH concentration across collection tube types. ACTH concentrations were highest in ethylenediaminetetraacetic acid (EDTA) tubes and lowest in serum separator tubes at both temperatures. Twenty-four hour storage temperature did not have a statistically significant impact on mean ACTH concentration. There was no significant difference in ACTH concentration between the first and last drawn EDTA tubes. Bottlenose dolphin whole blood should be collected in EDTA for ACTH measurement. Samples may be refrigerated and shipped on cold packs if assayed within 24 h of collection or immediately frozen and shipped. When multiple samples are collected in various collection tubes at the same time, the order in which tubes are filled does not affect the ACTH measurement.

Stress response of wild bottlenose dolphins (Tursiops truncatus) during capture–release health assessment studies

“…Further details are described in Lewis et al, 2013. The assay range is 12-1250 pg/mL and analytical sensitivity is 9 pg/mL.…”

Polish Journal of Veterinary Sciences

“…Accordingly, the pT concentration was lower than the control (plasma stored in a freezer at -20°C). Degradation caused by blood enzymes is a factor for the instability of hormones (Lewis et al 2013).…”

“…Moreover, during plasma or serum preparation, delayed time of centrifugation of blood samples leads to variation in the concentrations of progesterone (De Castro et al 2004), adrenocorticotrophic hormone (Preissner et al 2004, Christensen et al 2016, and parathyroid hormones (Omar et al 2001). Moreover, storage conditions can also affect the stability of hormone concentrations (Lewis et al 2013, Prutton et al 2015. Having seen the evidence above, it is thus important to evaluate if these pre-analytical factors introduce systematic bias on measuring T concentrations in goats.…”