Award Number: N00014-14-1-0425 https://www.sfos.uaf.edu LONG-TERM GOALS To promote the use of endocrine techniques to advance our knowledge of the physiology of cetaceans and their responses to change in their environment. OBJECTIVES Two objectives are being nested under the overall goal of developing techniques that can be used for the detection of stress and pregnancy in large whales. 1)The first objective is to develop and conduct analytical and preliminary biological validations of pregnancy and stress hormones for large whales (humpback whales, blue whales, and possibly insular false killer whales). 2) The second objective is to complete the biological validation using archived samples from whales with known life history or behaviors. APPROACH The project has a two year approach in which the first year will serve as the proof of concept and the second year will complete the validation and add in life history and behavior data. Details of each year's activities are separated below. Year 1-Commercially available radioimmunoassays (RIA) previously validated in other marine mammal species will be validated for corticosterone or cortisol and progesterone in humpback whale blubber. The validation will follow two forms, analytical and biological, described below. Analytical Validation: The analytical validation will be conducted using standard methods of parallelism, accuracy and metabolite identification using high-pressure liquid chromatography (HPLC). Briefly, pooled blubber extract from animals of known gender will be serially diluted 1:2 and run as samples, in duplicate, in the assay to determine displacement by the pool. Accuracy of the assay will be determined by spiking the pool at the appropriate dilution with known amounts of hormone then performing the assays. Mass of the pool will be calculated and subtracted from the mass measured