VALIDATION OF AN INTERLEUKIN-1 BETA ASSAY FOR ASSESSING IMMUNE FUNCTION IN SALMONIDS
Aaron Frenette, University of Waterloo
Aaron Frenette1, Tania Rodríguez-Ramos1, Shawna Semple1, Fabio Zanuzzo2, Cheryl Soulliere1, Jack Iwanczyk3, Jessi Rix4, Devyn Ramsay2, Kurt Gamperl2 and Brian Dixon1
1 Department of Biology, University of Waterloo, Waterloo, ON,
2 Department of Ocean Sciences, Memorial University of Newfoundland, St. John’s, NL,
3 Cedarlane Laboratories Limited, Burlington, ON,
4 Somru BioScience Inc., Charlottetown, PE
Salmonid aquaculture in Canada faces several climate change-related challenges, including increasing water temperatures and hypoxic events, which may exacerbate bacterial and viral diseases. However, functional (and quantitative) immunological tools are needed if the industry is to select efficacious vaccines/treatments and to improve fish health/disease management, and we are to better understand how the fish immune system responds to such challenges. Genes encoding small signaling proteins (cytokines) that regulate immune/stress responses have been identified in many fishes, but their biological function(s) have not been elucidated. Herein, we report on the use of a quantitative enzyme-linked immunosorbent assay (ELISA) to measure interleukin-1 beta (IL- 1β) in rainbow trout (Oncorhynchus mykiss). IL-1β gene transcript levels, measured by qPCR, were consistent with an increase in IL-1β protein production/secretion in both cell culture (monocyte/macrophage cell line) and in primary spleen cells stimulated with heat-killed Vibrio anguillarum. In vivo assessment of the response of trout to acute heat-stress revealed that the expression of IL-1β transcripts in whole blood was consistent with the secretion of IL-1β protein in the plasma. In addition, assay validation was demonstrated by being able to identify putative IL-1β precursor and mature versions from fish tissues and cell cultures using Western blots. This research validates the use (effectiveness) of this assay, and suggests that the biological function(s) of IL-1β may be analogous to the mammalian paradigm for signaling and activation.