Enhancing reproducibility in stable isotope analysis (SIA) of fish eye lenses: A comparison between lamina number and diameter.

Analyzing stable isotopes in archival tissues, such as fish eye lenses, is used to document shifts in feeding ecology, diet, habitat use, and to reconstruct life history. Fish eye lenses grow throughout their ontogeny, forming multiple sequential layers, or laminae. These laminae represent the chronology of the fish's life, much like tree rings, which record environmental conditions over time. Lenses are protein-rich, which makes them an ideal structure for analyzing light isotopes such as δ¹³C, δ¹⁵N, and δ34S. These light isotopes are primarily integrated into the lens tissue through the fish's diet, where they are bound to amino acid structures during protein synthesis. As research begins to emerge using eye lenses to reconstruct the life histories of fishes, the need for a reproducible method of delamination grows. For this study, each researcher independently delaminated one lens from each of the 10 adult Chinook Salmon (Oncorhynchus tshawytscha). Lens lamina number, diameter (mm), and mass (mg) of each lamina were recorded. Laminae were then submitted for stable isotope analysis of both δ¹³C and δ¹⁵N. Isotope values were used as a validation to compare delamination patterns between researchers. δ¹³C and δ¹⁵N values from the lenses were then plotted using both the assigned lamina number and lens diameter to compare the difference between researchers. Analysis based on lamina number showed significant shifts in isotope values and variability in lamina counts between researchers. However, when lens diameter was used instead of lamina number, isotope patterns throughout the lenses of the same fish were nearly identical. Using lens diameter removes subjectivity between researchers, thereby increasing the reproducibility of the technique and providing a more robust interpretation of the data.