12Otolith microchemistry is a widely used technique for elucidating life history patterns in 13 fishes. This typically involves sectioning the otolith and collecting elemental signatures 14 via laser ablation. But this requires time-intensive handling that may influence results. 15As an alternative to traditional cut/polish/ablate techniques, we tested depth-profiling 16 laser ablation, which offers reduced handling and contamination risk. To validate depth-17 profiling as a robust method for collecting trace element otolith microchemistry data, 18 we constructed composite otoliths using otolith materials from fishes of different 19 origins (freshwater, seawater). Test ablations were conducted on composite 20 diadromous otoliths at a range of spot sizes and pit depths. We measured tailing and 21 fractionation effects in the following elements; Na, Mg, K, Mn, Zn, Rb, Sr, and Ba. Given 22 appropriate instrument parameters, depth-profiling can accurately collect elemental 23 concentration data both between and within top and bottom layers of an otolith 24 composite across a range of spot sizes and pit depths. Analytical power and lag effects 25 were dependent on spot size, highlighting the importance of optimizing spot size based 26 on sample morphology and instrument parameters. 27 28

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