Dissolved nickel (Ni) typically displays a ‘nutrient-like’ vertical profile in the ocean, with lower concentrations in surface waters and higher concentrations in deep waters, similar to other micronutrient metals such as iron and zinc. Vertical profiles of Ni show particular similarities to profiles of the macronutrients phosphate and silicic acid, suggesting that diatoms play an important role in mediating the vertical distribution of this metal. We performed synchrotron x-ray fluorescence (SXRF) analysis on individual phytoplankton cells collected from stations in the equatorial Pacific Ocean and from nutrient-addition incubation experiments conducted on the same cruise. Diatoms were enriched in Ni twofold to fivefold relative to picoplankton and flagellated cells. Changes in cellular quotas of Si, P and Ni observed in diatoms growing in response to Fe and Si additions were used to estimate the Ni:P (0.52 ± 0.10 mmol/mol) and Ni:Si (28 ± 13 μmol/mol) ratios of internal biomass and the frustule, respectively. Elevated internal Ni:P suggests a heightened role for urease or the Ni isoform of superoxide dismutase in diatoms (similar to cyanobacteria), while Ni associated with the frustule appears to contribute an additional 50% of cellular Ni found in the diatoms. The derived Ni:Si ratio for frustule material is comparable to Ni:Si ratios in published nutrient profiles, confirming the dominant role that diatoms play in ocean Ni biogeochemistry. While a molecular explanation for the association of Ni with frustules remains to be determined, this study demonstrates the unique biogeochemical insight that can be gained from microanalytical element analysis.