Larvae of Mytilus spp. are among the most Cu sensitive marine species. In this study we assessed the combined effect of salinity and dissolved organic carbon (DOC) on Cu accumulation on mussel larvae. Larvae were exposed for 48 h to three Cu concentrations in each of nine salinity/DOC treatments. Synchrotron radiation X-ray fluorescence was used to determine the Cu concentration in 36 individual larvae with a spatial resolution of 10 × 10 μm. Cu body burden concentrations varied between 1.1 and 27.6 μg/g DW larvae across all treatments and Cu was homogeneously distributed at this spatial resolution level. Our results indicate decreasing Cu accumulation with increasing DOC concentrations which can be explained by an increase in Cu complexation. In contrast, salinity had a nonlinear effect on Cu. This cannot be explained by copper speciation or competition processes and suggests a salinity-induced alteration in physiology.