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Many freshwater habitats around the world suffered dramatic water chemistry changes over the last century mostly due to anthropogenic activities, including an overall reduction in pH due to high sulfur emissions and unsustainable forestry practices. One consequence of this change in water chemistry is a drop in available calcium concentration, which creates problems for aquatic organisms that rely on dissolved calcium to build their exo- or endoskeletons and reinforce their carapace during regular molts. Daphnia populations in shield lakes in northern Ontario are also exposed to other stressors, including copper, which persists at high concentrations in many of these freshwater lakes and ponds due to mining and other human activities. Copper toxicity on animals is influenced by the availability of other competing ions, such as calcium. Using our newly developed high throughput toxicity screening system, we show that mortality of Daphnia pulex increases with exposure to low calcium (0.05 mg L-1) and high copper (300 µg L-1). When these two stressors were combined, we found that copper was less toxic at high calcium concentrations, indicating a protective effect of calcium against copper toxicity. We then established basic calcium uptake kinetics in D. pulex using radioactive tracer 45Ca and provide evidence that copper, at environmentally relevant concentrations, competes with calcium uptake based on Km and Vmax. Our data show that both calcium decline and copper increase in aquatic ecosystems may negatively impact natural Daphnia populations, and that interactions between these two metals may occur in natural environments that result in fitness consequences for zooplankton.