Effects of chronic copper exposure on a suite of indicators were examined: acute toxicity, acclimation, growth, sprint performance, whole-body electrolytes, tissue residues, and gill copper binding characteristics. Juvenile rainbow trout were exposed for 30 d to waterborne copper in hard water (hardness = 120 mg/L as CaCO3, pH = 8.0, Cu = 20 and 60 μg/L) and soft water (hardness = 20 mg/L as CaCO3, pH = 7.2, Cu = 1 and 2 μg/L). Significant acclimation to the metal occurred only in fish exposed to 60 μg/L, as seen by an approx. twofold increase in 96-h LC50 (153 vs 91 μg Cu/L). Chronic copper exposure had little or no effect on survival, growth, or swimming performance in either water hardness, nor was there any initial whole-body electrolyte loss (Na+ and Cl−). The present data suggest that the availability of food (3% wet body weight/day, distributed as three 1% meals) prevented growth inhibition and initial ion losses that usually result from Cu exposure. Elevated metal burdens in the gills and livers of exposed fish were measures of chronic copper exposure but not of effect. Initial gill binding experiments revealed the necessity of using radiolabeled Cu (64Cu) to detect newly accumulated Cu against gill background levels. Using this method, we verified the presence of saturable Cu-binding sites in the gills of juvenile rainbow trout and were able to make estimates of copper-binding affinity (log Kgill=Cu) and capacity (Bmax). Furthermore, we showed that both chronic exposure to Cu and to low water calcium had important effects on the Cu-binding characteristics of the gills.