Current toxicity tests with microalgae are often criticized as being overly sensitive to metals because algae are cultured in metal-deficient media. If such bioassays overestimate copper toxicity in surface waters, the relevance of water quality guidelines derived from these tests is questionable. In this study, the effect of acclimation to copper at environmentally relevant concentrations, on the sensitivity of the marine diatom Nitzschia closterium and the freshwater green alga Chlorella sp. to copper and zinc was examined. N. closterium was acclimated in culture medium containing 5 or 25 g Cu L-1 for 200 days, while Chlorella sp. was acclimated in medium containing 2 g Cu L-1 for 100 days. Changes in algal growth rates and copper and zinc tolerance were monitored using standard growth inhibition toxicity tests in minimal medium over 72 h. Neither of the two acclimated N. closterium cultures had increased zinc or copper tolerance compared with that of the nonacclimated algae, nor were there any changes in control growth rates. Similarly, no changes in copper tolerance or control growth rates were observed for the acclimated Chlorella sp. culture. This was supported by measurements of intracellular and extracellular copper which confirmed that there were no differences in copper accumulation in either acclimated or nonacclimated algae. These results suggest that these algae grown in standard culture media are generally no more sensitive than algae grown in a metal-enriched medium. This supports the continued use of current laboratory bioassays with microalgae, as part of a suite of tests for assessing metal bioavailability, for use in ecological risk assessments and for providing data for the derivation of water quality guidelines. Copyright 2007 Wiley Periodicals, Inc. Environ Toxicol 22: 234-244, 2007.