Invasive species are commonly thought to have broad tolerances that enable them to colonize new habitats, but this assumption has rarely been tested. In particular, the relative importance of acclimation (plasticity) and adaptation for invasion success are poorly understood. This study examined effects of short-term and developmental acclimation on adult salinity tolerance in the copepod Eurytemora affinis. This microcrustacean occurs in estuarine and salt marsh habitats but has invaded freshwater habitats within the past century. Effects of short-term acclimation were determined by comparing adult survival in response to acute versus gradual salinity change to low salinity (fresh water). Effects of developmental acclimation on adult tolerance were determined using a split-brood 4 x 2 factorial experimental design for one brackish-water population from Edgartown Great Pond, Massachusetts. Twenty full-sib clutches were split and reared at four salinities (fresh, 5, 10, and 27 practical salinity units [PSU]). On reaching adulthood, clutches from three of the salinity treatments (no survivors at fresh) were split into low- (fresh) and high- (40 PSU) salinity stress treatments, at which survival was measured for 24 h. Short-term acclimation of adults did not appear to have a long-term affect on low-salinity tolerance, given that gradual transfers to fresh water enhanced survival relative to acute transfers in the short term (after 7 h) but not over a longer period of 8 d. Developmental acclimation had contrasting effects on low- versus high-salinity tolerance. Namely, rearing salinity had a significant effect on tolerance of high-salinity (40 PSU) stress but no significant effect on tolerance of low-salinity (freshwater) stress. In addition, there was a significant effect of clutch on survival under freshwater conditions, indicating a genetic component to low-salinity tolerance but no significant clutch effect in response to high salinity. While developmental acclimation might enhance survival at higher salinities, the minimal effect of acclimation and significant effect of clutch on low-salinity tolerance suggest the importance of natural selection during freshwater invasion events.