Erythrocyte osmotic fragility (EOF) is a measure of erythrocyte strength and its ability to withstand varying osmotic gradients. Erythrocyte osmotic fragility was compared among a variety of terrestrial ectotherms, aquatic/semiaquatic ectotherms and endotherms. We hypothesized that EOF should be lowest in those animals that are aquatic and whose red blood cells would likely be exposed to hypotonic conditions (i.e., some amphibians and reptiles). The greatest erythrocyte osmotic resistance (i.e., least fragile erythrocytes) was found in the bullfrog (Rana catesbeiana), followed in decreasing order by the larval tiger salamander (Ambystoma tigrinum), marine toad (Bufo marinus), ornate box turtle (Terrepene ornata ornata), painted turtle (Chrysemys picta bellii), red-eared slider (Trachemys scripta elegans), red-sided garter snake (Thamnophis sirtalis parietalis), domestic chicken (Gallus gallus), cotton rat (Sigmodon hispidus), Sprague-Dawley rat (Rattus norvegicus), and eastern woodrat (Neotoma floridana). A significantly higher erythrocyte osmotic resistance was detected in the aquatic/semiaquatic amphibians (R. catesbeiana and A. tigrinum) than in the terrestrial amphibian (B. marinus). However, among reptilian species, no significant differences were noted between terrestrial and aquatic/semiaquatic species. The erythrocytes of ectotherms possessed a greater osmotic resistance than those of endotherms. Such differences might be necessary because of exposure to wide temperature ranges in ectotherms and/or the long erythrocyte life span of ectothermic red blood cells relative to those of endotherms. Nucleated erythrocytes were more osmotically resistant than were non-nucleated erythrocytes. Additionally, erythrocyte osmotic resistance was correlated with erythrocyte mean cell volume. Larger erythrocytes were more osmotically resistant than smaller erythrocytes. Polynomial regression equations computed to approximate these data showed the cubic polynomial to be an excellent fit when data for the larval tiger salamander were excluded. Our data suggest that erythrocyte volume may play a significant role in erythrocyte osmotic resistance.