Dimethyl sulfoxide (DMSO) is commonly used as a solvent for organic compounds but its toxic properties can affect both in vitro and in vivo studies. Thus, evaluation of DMSO toxicity must be performed on the model organism before its application as a solvent in the target studies. Present study aimed to determine the lethal concentration required to kill 50% of the population in a given period of time (LC50), no observed effect concentration (NOEC), and lowest observed effect concentration (LOEC) values for DMSO on Drosophila melanogaster which is frequently used as a model for toxicity studies. Twelve different concentrations of DMSO were tested on three-day old D. melanogaster larvae for 12 days. At the end of the life cycle, number of live hatched adults and un-hatched pupae were counted. Based on probit analysis 12 days LOEC was 0.04% v/v, 12 days NOEC was ˂ 0.04% v/v and 12 days LC50 was 0.42% v/v. The results indicate that DMSO may be more toxic to D. melanogaster than it was initially considered.