Introduction: Mycotoxins are among the environmental chemical agents with toxic effects on animals and humans. Aflatoxins (AF) are a major group of mycotoxins produced following the growth of some species of Aspergillus flavus and Aspergillus parasiticus, particularly in agricultural products. Aflatoxin M1 (AFM1) is secreted to milk and remains there since the process of pasteurization and sterilization has little effect on this metabolite. The aim of this study was to investigate the natural occurrence of AFM1 in raw milk in Behbahan area, which is an area of highdensity livestock production in southwest of Iran. The other aim was to compare these obtained levels with the accepted level recommended by the European community (50 ng/L). Methods: In the present study, 90 samples of raw milk were collected from five dairy farms in Behbahan region, Khuzestan province, Iran. The AFM1 was analyzed using the Enzyme Linked Immune Sorbent Assay (ELISA) method by ELISA kits from Euro Clone, Italy. Data were analyzed by ANOVA and the results were reported as mean ± SE. The significant level was set at p ≤ 0. 05. Results: The concentration of aflatoxin M1 in (15)16. 54% of raw milk was higher than maximum tolerance limit accepted by European Union /Codex Alimentarius Commission (50 ng/l). The results of this study showed that all 90 investigated raw milk samples were contaminated with AFM1 (100%). Conclusion: A significant difference was observed between concentration means of AFM1 in various studied farms at P < 0. 05. Consequently, high occurrence of AFM1 in raw milk samples was considered as a possible hazard for human health. However, further longitudinal studies are required on milk samples and other dairy products in Behbahan as well as other regions of Iran to estimate the representative intake of AFM1 in Iran.