Background : Propolis is a dark and sticky substance produced by bees. Propolis in the world of health was known to have benefits as anti-inflammatory, antioxidant, antibacterial, antiviral, antifungal, and antitumor. However, propolis has the potential of being toxic if used as herbal medicine. Purpose : This study aim is to tests the effect of cytotoxic ethanol extract of propolis (EEP) on viability of cultured fibroblast cells.. Method : The design of this study was pure laboratory experimental in vitro. Propolis used comes from Apis trigona bees and extracted using 40% ethanol. EEP is then used as a treatment on Human Dermal Fibroblast adult (HDFa) cells. The cytotoxic effects of EEP were analyzed based on viability of fibroblast cells calculated by the method of MTT assay. Results : EEP with a concentration of 500 μg / mL had the highest cytotoxic effect because it resulted in a culture of fibroblasts having viability of 8.44%. EEP-treated fibroblast cells show a morphological change observed under a microscope. Morphological changes show few cells become smaller and rounder. Statistical analysis was done by using Spearman correlation test. The coefficient of correlation shows (r = 0,839) which means there is a strong and opposite relationship between giving EEP various concentrations and viability of fibroblast cells. Conclusion : EEP showed the presence of dose-dependent cytotoxicity on fibroblasts' viability.