The filling materials utilized in root canal therapy have a long-term retention in the oral cavity. Safety and biocompatibility of the filing materials always draw people’s attention. The biocompatibility of these materials has a direct impact on the quality of root canal filling and periapical tissue repair response. Biocompatibility tests, including histology, cytology, molecular genetics, and biological evaluation of different levels, cytotoxicity, and genotoxicity of materials, have been successfully utilized to evaluate the biocompatibility of the materials. This study is a review about the cytotoxicity and genotoxicity of root canal filling materials.