The main cause of endodontic disease is bacteria. Disinfection is presently achieved by cleaning the root canal system prior to obturation. Following setting, root canal filling is devoid of any antibacterial effect. Endodontic sealers with antimicrobial properties yet biocompatible may enhance root canal therapy. For this purpose, quaternized polyethylenimine nanoparticles which are antibacterial polymers, biocompatible, nonvolatile, and stable may be used. The aim of the present study was to examine the impact of added QPEI on the cytotoxicity of AH Plus, Epiphany, and GuttaFlow endodontic sealers. The effect of these sealers on the proliferation of RAW 264.7 macrophage and L-929 fibroblast cell lines and on the production of TNF from macrophages was examined. Cell vitality was evaluated using a colorimetric XTT assay. The presence of cytokines was determined by two-site ELISA. Results show that QPEI at 1 concentration does not impair the basic properties of the examined sealers in both macrophages and fibroblast cell lines. Incorporation of 1 QPEI into the sealers did not impair their biocompatibility. QPEI is a potential clinical candidate to improve antibacterial activity of sealers without increasing cytotoxicity.