The aim of this study was to evaluate the antibacterial activity of a recently introduced glass ionomer endodontic sealer, Ketac Endo (KE), compared to a commonly used ZOE-based endodontic sealer, Roth's cement (RC). With the use of E. faecalis as a test organism, the agar diffusion test (ADT) and direct contact test (DCT) were performed. For the DCT, the sealers were mixed and placed on the side wall of microtiter plate wells. A 10 microL bacterial suspension was placed on the tested material samples 20 min, 24 h, and 7 days after mixing. Bacteria were allowed to directly contact the sealers for 1 h at 37 degrees C. Fresh media was then added and bacterial growth was measured every 30 min over 15 h. The results showed that in the ADT, freshly mixed KE exhibited a twofold greater inhibition zone than RC (p < 0.05); in the DCT, freshly mixed KE and RC completely inhibited bacterial growth. The 24-h and 7-day samples of KE showed no antibacterial activity, whereas RC continued to exhibit a strong effect with similar samples. It has been concluded that KE possesses a short-acting very potent and diffusable antibacterial activity, whereas RC extends its effect over 7 days after setting.