This in vitro study evaluated the seal created by varying depths of mineral trioxide aggregate (MTA) plugs placed in an orthograde fashion in five groups of 10 teeth. One group received a 2 mm thick orthograde apical plug of MTA, the second group a 5 mm apical MTA plug, and the third group a 2 mm apical MTA plug with a second 2 mm increment, 24 h later. The remaining portion of the canal in these groups was left unfilled. Group four received a 2 mm MTA plug that set for 24 h and the canal was then back-filled with gutta-percha and eugenol based sealer. Group five was a positive control without an MTA plug. The apical seal was tested using a bacterial leakage model of Actinomyces viscosus. Results showed a statistically significant difference in only the 5 mm apical plug, which completely prevented bacterial leakage.