This in vitro study evaluated the effect of smear layer removal on adhesion to human dentine of Ketac-Endo, Grossman, and Sealer 26 root canal sealers. A total of 60 extracted human maxillary and mandibular molars with their crowns ground flat were used. The teeth were divided into two groups: Group 1, the dentine surface received no treatment; Group 2, EDTA-C was applied to the dentine surface for 5 min. Ten samples were tested for each sealer and each group. Adhesion was measured with a Universal Testing Machine. Sealer 26 showed greater adhesion both with and without smear layer (p < 0.01). Ketac-Endo and Grossman sealers were statistically equal, having the lowest values of adhesion. Application of EDTA-C did not alter the adhesion of Ketac-Endo and Grossman sealers. However, the use of EDTA-C increased the adhesion of Sealer 26. The epoxy resin-based root canal sealer (Sealer 26) adhered better to dentine prepared with and without EDTA-C than glass-ionomer-based sealers and the zinc oxide eugenol-based (Ketac-Endo and Grossman) sealers.