The aim of the present in vitro study was to determine the penetration depth of dentine bonding agents into carious dentine as a possible strategy in caries treatment of deep lesions. Forty-eight extracted carious teeth were used for the experiments and divided in four groups. In a split-tooth design, one half of each tooth was treated without caries excavation, and the second half after caries excavation. In group 1 the teeth were treated with Prime & Bond 2.0 without phosphoric acid etching, and in group 2 with phosphoric acid etching. In group 3, the teeth were treated with All-Bond 2 without phosphoric acid etching, and in group 4 with phosphoric acid etching. Bonding agents were marked with an orange dye prior to application. The composition of the bonding agents had no significant influence on penetration depths. The mean thickness of the outer, necrotic caries zone in unexcavated specimens was 117 microm (+/-126). Bonding agents penetrated significantly deeper into unexcavated carious dentine (mean 124 +/- 76 microm) compared with excavated dentine (11 +/- 14 microm) and sound dentine (3 +/- 4 microm). Acid etching significantly reduced the thickness of the outer caries zone, and the penetration depth of the bonding agents into unexcavated carious specimens by approximately 50%, and increased it into sound dentine. The results indicate that dentine bonding agents could completely penetrate the unexcavated outer carious dentine in 90% of specimens. Further studies are required to determine whether further caries activity is arrested after treatment with dentine bonding procedures.