The presence of glycosaminoglycans (GAGs) and their contribution to mechanical properties of the cementum-dentin junction (CDJ) were investigated using nanometer scale characterization techniques. Five to two millimeter thick transverse sections from the apical ends of human molars were ultrasectioned at room temperature under wet conditions using a diamond knife and an ultramicrotome. The structure of the CDJ under dry and wet conditions before and after digestion of GAGs and collagen fibrils was studied using an atomic force microscope (AFM). The mechanical properties of the untreated and enzyme treated CDJ under wet conditions were studied using an AFM-based nanoindenter. GAG digestion was performed for 1, 3, and 5 h at 37 degrees C using chondroitinase-ABC. Collagen fibril digestion was performed for 24 and 48 h at 37 degrees C using collagenase. As reported previously, AFM scans of dry untreated CDJ (control) revealed a valley, which transformed into a peak under wet conditions. The height differences relative to cementum and dentin of untreated and treated CDJ were determined by measuring the CDJ profile under dry and wet conditions. The depth of the valley of GAG and collagen-digested CDJ was greater than that of undigested CDJ under dry conditions. The height of the peak of GAG-digested CDJ was significantly higher than that of the undigested CDJ under wet conditions. The collagen-digested CDJ under wet conditions is assumed to form a valley because of the removal of collagen fibrils from the CDJ. However, the depth of the valley was lower compared to the depth under dry conditions. Wet AFM-based nanoindentation showed that the elastic modulus and hardness of control (3.3 +/- 12 and 0.08 +/- 0.03 GPa) were significantly higher (ANOVA & SNK, P < 0.05) than chondroitinase-ABC treated CDJ (0.9 +/- 0.4 and 0.02 +/- 0.004 GPa) and collagenase treated CDJ (1.5 +/- 0.6 and 0.04 +/- 0.01 GPa). No significant difference in mechanical properties between chondroitinase-ABC and collagenase treated CDJ was observed. Based on the results it was concluded that the 10 50 pm wide CDJ is a composite that includes, chondroitin-4-sulfate, chondroitin-6-sulfate. and possibly dermatan sulfate, and collagen fibrils. The association of GAGs with the collagen fibrils provides the observed controlled hydration and partially contributes toward the stiffness of the CDJ under wet conditions. (C) 2005 Elsevier Inc. All rights reserved.