Clinical studies have shown a connection between the reduction of micro-organisms in the root canal and the degree of apical enlargement. The aim of this study was to create a non-invasive 3D model for monitoring the apical enlargement in molar teeth before and after mechanical instrumentation. Extracted molar teeth were embedded in resin and kept moisturized. Each root canal was instrumented with NiTi instruments to #40. The teeth were scanned before and after instrumentation, using a microCT40 micro-tomography (SCANCO Medical AG, Bassersdorf, Switzerland). The reconstructed slices were imported into Analyze7.0. In-house software, Landmarker, was applied to calculate the distance between two equal points on the superimposed 3D models. Fig. 1 exemplifies the results of the reconstructed root canals before and after instrumentation. In Fig. 2 the colour scale indicates increasing dentin removal during instrumentation. In conclusion, the method is useful for quantitative analysis of apical enlargement during root canal instrumentation.