The aim of the present study was to investigate laser-fluorescence effects on the root surface. The possibility of detecting subgingival calculus should be evaluated to control complete calculus removal after scaling and root planing. Twenty freshly extracted human teeth, partially covered with calculus on the root surface, were fixed on a translation stage. The specimens were irradiated by light from a laser source (diode laser, <1 mW) with a wavelength of 655 nm using the Diagnodent trade mark -system. To simulate the conditions in a periodontal pocket during the measurement procedure, the teeth were covered by a layer of physiological saline solution or blood in order to determine the influence of different inflammatory fluids. The root surface was scanned on a line of about 1 cm in steps of 0.5 mm. The start and end was marked by a notch. The results of the laser-fluorescence detection were compared to clinical and histological findings. Clinically apparent calculus on the root surface was always accompanied with a statistically significant increase of laser-fluorescence values ( p < 0.05). These values were not influenced by the different fluids ( p < 0.05). The laser-fluorescence values were reproducible ( p > 0.05). After scaling, when no debris remained at the root surface, laser-fluorescence values decreased significantly ( p < 0.05). The histological findings supported the clinical results. In conclusion, the present study indicates that laser-fluorescence values on the root surface are strongly correlated to the presence of calculus. This fluorescence effect can basically be used to develop a new detecting system for subgingival calculus in periodontal pockets.