A smear layer is created during the cleaning and shaping of root canal systems. The Nd:YAG laser has been shown to be effective in removing that smear layer and any tissue remnants from prepared root canal systems suggesting that it may aid in root canal sterilization without detrimental thermal effects to adjacent tissues. The root canal system of 72 single-rooted teeth was conventionally prepared and sterilized using gamma radiation. The teeth were divided into three groups of 24 each, 8 of which were inoculated only with sterile broth and remained as negative controls. Sixteen teeth of each group were inoculated with one of three organisms of 106 to 1010 CFU/(mu) l: B subtilis (BS), E. coli (EC) and S. marcescens (SM) (10 (mu) l). Eight in each group were not treated further and served as positive controls. Sixteen test teeth were treated with the laser three times using each exposure parameter: 1 W, 10 Hz pulses per second (pps); 2 W, 20 Hz; and 3 W, 30 Hz inserted to the radiographic apex. Laser exposures were completed while withdrawing the fiber from the root canal system. At completion of laser exposure, all teeth were cultured, using sterile paper points and plated on brain heat infusion agar. Three cultures were taken for each tooth, the plates incubated for 72 hours, and read for the presence of growth of colony-forming units. The laser was able to reduce the number of organisms placed in root canal systems, and suggests that the laser may be used in root canal therapy for bacterial reduction and cleaning of the root canal space.