Scaling and root planing are one of the most commonly used procedures for the treatment of periodontal diseases. Removal of calculus using conventional hand instruments is incomplete and rather time-consuming. In search for more efficient and less difficult instrumentation, investigators have proposed lasers as alternatives or adjuncts for scaling and root planing. The aim of the present study was to compare the effectiveness of subgingival scaling and root planing with erbium: yttrium, aluminium, garnet (Er:YAG) laser and hand instrumentation in vitro. The mesial and distal surfaces of 15 periodontal loosed extracted teeth were treated randomly either by hand instrumentation or by Er:YAG laser irradiation. After choosing the "very long pulse mode" (pulse duration of about 700 micros), the output energy of 160 mJ with 920-microm beam diameter (RO7 Perio tip, Fidelis, Fotona, Slovenia) and frequency of 12 Hz were selected, both according to the best results of past studies. In addition, air water spray was used during the procedures. The morphology of the root surface was evaluated by three observers with a scanning electron microscopy in magnifications of 50x and 400x. The result of this setting showed that the rate of remained roughness on treated root surfaces in two groups of hand instruments and Er:YAG laser had a meaningful difference: The surface roughness in Er:YAG laser group was more than in hand instruments group. The present study could demonstrate the in vitro capability of the Er:YAG laser for scaling and root planing in periodontitis, although the effectiveness of this setting did not reach that achieved by hand instrumentation. It could be concluded that lower frequency and long pulse duration maybe more suitable for the micro-morphology of root surface after treatment. This theory is going to be tested with the same laser instrument in the next study.