The aim of this clinical study was to compare the treatment outcome following root surface conditioning using an EDTA gel preparation in conjunction with surgical therapy with that following conventional flap surgery in periodontal intraosseous defects. 36 patients, each of them contributing one intraosseous defect ≥4 mm in depth participated. Defect sites had a probing pocket depth ≥5 mm and bled on probing following hygienic treatment phase. No furcation involvement or endodontic complications were present. In the EDTA group, 18 consecutive patients, defects were treated by root conditioning with EDTA gel for 3 minutes in combination with surgical therapy. In the control group, 18 patients, conventional flap surgery was performed without root conditioning. Chlorhexidine rinsings 0.2% were prescribed following surgery for 2-3 weeks with modified oral hygiene instruction. A strict recall program was implemented including professional prophylaxis and oral hygiene reinforcement every 4-6 weeks until 6-month re-evaluation. Baseline probing pocket depths and defect depths of 7.1±1.3 mm and 6.9±1.6 mm in the EDTA group and 7.6±1.9 mm and 6.6±1.7 mm, respectively, in the control group were measured. 6-month clinical results showed a significant probing attachment level gain of 1.8±1.5 mm and 1.0±1.7 mm in the EDTA and control groups respectively. A probing bone gain of 1.0±1.3 mm in the EDTA group was measured with a non-significant gain of 0.4±1.2 mm in the control group. Radiographic analysis confirmed these results. There were no statistically significant differences in treatment outcome between the group treated by root conditioning in combination with flap surgery and conventional flap surgery alone.