Objective: The aim of this study was to evaluate whether the use of a gel applied for 1 × 45 minutes would have the same bleaching rate and tooth sensitivity levels when compared with 3 × 15-minute applications. Methods: In-office bleaching was performed in 30 participants with 35% hydrogen peroxide gel. In one group (n=15; 3×15 minutes), the bleaching agent was refreshed every 15 minutes, three times at each bleaching appointment. In the other group (n=15; 1×45 minutes) the gel was left undisturbed on the buccal surfaces of all teeth for 45 minutes at each bleaching appointment. This protocol was repeated after one week. The bleaching evaluation was completed by two blinded, calibrated evaluators who compared the baseline color of the maxillary anterior teeth with a valueoriented shade guide after each period. The patients recorded the tooth sensitivity on a 0-4 scale. The color changes were evaluated by appropriate tests (α=0.05). The percentage of patients with tooth sensitivity and its intensity were also statistically analyzed (α=0.05), respectively. Results: The use of gel for a single 45-minute period (1×45 minutes) decreased the bleaching efficacy (p<0.05) 86.7%, and 100% of patients from the 3 × 15-minute and 1 × 45-minute groups, respectively, experienced tooth sensi- tivity (p=0.22). The intensity of sensitivity was lower for the 3 × 15-minute applications (p=0.04). Conclusions: A 35% hydrogen peroxide gel for in-office bleaching preferably should be applied in three 15-minute applications because 1 × 45 minutes reduces the bleaching speed and slightly increases the intensity of tooth sensitivity. © 2011 Operative Dentistry, Inc.