This in vitro study aimed to evaluate the effect of two alternative clinical protocols on the tooth bleaching effectiveness of 37% carbamide peroxide. Thirty enamel-dentin blocks from bovine incisors were stained with a coffee solution for 1 week. After color measurement at baseline with a portable spectrophotometer, the blocks were bleached during two 45-min applications of 37% carbamide peroxide with a 2-day interval between bleaching procedures, according to one of the following protocols: 1) Control: placement of bleaching agent over dried enamel; 2) Moist enamel: placement of bleaching agent over enamel previously humidified with a damp gauze for 1 min; 3) Moist cotton pellet: covering the bleaching agent with a moist cotton pellet during the entire bleaching procedure. The tooth color was assessed 2 days after each bleaching procedure. The data of ΔL, Δa, Δb, and ΔE were individually analyzed using ANOVA and Tukey's test (α = 0.05). In general, applying the bleaching agent to the moist enamel resulted in lower Δa and Δb values, and higher ΔL and ΔE values than the control. No difference was observed between the control and the protocol of covering the bleaching agent with a moist cotton pellet. Regardless of the protocol, the second bleaching procedure resulted in a further increase in ΔL and ΔE, and a reduction in Δa. A further decrease in Δb was observed only for the moist enamel protocol. In conclusion, humidifying the enamel with a damp gauze prior to the placement of 37% carbamide peroxide improved the effectiveness of the bleaching.