The aim of this in vitro study was to present a method using confocal laser scanning microscopy for three-dimensional analysis of human dental enamel subjected to ceramic bracket debonding. The labial enamel surfaces of three upper central incisors were prepared and mounted in the form of standardized specimens. A sample repositioning protocol was established to enable surface measurement and analysis before and after bracket debonding. Observations were made of representative areas measuring 1,280 × 1,280 μm2, in the center of the enamel samples, as well as of the total topography (2,500 × 3,500 μm) of the bonding areas provided by the equipment software. Noncontact three-dimensional high-resolution image analyses revealed the capabilities of the employed technique and methodology to permit the examination of specific characteristics and alterations on the surfaces, before and after the debonding and finishing procedures. The new protocol was effective to provide qualitative and quantitative assessments of changes on the same dental surfaces at different trial times. The methodology constitutes a feasible tool for revealing the effects of debonding of ceramic brackets on sound and previously injured dental enamel surfaces.