Purpose: The objectives of this study were (1) to compare the hybridization effectiveness of two adhesive systems that are applied in respectively three and two steps, and {2} to determine the best resin-dentin interface preparation technique for atomic force microscopy (AFM). Materials and Methods: The resin-dentin interface produced by the three-step OptiBond Dual-Cure (Kerr} and its simplified two-step successor OptiBond Solo (Kerr)was ultramorphologically examined using transmission electron microscopy (TEM) and AFM. Four different methods were used to prepare interface specimens for AFM: (1) polishing to a 0.1pm finish with a silicon oxide suspension, (2) polishing to a 0.05-pm finish with an aluminum oxide suspension, (3) argon-ion etching, and (4) sectioning with a diamond knife. Results: Both TEM and AFM demonstrated that some collapse of the exposed collagen fibril network, due to gentle postconditioning air-drying of the dentin surface, may not have been totally recovered through hybridization by the two-step adhesive formulation as opposed to the three-step precursor. From the four interface preparation methods, only diamond-knife sectioning revealed sufficient ultramorphologic detail and high resolution that can capitalize on the high resolution offered by AFM. Conclusion: First, the findings suggest that simplifying the application procedure of adhesives by combining the primer and adhesive resin into a single application step may reduce hybridization effectiveness. Future research should confirm this effect for other two- versus three-step adhesive systems. Second, diamond-knife sectioning should be used for future topographic imaging and physicomechanical testing of resin-dentin interfaces by AFM.