Purpose This study evaluated the effect of different surface treatments on tensile bond strength of acrylic denture teeth to prepolymerized computer-aided design/computer-aided manufacturing (CAD/CAM) denture base resin (PMMA) under two different storage conditions. Patients and methods 160 test specimens were used in this study, each specimen has two parts: the first part: is an acrylic cross-linked maxillary central incisor denture tooth embedded in acrylic resin blocks, and the second part: is cone shape (5 mm × 5 mm × 3 mm) specimens prepared from prepolymerized PMMA discs by computer numerically controlled (CNC) milling machine under water cooling. Test specimens were divided into four main groups 40 each according to tooth surface treatment. Group I: no sandblasting. Group II: sandblasted using 50 μm aluminum oxide. Group III: sandblasted using 150 μm Al2O3. Group IV: sandblasted using 200 μm Al2O3. Each group was divided into 2 subgroups (A, B) 20 each, subgroup A used resin cement bond first part with the second part. Subgroup B used a bonding agent to bond the first part with the second part. Each subgroup was divided into 10 specimens that were stored in 50 hours of distilled water storage and 10 specimens were thermocycling. All specimens were then subjected to a tensile load using testing machine at a crosshead speed of 1 mm/min until debonding. The resulting debonding forces were recorded and statistically analyzed. For all specimens, the interface was inspected. The interface failures were classified into adhesive and cohesive failures. The data were analyzed using a three-way Analysis of Variance (ANOVA), followed by Tukey’s test. Results The highest mean value tensile bond strength was recorded for GIV (6.5 ± 1.46). Sandblasting was improving the tensile bond strength value of all the specimens. Bonding agents without and with surface treatment significantly improved the tensile bond strength of all specimens. The thermocycling significantly decreased the tensile bond strength of all specimens. Conclusion Sandblasting of denture teeth followed by bonding agent improved the tensile bond strength to CAD/CAM denture base. Thermocycling decreased the tensile bond strength of acrylic denture teeth to CAD/CAM denture base.