Objective: The objective of the study was to compare the diagnostic accuracy of cone beam computed tomography and three intraoral radiographic systems in the detection of in vitro caries lesions. Material and Methods: One hundred teeth (46 molars and 54 premolars) were evaluated, including 176 proximal surfaces and 90 occlusal surfaces, with or without dental caries lesions. Digital images of all teeth were obtained using specific intraoral radiographs, VistaScan DürrDental®phosphor-plate radiography, XIOS XG Sirona® digital sensor radiography, and CBCT I-CAT™. Observers evaluated the images for the detection of caries lesions. The teeth were clinically sectioned and stereomicroscopy served as a validation tool. The relationship of sensitivity and specificity between all systems was determined through the ROC curve using Az values. Results: The values of the area under the curve (Az) selected for the CBCT I-CAT™ system were 0.89 (0.84-0.93), for conventional radiography 0.71 (0.66-0.76), digital sensor radiography 0.74 (0.70-0.78) and digital radiography with phosphor-plates 0.73 (0.69-0.77). Statistically significant differences were found between the CBCT I-CAT™ system and intraoral radiographic systems (p<0.01). The sensitivity and specificity values for the CBCT I-CAT™ were 0.84 and 0.93 respectively. Conclusion: CBCT has a high sensitivity and specificity compared to intraoral radiographic systems for the diagnosis of dental caries lesions in vitro.