The aim of this validation study was to comprehensively evaluate the performance and generalization capability of a deep learning-based periapical lesion detection algorithm on a clinically representative cone-beam computed tomography (CBCT) dataset and test for non-inferiority. The evaluation involved 195 CBCT images of adult upper and lower jaws, where sensitivity and specificity metrics were calculated for all teeth, stratified by jaw, and stratified by tooth type. Furthermore, each lesion was assigned a periapical index score based on its size to enable a score-based evaluation. Non-inferiority tests were conducted with proportions of 90% for sensitivity and 82% for specificity. The algorithm achieved an overall sensitivity of 86.7% and a specificity of 84.3%. The non-inferiority test indicated the rejection of the null hypothesis for specificity but not for sensitivity. However, when excluding lesions with a periapical index score of one (i.e., very small lesions), the sensitivity improved to 90.4%. Despite the challenges posed by the dataset, the algorithm demonstrated promising results. Nevertheless, further improvements are needed to enhance the algorithm’s robustness, particularly in detecting very small lesions and the handling of artifacts and outliers commonly encountered in real-world clinical scenarios.