Background: Despite its pivotal role, the use of bone age is limited due to a lack of physician expertise, time, and access. Objective: To develop and validate a mobile application-based bone age assessment tool. Study Design: The study involved the selection of standardized images (from 307 radiographs), delineation (90 radiographs), evaluation (200 radiographs) and incorporation of regions of interest on the application, and validation against the Tanner-Whitehouse 3 method by an expert (252 radiographs) and non-expert users (110 radiographs). Results: The application-based assessment by expert users had an absolute standardized difference of 4.7 months (95% confidence interval; 4.2-5.2 months), a relative standardized difference deviation of 4.4% (3.9-4.9%), and similar intraindividual [2.8 (1.9-3.7) months versus 3.6 (2.8-4.5) months] and interindividual variation [4.2 (3.3-5.0) versus 4.2 (3.3-5.2) months] compared to the TW3 method. Non-expert user assessment had an absolute standardized difference of 6.7 months (5.6-7.7 months) and a relative standardized difference deviation of 5.8% (4.7-6.9%). The mean test time was lower for application than TW3 for both expert [1.3 (0.2) versus 3.1 (1.0) minutes, p<0.001] and non-expert users [2.8 (1.0) versus 5.0 (1.4) minutes, p<0.001]. Discussion: Our study confirms the accuracy of mobile application-based bone age assessment. This, along with good precision, reduced complexity, and lower time requirement, suggests a potential for its widespread implementation.