EstomatoNet was created in the south of Brazil to provides specialist support over a web-based platform to primary care dentists for diagnosis of oral lesions. To evaluate the usability of EstomatoNet and to identify user perceptions regarding their expectations and difficulties with the system; and to compare the perceptions of regular users of the service to those of first-time users. Sixteen dentists were selected for the study: 8 were frequent users of EstomatoNet and 8 were residents who had never used the Platform. To assess usability, participants were required to request telediagnosis support for a fictional case provided by the research team. During the process of uploading the information and sending the request, users were asked to "think out loud," expressing their perceptions. The session was observed by an examiner with remote access to the user's screen (via Skype). After the simulation, users completed the System Usability Scale (SyUS), a validated tool with scores ranging from 0 to 100. The mean SyUS score assigned by frequent users was 84.7±6.6, vs. 82.2±9.3 for residents (satisfactory usability: score above 68). The difference between the groups was not statistically significant (Student t test, P = .55). The residents group took longer (347.1±101.1s) to complete the task than frequent users (252.8±80.3s); however, the difference between the groups was not statistically significant (Student t test, P = .06). In their subjective evaluation, users suggested the inclusion of a field to add further information on outcomes and resolution of the case and changes in the position of the "Send" button to improve workflow. The present results indicate satisfactory usability of EstomatoNet. The Platform seems to meet the needs of users regardless of how experienced they are; nevertheless, a few minor changes in some steps would improve the tool.