Optimizing the usability of e-learning materials is necessary to reduce extraneous cognitive load and maximize their potential educational impact. However, this is often neglected, especially when time and other resources are limited. We conducted a randomized trial to investigate whether a usability evaluation of our multimedia e-learning resource, followed by fixing of all problems identified, would translate into improvements in usability parameters and learning by medical residents. Two iterations of our e-learning resource ["version 1" (V1) and "version 2" (V2)] were compared. V1 was the first fully functional version and V2 was the revised version after all identified usability problems were addressed. Residents in internal medicine and anesthesiology were randomly assigned to one of the versions. Usability was evaluated by having participants complete a user satisfaction questionnaire and by recording and analyzing their interactions with the application. The effect on learning was assessed by questions designed to test the retention and transfer of knowledge. Participants reported high levels of satisfaction with both versions, with good ratings on the System Usability Scale and adjective rating scale. In contrast, analysis of video recordings revealed significant differences in the occurrence of serious usability problems between the two versions, in particular in the interactive HandsOn case with its treatment simulation, where there was a median of five serious problem instances (range: 0-50) recorded per participant for V1 and zero instances (range: 0-1) for V2 (P < 0.001). There were no differences in tests of retention or transfer of knowledge between the two versions. In conclusion, usability evaluation followed by a redesign of our e-learning resource resulted in significant improvements in usability. This is likely to translate into improved motivation and willingness to engage with the learning material. In this population of relatively high-knowledge participants, learning scores were similar across the two versions.