The application of three‐dimensional (3D) visualization technology has generated interest due to its potential to augment or even replace cadaver use in anatomical education. By adopting stereoscopic 3D digital technologies, educational institutions may be able to mitigate issues related to the resource intensiveness of cadaver use such as prosection availability, body donation and the infrastructure to maintain physical specimens. We previously developed a smartphone application, titled VRBR, which uses an inexpensive Google Cardboard headset to display 2D or stereoscopic 3D images of cadaveric and plastinated specimens and plastic models for learning and testing anatomical knowledge. The purpose of this study was to compare the effectiveness and validity of stereoscopic 3D images, 2D images and cadaveric specimens in testing anatomical knowledge with practical examinations (OSPEs) for undergraduate students with prior anatomy education. As cadaveric specimens and stereoscopic 3D images are both stereoscopic, we hypothesized that participants would perform similarly between those modalities, and more poorly in 2D. Students who had previously completed an undergraduate anatomy and physiology course (N = 60) were randomized to one of three testing groups (A, B or C). Each testing group was administered OSPEs in three distinct modalities: VRBR 2D images, VRBR stereoscopic 3D images or cadaveric specimens. In total, each participant answered 45 anatomy‐based questions (15 per modality, test order randomized across groups) and completed a questionnaire assessing cybersickness and user satisfaction. Participants completed a stereo fly test and mental rotation test to assess their stereoacuity and visuospatial ability, respectively. These are potential covariates of the testing outcome when assessing spatially complex objects of varying depth. This study was approved by the Hamilton Integrated Research Ethics Board. In regard to cybersickness, a small fraction reported nausea (7.7%), vertigo (7.7%), and dizziness (15.4%) while responses for headache (23.1%) and fatigue (23.1%) were moderate and general discomfort (61.5%) and eyestrain (76.9%) were more prevalent. Preliminary data (N = 13) assessed via two‐way ANOVA, suggested that there were no statistically significant effects of test version (F(1, 22) = 0.005, p = 0.945) or stereoscopy (F(1, 22) = 0.009, p = 0.728). If this trend persisted through completion of the study, it may suggest that stereoscopy, as provided by the headset, does not improve the effectiveness of digital tools for testing anatomy knowledge recall. Complete data collection and analysis in February 2020 will provide a more complete picture of the validity of stereoscopy in testing anatomy knowledge recall. With ongoing analysis, the results from this study may elucidate the effect of stereopsis in testing anatomical knowledge and guide the future development of undergraduate anatomy education curricula.Support or Funding InformationFunded by the Education Program in Anatomy.