Reaction time (RT) is a fundamental cognitive function impacting daily life and sports activities. Despite the increasing use of virtual reality (VR) for cognitive assessments, its efficacy for RT assessments remains uncertain. We aimed to develop and validate a novel RT test in VR, and to explore the potential contributions of VR technology to RT assessment. Forty-eight participants completed a computerized RT test (COM-RT) and a novel RT test in VR (VR-RT). The latter replicated the COM-RT test, assessing simple and choice RTs. Additionally, it introduced more complex conditions: reaching to touch static stimuli in known or unknown locations or moving stimuli. Correlations and differences between the tests and tasks were examined. Moderate-to-strong linear correlations were found between the tests for the simple (r ≥ 0.642) and choice (r ≥ 0.736) tasks, and the difference between them (r ≥ 0.677) (p < 0.001). However, RTs were significantly longer in the VR-RT test compared with the COM-RT test (p < 0.001). Significant differences were found among the RT-VR tasks (p < 0.001): reaching to touch stimuli involved longer RTs compared with pressing a button, and RT was even longer when stimuli appeared in unexpected locations. Moving stimuli were associated with shorter RT. Additionally, movement velocity was significantly higher when reaching for dynamic versus static stimuli in known or unknown locations (p < 0.001). VR is valid for RT measurement, yet its outcomes should be interpreted within its framework rather than in comparison to computer assessments. Furthermore, VR offers additional possibilities, such as using lifelike stimuli and measuring kinematics.