Ocular assessment tools provide insights into neurological functioning when diagnosing sport-related concussions (SRCs). Emerging technologies such as EyeGuide Focus have attempted to improve on pre-existing clinical tools however the reliability of the tool is currently uncertain and it is unclear if physical exertion may affect the validity of the device as seen with similar ocular assessments. The aim of this study was to determine if EyeGuide Focus accuracy is affected by high-intensity, intermittent exercise. Seven amateur female rugby union athletes were recruited (Mean age [± SD] = 23 ± 5 years; stature = 172 ± 4 cm, mass = 87 ± 20 kg). Athletes completed baseline testing on the unit which included two to four familiarisation trials and three official tests on the unit measured in arbitrary units (AUs). Testing was completed in a dark room free from distraction, and participants wore noise-cancelling headphones (BOSE QC45) during assessment. The athlete then performed a brief warmup before beginning the exercise protocol by Whyte et al. (2022, Phys Therapy Sport, 58, 126-133) which involves repeated circuits of acceleration, deceleration, backpedalling, change of direction, jumping, and side shuffling. This protocol has previously displayed significant effects on Vestibular Ocular Motor Screening testing. Athletes repeated circuits until they reported 17-18/20 on the Borg Scale with 30s between each circuit. Athletes then completed three more trials at five- and 10-mins post-exercise cessation. Data were exported to Microsoft Excel and Statistical Packages for the Social Sciences for analysis. Athletes completed an average of 03:15 ± 23 s total work in circuits. Significant differences existed between athlete’s heart rate at rest and post-warmup, post individual circuits, and five mins post-test (P < 0.05) but not 10 mins post-test (P > 0.05). Mean EyeGuide Focus scores were 23443.29 ± 8082.18, 35684.04 ± 13279.40, and 22927.64 ± 6377.61 AUs at baseline, 5 mins post-test, and 10 mins post-test respectively. Scores were significantly different between baseline and 5 mins post-test (P < 0.001), and between 5- and 10-mins post-test (P < 0.001), but not baseline and 10 mins post-test (P > 0.848). Three mins of high-intensity, intermittent exercise was sufficient to induce significant changes in amateur female athletes’ EyeGuide Focus scores, returning to baseline after 10 mins of rest. The findings indicate that physical activity can impact on EyeGuide Focus scores and thus sufficient rest is required when assessing athletes for SRCs. Future research should investigate if sport-related contact or collisions also impact scoring on this device.