Fatigue is a frequent symptom following acquired brain injury. Assessment is usually based on self-report, but additional objective measures are advised. The Psychomotor Vigilance Test has previously been found to be sensitive to the effects of acquired brain injury and performance on this test may relate to fatigue and sleepiness. This study examined whether Psychomotor Vigilance Test performance in people with acquired brain injury is associated with fatigue, after controlling for daytime sleepiness, sleep quality, and mood. The study found that performance on this test is related to fatigue, but also to sleepiness and mood. The Psychomotor Vigilance Test therefore cannot be used as a specific measure for fatigue, but could be used to measure changes or improvement of symptoms including fatigue, mood and sleepiness. The Psychomotor Vigilance Test thus has the potential to be implemented as an objective measure to evaluate these symptoms following acquired brain injury in both research and clinical practice.