Level crossing safety is a well-researched safety issue worldwide, but little attention has been placed on the safety benefits of using train horns when a train approaches a level crossing. Given train horns' adverse effects on the health and well-being of residents living near rail tracks, the use of train horns must be beneficial to safety. The current study sought to determine in a laboratory environment whether road users (N = 31) can detect the range of train horns observed in Australia in terms of loudness and duration, using high-definition audio recordings from railway crossings. A repeated measures design was used to evaluate the effects of key factors likely to influence the detectability of train horns, including, visual and auditory distractive tasks, hearing loss and environmental noise (crossing bells). Train horn detectability was assessed based on participants' accuracy and reaction times. Results indicated the duration of the train horn had the most influential effect on the detectability of train horns, with short-duration train horns less likely to be detected. The presence of bells at a crossing was the second most important factor that limited train horn detection. Train horn loudness also affected detectability: faint blasts were less likely to be noticed, while loudest blasts were more likely to be noticed. However, loud horns reduced the ability to detect the side from which the train was approaching and may result in longer times to detect the train, in the field. The auditory distractive task reduced the train horn detection accuracy and increased reaction time. However, the visual distractive task and medium to severe hearing loss were not found to affect train horn detection. This laboratory study is the first to provide a broad understanding of the factors that affect the detectability of Australian train horns by road users. The findings from this study provide important insights into ways to reduce the use and modify the practice to mitigate the negative effects of train horns while maintaining the safety of road users.