Teachers are ideally placed to identify and refer pupils who self-injure, but are often unaware when pupils self-injure or unsure how to respond. The aims of this study were to explore and compare pre-service and in-service teachers’ knowledge and attitudes towards self-injury, and their confidence responding to pupils who self-injure. Pre-service teachers (n = 267) and in-service teachers (n = 261) completed self-report questionnaires. Prior education regarding self-injury was positively related to knowledge and confidence, while pre-service teachers were more confident than in-service teachers in their ability to cope with legal and school regulations. Thematic analysis of open-ended questions indicated that although pre- and in-service teachers are concerned about pupils who self-injure and are willing to help these students, they feel ill-informed about self-injury and requested school policies and additional education regarding the behaviour. Results have implications for educational programmes that prepare pre- and in-service teachers to identify and respond to pupils who self-injure.