In the last 20 years, humane education (HE) has undergone a paradigm shift from a simple animal welfare education model to a teacher-led positive education approach that cultivates kindness, love, and empathy in children and youth. While teachers’ humane attitudes are considered a prerequisite for HE program success in school settings, their attitudes toward animals have received little attention to date. This study investigated pre-service teachers’ humane attitudes toward animals and assessed whether and to what extent these attitudes are correlated with empathetic tendencies as well as variables such as gender and current and previous pet ownership. A total of 226 pre-service teachers in Hong Kong completed the Animal Attitude Scale (AAS) and the Davis Interpersonal Reactivity Index (IRI). The results showed that pre-service teachers showed moderate concern for the humane treatment of animals, and a difference in humane attitude was identified based on sex: female pre-service teachers tended to express more humane attitudes than their male counterparts. Moreover, pre-service teachers with greater tendencies toward warmth and compassion demonstrated a more welfare-oriented attitude toward animals. The results of the current study support using HE as a positive education approach and imply that the provision of training for pre-service teachers and teachers should be given higher priority by humane educators. Humane educators are recommended to recruit pre-service teachers as HE program instructors in their research projects. Copyright © 2020 ISAZ.