Anxiety relating to a multitude of ecological crises, or eco-anxiety, is a subject of growing research significance. We used a multi-study mixed-methods design to explore eco-anxiety in Australia and New Zealand, validating a new eco-anxiety scale. In Study One, we developed and tested a 7-item eco-anxiety scale (n = 334), finding that this captured some, but not all, experiences of eco-anxiety. We found that people were anxious about a range of environmental conditions and their personal negative impact on the planet. Notably, people’s anxiety about different environmental conditions (e.g., climate change, environmental degradation, pollution) were inter-connected, lending support for the existence of eco-anxiety (a broader construct that encompasses climate change anxiety). These results informed further scale development in Study Two. Exploratory (n = 365) and Confirmatory Factor Analysis (n = 370) supported a final 13-item scale that captured four dimensions of eco-anxiety: affective symptoms, rumination, behavioural symptoms, and anxiety about one’s negative impact on the planet, which were each distinct from stress, anxiety and depression. A further longitudinal sample (n = 189) established the stability of these factors across time. Findings support eco-anxiety as a quantifiable psychological experience, reliably measured using our 13-item eco-anxiety scale, and differentiated from mental health outcomes.