Spatial planning is concerned with spatial impacts of decision-making in sector policies and actions in public and private sectors. It aims to consider and coordinate these impacts in a way that leads to sustainable spatial development. Up to now, the consideration of biodiversity and ecosystem services in spatial planning has been based on environmental objectives and targets as derived from international, European Union, national or subnational government directives and legislation as democratically legitimized aims of land development. Biodiversity and ecosystem service impact assessment of spatial planning includes all the basic elements of an ordinary impact assessment: ecological, social, cultural and economic impacts from the point of view of biodiversity and ecosystem services. Tools for assessing impacts of different spatial planning alternatives on biodiversity and ecosystem services are needed to find the most optimal solutions during the planning process. Comprehension of best methodologies in assessing the impacts is constantly evolving, and there is already much experience of feasible methods. Because spatial planning is about allocation of space to different functions, spatially explicit analysis and impact visualization methods are often found to be the most useful. In addition, collaboration and public participation are seen as elementary for spatial planning, and GIS-based methods have been developed for that purpose too. This chapter presents key issues related to including biodiversity and ecosystem services in impact assessment of spatial planning, introduces some useful tools for assessing these impacts, and showcases three real-life spatial planning processes where impacts have been assessed using various tools. Finally, a checklist for implementing impact assessment in practice is provided.