This guidebook provides theoretical and practical information for using a variety of isotope tracers for dating old groundwater, i.e. water stored in geological formations for periods ranging from about 1000 to one million years. Theoretical underpinnings of the methods and guidelines for their use in different hydrogeological environments are described. The guidebook also presents a number of case studies providing insight into how various isotopes have been used in aquifers around the world. The methods, findings and conclusions presented in this publication will enable students and practicing groundwater scientists to evaluate the use of isotope dating tools for specific issues related to the assessment and management of groundwater resources. In addition, the guidebook will be of use to the scientific community interested in issues related to radioactive waste disposal in geological repositories.