The number of ecohydrological studies involving water stable isotope measurements has been increasing steadily due to technological (e.g., field-deployable laser spectroscopy and cheaper instruments) and methodological (i.e., tracer approaches or improvements in root water uptake models) advances in recent years. This enables researchers from a broad scientific background to incorporate water-isotope-based methods into their studies. Several isotope effects are currently not fully understood but might be essential when investigating root water uptake depths of vegetation and separating isotope processes in the soil–vegetation–atmosphere continuum. Different viewpoints exist on (i) extraction methods for soil and plant water and methodological artifacts potentially introduced by them, (ii) the pools of water (mobile vs. immobile) measured with those methods, and (iii) spatial variability and temporal dynamics of the water isotope composition of different compartments in terrestrial ecosystems. In situ methods have been proposed as an innovative and necessary way to address these issues and are required in order to disentangle isotope effects and take them into account when studying root water uptake depths of plants and for studying soil–plant–atmosphere interaction based on water stable isotopes. Herein, we review the current status of in situ measurements of water stable isotopes in soils and plants, point out current issues and highlight the potential for future research. Moreover, we put a strong focus and incorporate practical aspects into this review in order to provide a guideline for researchers with limited previous experience with in situ methods. We also include a section on opportunities for incorporating data obtained with described in situ methods into existing isotope-enabled ecohydrological models and provide examples illustrating potential benefits of doing so. Finally, we propose an integrated methodology for measuring both soil and plant water isotopes in situ when carrying out studies at the soil–vegetation–atmosphere continuum. Several authors have shown that reliable data can be generated in the field using in situ methods for measuring the soil water isotope composition. For transpiration, reliable methods also exist but are not common in ecohydrological field studies due to the required effort. Little attention has been paid to in situ xylem water isotope measurements. Research needs to focus on improving and further developing those methods. There is a need for a consistent and combined (soils and plants) methodology for ecohydrological studies. Such systems should be designed and adapted to the environment to be studied. We further conclude that many studies currently might not rely on in situ methods extensively because of the technical difficulty and existing methodological uncertainties. Future research needs to aim on developing a simplified approach that provides a reasonable trade-off between practicability and precision and accuracy. How to cite. Beyer, M., Kühnhammer, K., and Dubbert, M.: In situ measurements of soil and plant water isotopes: a review of approaches, practical considerations and a vision for the future, Hydrol. Earth Syst. Sci., 24, 4413–4440, https://doi.org/10.5194/hess-24-4413-2020, 2020.