In today’s world shaped by technology and the natural sciences, knowledge and skills related to experimentation are crucial, especially given growing public debates about science-related topics. Despite a strong emphasis on experimentation in science curricula worldwide, learners still encounter diverse challenges when experimenting. This paper provides a systematic review of empirical research on learners’ challenges during the following inquiry phases: stating research questions, generating hypotheses, planning and conducting an experiment, analysing data and drawing conclusions. A database search and an analysis of two prior narrative literature reviews identified 368 studies, of which 66 were used for further analyses after screening for eligibility using specific inclusion criteria. The analyses revealed 43 challenges at the conceptual, procedural and epistemic level that not only elementary but even university students face during experimentation. Additionally, cognitive biases and preconceptions are identified as a source of such challenges. Overall, the findings demonstrate a lack of in-depth research on stating research questions despite its importance for experimentation, whilst learners’ abilities in the other inquiry phases have been intensively investigated. The results offer valuable information for science education research and provide a basis for tailored scaffolding in the science classroom or the design of effective instructional interventions.