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Oxygen diffusion limitation and metabolic oxygen consumption by embedded cells produce areas of hypoxia in the tissue\/matrix. However, reliable systems to detect oxygen gradients and cellular response to hypoxia in 3D cell culture systems are still missing. In this study, we developed a system for visualization of oxygen gradients in 3D using human adipose tissue\u2013derived mesenchymal stem cells (hAD\u2010MSCs) modified to stably express a fluorescent genetically engineered hypoxia sensor HRE\u2010dUnaG. Modified cells retained their stem cell characteristics in terms of proliferation and differentiation capacity. The hypoxia\u2010reporter cells were evaluated by fluorescence microscopy and flow cytometry under variable oxygen levels (2.5%, 5%, and 7.5% O<jats:sub>2<\/jats:sub>). We demonstrated that reporter hAD\u2010MSCs output is sensitive to different oxygen levels and displays fast decay kinetics after reoxygenation. Additionally, the reporter cells were encapsulated in bulk hydrogels with a variable cell number, to investigate the sensor response in model 3D cell culture applications. The use of hypoxia\u2010reporting cells based on MSCs represents a valuable tool for approaching the genuine in vivo cellular microenvironment and will allow a better understanding of the regenerative potential of AD\u2010MSCs.<\/jats:p>","DOI":"10.1002\/bit.27503","type":"journal-article","created":{"date-parts":[[2020,7,15]],"date-time":"2020-07-15T10:58:02Z","timestamp":1594810682000},"page":"3265-3276","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":29,"title":["Live reporting for hypoxia: Hypoxia sensor\u2013modified mesenchymal stem cells as in vitro reporters"],"prefix":"10.1002","volume":"117","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5337-299X","authenticated-orcid":false,"given":"Carola","family":"Schmitz","sequence":"first","affiliation":[{"name":"Institute of Technical Chemistry Gottfried Wilhelm Leibniz University Hannover  Hannover 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