{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,17]],"date-time":"2026-08-17T21:54:24Z","timestamp":1787003664451,"version":"build-2736575974"},"reference-count":32,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2026,7,21]],"date-time":"2026-07-21T00:00:00Z","timestamp":1784592000000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2026,7,21]],"date-time":"2026-07-21T00:00:00Z","timestamp":1784592000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Sci Rep"],"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Human 3D in vitro models are increasingly adopted in preclinical drug discovery, as they better recapitulate tissue-like architecture, cell-cell interactions, and organ-specific functions compared with conventional 2D cultures. In parallel, initiatives such as the FDA Modernization Act 2.0 and the proposed 3.0 encourage the use of human-relevant in vitro systems as New Approach Methodologies (NAMs) alongside animal models. However, most 3D cell-based workflows rely on manual protocols that are difficult to standardize, producing variable spheroid morphology and fragmented culture steps that impair assay robustness and cross-study comparability. To address these challenges, we present the MO:BOT, a modular benchtop platform designed to standardize and automate complex 3D cell-based workflows. The MO:BOT integrates critical steps of 3D cell culture, including accurate cell seeding, precise medium exchange, on-deck image-based quality control, scalable compound dosing, and downstream assays. Here, we describe a proof-of-concept automated protocol to generate HepG2 liver spheroids and perform acetaminophen (APAP) drug-response testing. Compared with a manual workflow, the MO:BOT not only reduces hands-on time but also minimizes well-to-well variability in liver spheroid size and improves overall spheroid viability. We demonstrate that standardized, well-tuned pipetting routines can maintain the integrity of delicate 3D structures. An MO:BOT-automated APAP dose-exposure experiment with seven concentrations yields a clear sigmoidal cell-toxicity profile with matching changes in viability, LDH release, and ALT activity, demonstrating that improved spheroid uniformity enhances the sensitivity and robustness of downstream assays. The MO:BOT advances the standardization, scalability, and reproducibility of 3D in vitro models, supporting their broader adoption in preclinical research.<\/jats:p>","DOI":"10.1038\/s41598-026-58939-4","type":"journal-article","created":{"date-parts":[[2026,7,21]],"date-time":"2026-07-21T09:02:14Z","timestamp":1784624534000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Automation of 3D liver spheroid generation and acetaminophen dose\u2013response on the MO:BOT enhances assay robustness and precision"],"prefix":"10.1038","volume":"16","author":[{"given":"Dana","family":"Hellmold","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Daniel S.","family":"Ziemianowicz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Frowin","family":"Ellermann","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Philipp","family":"Depperschmidt","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Max","family":"Appold","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ahmed S.","family":"Omar","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jonathan","family":"Kurz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Frank","family":"Krieg-Schneider","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Thorben Pascal","family":"Hoppe","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David","family":"Hackenberger","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lukas","family":"Gaats","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julia","family":"Vallverd\u00fa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2026,7,21]]},"reference":[{"issue":"8","key":"58939_CR1","doi-asserted-by":"publisher","first-page":"545","DOI":"10.1038\/s44222-023-00063-3","volume":"1","author":"A Loewa","year":"2023","unstructured":"Loewa, A., Feng, J. J. & Hedtrich, S. Human disease models in drug development. Nat. Rev. Bioeng. 1 (8), 545\u2013559. https:\/\/doi.org\/10.1038\/s44222-023-00063-3 (2023).","journal-title":"Nat. Rev. Bioeng."},{"issue":"21","key":"58939_CR2","doi-asserted-by":"publisher","first-page":"e175824","DOI":"10.1172\/JCI175824","volume":"133","author":"PJH Zushin","year":"2023","unstructured":"Zushin, P. J. H., Mukherjee, S. & Wu, J. C. FDA Modernization Act 2.0: transitioning beyond animal models with human cells, organoids, and AI\/ML-based approaches. J. Clin. Invest. 133 (21), e175824. https:\/\/doi.org\/10.1172\/JCI175824 (2023). PubMed PMID: 37909337; PubMed Central PMCID: PMC10617761.","journal-title":"J. Clin. Invest."},{"issue":"3","key":"58939_CR3","doi-asserted-by":"publisher","first-page":"449","DOI":"10.1111\/aor.14503","volume":"47","author":"JJ Han","year":"2023","unstructured":"Han, J. J. FDA Modernization Act 2.0 allows for alternatives to animal testing. Artif. Organs. 47 (3), 449\u2013450. https:\/\/doi.org\/10.1111\/aor.14503 (2023).","journal-title":"Artif. Organs"},{"key":"58939_CR4","unstructured":"U.S. Food and Drug Administration. Roadmap to Reducing Animal Testing in Preclinical Safety Studies [Internet]. 2025 Apr [cited 2026 Oct 5]. Available from: (2025). https:\/\/www.fda.gov\/files\/newsroom\/published\/roadmap_to_reducing_animal_testing_in_preclinical_safety_studies.pdf"},{"key":"58939_CR5","unstructured":"FDA Modernization Act 3.0 [Internet]. S.355. Available from: (2025). https:\/\/www.govinfo.gov\/app\/details\/BILLS-119s355is"},{"key":"58939_CR6","doi-asserted-by":"publisher","first-page":"33","DOI":"10.3389\/fmolb.2020.00033","volume":"7","author":"C Jensen","year":"2020","unstructured":"Jensen, C. & Teng, Y. Is It Time to Start Transitioning From 2D to 3D Cell Culture? Front. Mol. Biosci. 7, 33. https:\/\/doi.org\/10.3389\/fmolb.2020.00033 (2020).","journal-title":"Front. Mol. 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Biosci Rep 35(3), e00208. https:\/\/doi.org\/10.1042\/BSR20150034 (2015).","journal-title":"Biosci Rep"}],"container-title":["Scientific Reports"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-58939-4.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-58939-4","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-58939-4.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,7,21]],"date-time":"2026-07-21T09:02:20Z","timestamp":1784624540000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.nature.com\/articles\/s41598-026-58939-4"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,7,21]]},"references-count":32,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2026,12]]}},"alternative-id":["58939"],"URL":"https:\/\/doi.org\/10.1038\/s41598-026-58939-4","relation":{"has-preprint":[{"id-type":"doi","id":"10.21203\/rs.3.rs-9116861\/v1","asserted-by":"object"}]},"ISSN":["2045-2322"],"issn-type":[{"value":"2045-2322","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,7,21]]},"assertion":[{"value":"13 March 2026","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"17 June 2026","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"21 July 2026","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"All authors are employees of mo:re GmbH, a for-profit organization that develops an automated 3D cell culture and organoid platform, namely the MO:BOT, related to the subject of this research. The research reported here was conducted as part of the authors\u2019 employment at mo:re GmbH, which provided salary support and research funding. These relationships may be considered competing financial interests. The authors declare that they have no additional financial or non financial competing interests.","order":1,"name":"Ethics","label":"Competing interests","group":{"name":"EthicsHeading","label":"Declarations"}}],"article-number":"22707"}}