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. 2021 Feb 9;128(2):225–241. doi: 10.1007/s00702-021-02303-5

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© The Author(s) 2021

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — Generation of 5-HT specific neurons from human iPSC-derived 5-HT specific progenitors. A Our differentiation protocol generates βTUBIII + neurons (A, a stained in green) and GFAP + astrocytes (A, a–c, stained in red, examples marked by red arrows) following 5 weeks of neuronal maturation. 5-week-old neurons were additionally proven to be mature neurons illustrated by positive staining of (A, b) MAP2 (stained in green) and NeuN (stained in magenta) and (A, c) Tau (stained in green), and NeuN (stained in magenta). Mature neurons are marked by yellow arrows. Pictures were taken using epifluorescence microscopy. B All neurons were co-stained for an antibody against βTUBIII and TPH2, 5-HT, and 5-HTT after 4–5 weeks of neuronal maturation (differentiation week 7–8). Pictures were taken using confocal microscopy. Scale bar: 50 μm. Cell nuclei were counterstained with DAPI and Alexa Fluor 488 (βTUBIII, MAP2, Tau) and Alexa Fluor 555 (GFAP, TPH2, 5-HT, 5-HTT) and Alexa Fluor 647 (NeuN) were used to visualize target proteins. 5-HT specific neuronal differentiation was verified using the JMUi001-A iPS line (data not shown)