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. 2023 Jan 3;42(9):665–678. doi: 10.1038/s41388-022-02588-0

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© The Author(s) 2023

Open Access This 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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — A The workflow for labeling patient-derived HGSC cells with fluorescence markers, cell purification, organoid culture, and imaging-based studies. FACS, fluorescence-activated cell sorting, H&E, hematoxylin-eosin. B Representative type 2 MDA-HGSC-2414 HGSC organoid with fecundity cells captured by laser confocal microscopy with Z-axis scanning. a Reconstructed 3D image of nuclei (Z-scanning). b–e Merged fluorescence images of the cell membrane (green) and nuclei (magenta) from four Z-slices. Cyan arrows: PGCC nuclei; white arrows: fecundity cells. f The reconstructed 3D image demonstrates the overall cell membrane distribution and structures. Pixels were adjusted to 70% transparency to show the vesicle-rich region in the center of the organoid. g–j Color-coded LCK-GFP images. The images demonstrate the complex subcellular structures associated with membrane-rich vesicles and cell membrane borders of the fecundity cells. The Z-slice scales mark the position of the current image. For panels f–j: green arrow: cell membrane; red arrows: large membrane aggregate adjacent to fecundity cells; white arrows: solitary membrane aggregate in the cytoplasm. Bars equal 20 μm. C Key morphological characteristics of type 1 and type 2 organoids and the corresponding structures in patient tumors. From right to left: Live-cell imaging, phase contrast, fixed organoids, H&E-stained, bright field; Live-cell imaging, fluorescence laser confocal. LCK-GFP: green; H2B-mCherry: magenta; Fixed patient tumor tissue, wide-field epifluorescence. Immunofluorescence staining includes Na + /K + ATPase (epithelial cell membrane, red), Vimentin (mesenchymal cells, green), and DAPI (DNA, blue). Cyan arrow: giant nucleus; green arrows: fecundity cells. Bars equal 20 μm.