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. 2017 Sep 8;7:11028. doi: 10.1038/s41598-017-11403-w

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© The Author(s) 2017

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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Development of spontaneous and transplanted MMTV-PyMT tumours. (a–d) Immunofluorescence images of spontaneous MMTV-PyMT tumour development. Staining for; cytokeratin-5 (K5; red), cytokeratin-8 (K8; green) and DAPI (blue). Representative images are shown for: (a) Ductal structures with K8 positive luminal cells surrounded by K5 positive basal cell lining, (b) hyperplasia, (c) adenoma-like growth and (d) carcinoma-like growth. Scale bar: 50 µm. (e–f) Immunoflourescence images of organoids from primary PyMT tumour cells (e) or primary normal mammary epithelial cells (f) cultured in matrigel/collagen gels for 2 weeks. Cultures are stained for cytokeratin-5 (K5; cyan), cytokeratin-14 (K14; red), cytokeratin-8 (K8; green) and DAPI (blue). Scale bar: 200 µm. (g–i) Haemotoxylin and Eosin (H&E) staining of different stages of tumourigenesis from 10-week old MMTV-PyMT Mammary gland (FVB). Representative images of (g) hyperplasia, (h) adenoma transition and (i) carcinoma. Scale bar: 50 µm. (l–n) H&E staining of primary PyMT tumour 2 weeks post-transplantation in growth factor reduced matrigel. Representative images show ductal-lobular structures within the centre of the plug (l); transition from a ductal-lobular structure to more tumour-like growth towards the edge of the plug (m); carcinoma-like growth at plug’s edge (n) Scale bar: 50 µm. (o–p) Ki67 DAB staining (left panel) and immunofluorescence staining for cytokeratin-5 (K5; red) and DAPI (right panel) of ductal-lobular structures (o) and tumour-like growth (p). Scale bar: 50 µm.