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. 2019 Dec 11;10:5658. doi: 10.1038/s41467-019-13605-4

Fig. 3. 3D culture of endodermal organoids in ECM gel and Matrigel.

Fig. 3

a Human pediatric gastric enteroids in ECM gel. Scale bar 200 µm. b Planes of whole-mount immunofluorescence of 7-days gastric organoids showing both epithelial (zonula occludens-1, epithelial cadherin and actin) and gastric (ezrin and mucin-5AC) markers. Scale bar 50 µm. c Culture of liver ductal and hepatocyte human organoids in ECM gel, BME and Matrigel. Scale bar 500 µm. d Hepatocyte organoid viability. Mean ± S.D. (n = 8 organoid cultures). e Bright field and H&E images of mouse intestinal organoids in ECM gel and Matrigel. Scale bars 100 µm. f Immunofluorescence analysis of sections of mouse SI organoids in ECM gel and Matrigel, showing epithelial cadherin staining and proliferation marker Ki-67+. Scale bar 50 µm. g Immunohistochemical staining of mouse intestinal organoids in ECM gel and Matrigel. Scale bars 25 µm. h Forming mouse intestinal organoids per field of view at day 4 of culture in ECM gel and Matrigel over two passages. Mean ± S.D. (n = 12 organoid cultures). i Live/Dead assay of human pediatric SI organoids cultured in ECM gel and Matrigel. Calcein-AM shows living cells. Ethidium homodimer-1 shows dead cells. Scale bar 200 µm. j Quantification of vital cells from Live/Dead assay. Mean ± S.D. (n = 8 organoid cultures). k Morphology of eight consecutive passages over a period of 2 months of human pediatric SI organoids in ECM gels. Scale bar 300 µm. l Analyses of four consecutive passages of human pediatric SI organoids diameters at day 3 of culture in ECM gel and Matrigel. Mean ± S.D. (n ≈ 200 organoids). m Bright field of human fetal SI organoids in ECM gel. Scale bar 200 µm. n Whole-mount immunofluorescence Z-Planes of human fetal SI organoids showing crypt stem cell marker olfactomedin-4, crypt Paneth cell marker lysozyme, villi enterocyte marker keratin-20 and actin staining. Scale bar 100 µm. o Analyses of three consecutive passages of human fetal SI organoids diameters at day 3 of culture. (n ≈ 380 organoids). p Single-cell colony (arrows) formation capacity assessed over 3 days in disaggregated human fetal SI organoids in ECM gel and Matrigel. Scale bar 25 µm. Box plots are represented with the central line indicating the median of values, bounds of box indicating first and third quartiles, and whiskers to show minimum and maximum outside the first and third quartiles.