Research Article
Three-Dimensional Gastrointestinal Organoid Culture in Combination with Nerves or Fibroblasts: A Method to Characterize the Gastrointestinal Stem Cell Niche
Figure 5
Positive Dclk1 lineage shown in the enteric nervous system with GFP labeled neurons. Immunofluorescence of small intestine from Dclk1.Cre.GFP-Tomfl/flGFP mice. (a) DAPI for cell nuclei. (b) Positive GFP-fluorescence in neurons (Dclk1.Cre.GFP-Tomfl/flGFP label recombined cells of the Dclk1 lineage from birth on) invading organoids and single Dclk1-positive cells in epithelium, arrowhead indicating lineage of Dclk1-positive neurons, (c) Tom-Red ubiquitously expressed as marker protein complex. (d) Merge of (a)–(c), scale bars 100 μm. (e) Cultured neurons identified by positive GFP-fluorescence and overlapping with PGP9.5 staining as proof of lineage, last panel demonstrating lacZ stained neuronal structures next to smooth musculature within the GI tract, scale bar, 50 μm. (f) Overlapping fluorescence with GFAP as proof of lineage of glia within myenteric plexus, last panel showing lacZ stained glia within myenteric plexus, scale bar, 50 μm.
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