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Files in this Data Supplement:
Movie 1. Time-lapse imaging of unlabeled salivary gland. E13 SMG imaged at 203 magnification by phase-contrast time-lapse microscopy, as shown in Fig. 1B. Images were captured every 10 minutes for 20 hours. Dynamic epithelial movements are apparent.
Movie 2. Time-lapse imaging of an intact gland (containing both epithelium and mesenchyme) microinjected with adenovirus-GFP. An E13 SMG was microinjected with Ad-GFP and imaged at 203 magnification by time-lapse microscopy, as shown in Fig. 1D. Brightfield images (left) and images of GFP-labeled cells (right) were acquired every 10 minutes for 9.2 hours and are shown at 10 frames/second. Dynamic movements of GFP-labeled cells are observed.
Movie 3. Time-lapse imaging of individual cells of SMG epithelium moving during branching morphogenesis. E12 SMGs were labeled with Ad-GFP, and images were captured using a 203/0.75NA objective, as shown in Fig. 2B. Images were acquired at 12-minute intervals for 20 hours and are shown at 7.5 frames/second. Dynamic movements of GFP-labeled cells are apparent.
Movie 4. Time-lapse video of GFP-labeled epithelial cells and labeled FN during branching morphogenesis. E12 SMGs were labeled with GFP (green) and Alexa Fluor 647-FN (red), as shown in Fig. 3A. Confocal time-lapse images were acquired using a 103/0.3NA objective at 10-minute intervals for 14.5 hours and are shown at 10 frames/second. Dynamic movements of epithelial cells (green) are observed, particularly in cells that contact FN in the basement membrane (red).
Movie 5. Time-lapse imaging of control (left) and hydroxyurea-treated salivary glands (right). E12 SMGs were treated with 0.5 mM hydroxyurea (right) or vehicle control (left), and epithelial cells were labeled with GFP (green) and Alexa Fluor 647-FN (red). Confocal time-lapse images of red and green channels were acquired together with DIC images using a 203/0.75NA objective every 6 minutes for 16.7 hours and are shown at 15 frames/second. Cells treated with hydroxyurea show no change in cell migration relative to cells in the control SMG. Quantitation is shown in Fig. 4.
Movie 6. Time-lapse imaging of salivary epithelial cells late in development (P1). SMGs from P1 mice were labeled with GFP (green) and Alexa Fluor 647-FN (red), as shown in Fig. 5B. Confocal time-lapse images of red and green channels were acquired together with DIC images every 10 minutes with a 203/0.75NA objective for 13.3 hours and are shown at 10 frames/second. GFP-labeled acinar cells (green) surrounded by FN-incorporating mesenchyme cells (red) do not migrate.
Movie 7. Time-lapse imaging of washout of Alex Fluor 647-FN. E12 SMGs were treated with Ad-GFP and Alexa Fluor 647-FN for 6 hours and imaged by confocal microscopy for 1.8 hours. Then the Alexa Fluor 647-FN-containing medium was replaced with unlabeled medium, and imaging was continued. Three black frames were inserted at the time corresponding to the washout. Confocal time-lapse images were collected using a 203/0.75NA objective every 6 minutes for 16 hours and are shown at 15 frames/second. Alexa Fluor 647-FN accumulated particularly prominently at the leading tip (base) of the deepening clefts, as shown in Fig. 6
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