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Files in this Data Supplement:
Movie 1. Neural crest cells avoid stripes of substrate-bound ephrin-B2-Fc. Neural tubes were plated onto stripes of substrate-bound ephrin-B2-Fc on coverslips. Overnight, neural crest cells migrated out onto the coverslip and avoided the substrate-bound ephrin-B2-Fc distal to the neural tube. Neural crest cells distal to the neural tube were then examined by timelapse microscopy for 1 hour. Phase images of the cells were merged with a fluorescent image of the Texas Red-dextran-labelled ephrin-B2-Fc stripes (lighter, broader stripe, approximately 40 μm wide). Neural crest cells avoided the ephrin-B2-Fc stripes, although some cells did migrate across the stripes, often rounding up as they did so. Neural crest cells spread lamellipodia, but these protrusive structures tended to rapidly collapse on contact with ephrin-B2-Fc stripes. The internalisation of phase-bright vesicles was associated with collapse. Some neural crest cells appeared to exhibit a ‘love-hate’ relationship with the ephrin-B2-Fc stripes, migrating along their edges. These neural crest cells cycle between protrusion onto and withdrawal from these stripes, with withdrawal characterised by the formation of membrane ruffles.
Movie 2. Soluble, pre-clustered ephrin-B2-Fc decreases the stability of neural crest cell lamellipodia. Neural tubes were plated to fibronectin-coated coverslips and neural crest cells allowed to emerge overnight. Neural crest cells were examined for 30 minutes and then, following the addition of 2 μg/ml pre-clustered ephrin-B2-Fc, were examined by timelapse microscopy for an additional 60 minutes. This movie shows an individual neural crest cell (cropped from a larger movie) with its leading edge marked with a red line. A flash of white indicates when the pre-clustered ephrin-B2-Fc has been added. Following stimulation, the neural crest cell appears to speed up, in fact undergoing increased cycling between protrusion and collapse of its lamellipodia.
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