Coordinated cell movements drive embryonic development and tissue repair, and can also spread disease. Time-lapse microscopy is an integral part in the study of the cell biology of collective cell movements. Advances in imaging techniques enable monitoring dynamic cellular and molecular events in real time within living animals. Here, we demonstrate the use of spinning disk confocal microscopy to investigate coordinated cell movements and epithelial-to-mesenchymal-like transitions during embryonic wound closure in Drosophila. We describe image-based metrics to quantify the efficiency of collective cell migration. Finally, we show the application of super-resolution radial fluctuation microscopy to obtain multidimensional, super-resolution images of protrusive activity in collectively moving cells in vivo. Together, the methods presented here constitute a toolkit for the modern analysis of collective cell migration in living animals.
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