Zebrafish Husbandry and Embryonic Development

Wild type adult zebrafish (AB or TL strain) are maintained in the Aquatic Facility of the Luxembourg Centre for Systems Biomedicine and the Institute of Biology, Leiden University, according to standard protocols [39 ]. Zebrafish eggs were obtained by natural spawning on the day of each experiment, kept in 0.3X Danieau’s solution (14 mM NaCl, 2 mM KCl, 0.12 mM MgSO₄, 1.8 mM Ca(NO₃)₂, 1.5 mM HEPES pH 7.5 and 0.03 M methylene blue) or egg water (60 μg/ml sea salt, Sera Marin, Heinsberg, Germany), and staged by morphology (one-cell stage) for the injections. After each series of injections, the eggs were incubated at 28°C (±0.5) and evaluated up to 5 days post-fertilization (dpf).
Anaesthesia of larvae used for live imaging and COPAS [37 (link),40 (link)] analysis was done with 0.02% buffered Tricaine (3-aminobenzoic acid ethyl ester, Sigma-Aldrich) in egg water.