Imaging Axon Dynamics in Zebrafish

For live imaging, embryos were anesthetized in 0.02% tricaine and mounted in 1% low melting agarose in 10 mM HEPES E3 medium as previously described (Andersen et al., 2011 (link)). Live high speed imaging of EB3-GFP comets was performed with an Opterra Swept-Field confocal microscope (Bruker Nano Surfaces FM) equipped with a Nikon CFI Plan Apo VC 60x (NA 1.40) or 100x oil-immersion objective (NA 1.40). Embryos were imaged at stages between 18 and 26 hpf, while peripheral axons are initiating and arborizing. Z-stacks of 5–50 1-μm optical sections were captured at 2–9 s time intervals, for total durations between 3 and 20 min.