Apoptosis Assay

Apoptotic cells were detected by acridine orange (AO) staining in live larvae of the wild-type AB zebrafish line. Briefly, after E804 treatment (at 72 hpf), the larvae (n = 10) were washed with incubated with fish water for three times, incubated with 10 µg AO staining solution in the dark for 30 minutes at room temperature, and then washed with PBS for three times. Microscopy was performed on 10 tails of larvae anesthetized with 0.16% tricaine within 20 minutes by using a fluorescence microscope (Zeiss, Jena, Germany). The experiments were conducted in three replicates and all tests were repeated for three times.