3Z´s method consists of parallel screening of compounds in a sample of ~2500 zebrafish larvae. Larvae are placed in 96-well plates, 1 larva per well and acclimated for 24 hrs. Recording commences after acclimation and drugs are dispensed into the wells via a liquid handling robot. Larval behavior is recorded until 12:00 the following day. In a single run 50 molecules can be screened simultaneously (using three dose sizes and three independent CTRL groups).
Videos of larval behavior are tracked by an image analysis software and swim velocity data are converted into text files. These data are then analysed using a custom made software that yields a detailed readout of sleep and movement parameters.