OECD validation study to assess intra- and inter-laboratory reproducibility of the zebrafish embryo toxicity test for acute aquatic toxicity testing

- Zebrafish embryo acute toxicity test (ZFET) validated for aquatic toxicity testing.
- 20 chemicals tested in 11 laboratories with standardised ZFET protocol.
- ZFET shows good within- and between laboratory reproducibility (CV < 30%).
- ZFET available as OECD TG 236 and proposed for acute fish toxicity testing.
- Use of ZFET will reduce numbers of fish used in aquatic toxicity testing.

The OECD validation study of the zebrafish embryo acute toxicity test (ZFET) for acute aquatic toxicity testing evaluated the ZFET reproducibility by testing 20 chemicals at 5 different concentrations in 3 independent runs in at least 3 laboratories. Stock solutions and test concentrations were analytically confirmed for 11 chemicals. Newly fertilised zebrafish eggs (20/concentration and control) were exposed for 96 h to chemicals. Four apical endpoints were recorded daily as indicators of acute lethality: coagulation of the embryo, lack of somite formation, non-detachment of the tail bud from the yolk sac and lack of heartbeat. Results (LC50 values for 48/96 h exposure) show that the ZFET is a robust method with a good intra- and inter-laboratory reproducibility (CV < 30%) for most chemicals and laboratories. The reproducibility was lower (CV > 30%) for some very toxic or volatile chemicals, and chemicals tested close to their limit of solubility. The ZFET is now available as OECD Test Guideline 236.Considering the high predictive capacity of the ZFET demonstrated by Belanger et al. (2013) in their retrospective analysis of acute fish toxicity and fish embryo acute toxicity data, the ZFET is ready to be considered for acute fish toxicity for regulatory purposes.