This Test Guideline describes an assay that assesses early life-stage effects and potential adverse consequences of putative endocrine disrupting chemicals (e.g. oestrogens, androgens and steroidogenesis inhibitors) on fish sexual development. In the test, fish are exposed, from newly fertilized egg until the completion of sexual differentiation at about 60 days post hatch, to at least three concentrations of the test substance dissolved in water. In each treatment level and control(s) group(s), a minimum of four replicates is recommended. At termination of the test, two core endpoints are measured in each fish: vitellogenin concentration from head and tail or from blood sampling, and proportion of males, females, intersex and undifferentiated fish through gonadal histology. In fish species possessing a genetic sex marker, the genetic sex is identified to determine sex reversal in individual fish. The combination of the two core endocrine endpoints, vitellogenin concentration and phenotypic (and possibly genotypic) sex ratio, enable the test to indicate the mode of action of the test chemical.