| Assay | Fish species | Endpoints | Reference(s) |
| Corticosteroid secretion | Oncorhynchus mykiss | Corticosteroid secretion in response to ACTH | [106] |
| Rapid developmental toxicity HTS (in Tier 2) | Zebrafish | Morphological endpoints (edema, bent body axes, pigmentation anomalies, and organ malformations) | [40, 107-111] |
| Fish sex development test | Fathead minnow
Medaka
Zebrafish |
Designed to detect (anti-) estrogenic and (anti-) androgenic effects. Animals are exposed to test chemical before the onset of sexual differentiation.
Vitellogenin induction in males/inhibition in females. Gonadal histopathology Hormone levels Sex ratio Development of intersex |
[112]
[113]
[114] |
| Fish Two Generation Assay | Fathead minnow
Medaka Zebrafish |
Whole body, serum, tissue T4 levels | [115] |
| Locomotion medium throughput assay (in Tier 2) | Zebrafish | Can identify subtle developmental abnormalities between the nervous and musculoskeletal systems | [116] |
| Sex specific behavior | Zebrafish | Sex specific behaviors (aggressive: nipping, chasing, circling, avoiding, and reproductive: female association, spawning, chasing, and nipping) | [117] |
| Short-term reproduction assay/ 21-day fish assay | Fathead minnow
Medaka |
Designed to detect (anti-) estrogenic and (anti-) androgenic effects. Mature male and female fish will be monitored during a 21-day chemical exposure; survival, reproductive behavior, and secondary sexual characteristics will be observed while fecundity and fertilization success will be monitored daily. At termination of the assay, measurements will be made of a number of endpoints reflective of the status of the reproductive endocrine system, including the GSI, gonadal histology, and plasma concentrations of vitellogenin. | [115], [118]; |
| Transgenic reporter lines | Zebrafish | Current lines can detect estrogenic activity and aromatase induction. More transgenic lines are being developed. | |
| Assay | Amphibian species | Endpoints | Reference(s) |
| Corticosteroid secretion | X. laevis
Rana catesbeiana |
Corticosteroid secretion in response to ACTH | [123] |
| SEXDAMAX | X. laevis
X. tropicalis |
Sexual differentiation
Metamorphosis |
[124]
[125] |