The MolDarT is a novel short‐term assay for testing mechanism‐based molecular effects in developing zebrafish embryos. The objective of this study was to evaluate the inducibility of vitellogenin1 mRNA (Vtg1) by the estrogenically active compounds 17β‐Estradiol (E2), 17α‐Ethinylestradiol (EE2), Nonylphenol (NP), Bisphenol A (BPA), Cyproconazol, and the suspected xeno‐estrogen Atrazin in the MolDarT. Freshly fertilized zebrafish eggs were exposed semistatically for 120 h. Using reverse transcription real‐time PCR, the relative abundance of Vtg1 was measured. For EE2 a dose‐response relationship was established with EC50 = 60.7 ng/L (205 pM). Induction of Vtg1 was significant at concentrations of 84 pM EE2 (25 ng EE2/L) and above, 10 nM E2 (2.7 μg E2/L), 100 nM E2 (27 μg E2/L), 10 μM BPA (2280 μg BPA/L), and 15 μM BPA (3420 μg BPA/L). At NP concentrations of 0.75 μM (165 μg NP/L) and 1.5 μM (330 μg NP/L) Vtg1 was significantly down‐regulated. Both atrazine and cyproconazol showed no effect on relative Vtg1 abundance. With this study we further characterize the MolDarT assay and show its applicability for effect screening of compounds. © 2007 Wiley Periodicals, Inc. Environ Toxicol 22: 185–193, 2007.