Levels of neurotrophin-3 (NT-3) in the mouse brain were measured by a highly sensitive enzyme immunoassay (EIA). The monoclonal antibody, 3W3, was labeled with beta-galactosidase, followed by measurement of galactosidase activity. The detection limit of the EIA system was 0.4 pg/well (4 pg/ml). At 1 and 8 weeks of age, the highest level of NT-3 was detected in the hippocampus, a relatively high level also observed in the cerebellum. In contrast, in the cortex, the striatum, the diencephalon, the midbrain, and the brainstem, NT-3 levels were low. Furthermore, we examined the developmental changes of NT-3 level in the hippocampus and the cerebellum. In the hippocampus, the NT-3 levels were more than 20 ng/g tissue from 1 week to 14 weeks of age, but at 20 weeks of age the level decreased to about half. In the cerebellum, although the NT-3 level was high at 1 week of age, the levels were gradually decreased to one-fourth by 20 weeks of age. In peripheral tissues, a large amount of NT-3 protein was observed in the heart.