Age is a physiological condition that affects protein expression. We investigated differences in protein level between newborn and adult brains. Brain tissue extracts from male and female adult and neonatal rats were analyzed by two-dimensional gel electrophoresis. Gel comparison revealed the presence of many differences, of qualitative and quantitative nature, between a neonatal and adult brain. The most significant agerelated difference concerned alpha-fetoprotein, which was detected in the brain of neonatal rats only. The levels of 22 proteins, including dihydropyrimidinase-related proteins 1,3, and 4 and 14-3-3 proteins, were higher in the neonatal brain, whereas the levels of 28 proteins, including dihydropyrimidinase-related protein 2, dynamin-1 and many enzymes were higher in the adult brain. We did not detect a consistent sex-related difference in the brain proteins. An inconsistency was observed in the location of the spot representing glial fibrillary acidic protein in the male brain.