Abstract The properties of cytosolic thyroxine binding protein were studied in the cortex and cerebellum of the rat at different stages of postnatal development: 1. (1) Polyacrylamide-gel electrophoretic analysis showed that rat-brain cortex and cerebellum contain the same cytosolic thyroxine-binding protein which is very similar to the liver-corresponding entity. No changes in the electrophoretic mobility were seen during development in the 2 brain regions. In contrast, no defined triiodothyronine-binding component could be observed by the same technique. 2. (2) Kinetic analysis studies revealed that the equilibrium of binding is reached in ∼10 min whatever the brain region, the concentration of cytosolic protein and the stage of development. In all these cases saturation was obtained with the same thyroxine concentration (∼5 × 10 −7 M). Scatchard analysis also showed that whatever the experimental conditions, brain cytosolic protein contains a single class of thyroxine-binding sites with a K A of ∼8 × 10 7 M −1. 3. (3) Comparison of the K A during development showed that this constant remains unchanged from day 3 after birth until day 35 in both the cortex and the cerebellum. In contrast the number of binding sites significantly decreases in the cortex (∼2-fold; p < 0.001) from day 3 to 35 with an already significant decline from day 3 to 6 ( p < 0.001). In the cerebellum this decline was even more marked since almost no binding activity was left at adulthood. Comparison of cortex and cerebellum binding activities also showed that this latter region contains ∼ half the binding sites ( p < 0.001) at every stage of development studied.