A chemically defined serum-free medium is described that has been optimized for the selective growth of immature astrocytes starting from early postnatal mouse cerebellar cultures. The medium is a modification of one described recently [Fischer et al, 1982] and consists of epidermal growth factor (EGF, 10 nM), transferrin (10 micrograms/ml), insulin (10 micrograms/ml), and bovine serum albumin (BSA, 1 mg/ml) in BME with Earle's salts. None of these constituents of the medium alone nor combinations of two of them can stimulate the proliferation and survival of immature astrocytes. Only with a combination of EGF and transferrin together with either insulin or BSA can proliferation of the cells be obtained. For optimal growth all four constituents are needed. However, the immature astrocytes can only grow for a limited time period of about 2-3 wk in this medium. Therefore they can be selected for and characterized but not continuously cultivated in this medium.