A malignant astrocytoma was cultured from a tissue biopsy taken at surgical resection, and serially passaged 65 times over a period of 4 years. Early culture cells showed a variety of morphological types, but were all positively labelled with a polyclonal antibody directed against glial fibrillary acidic protein (GFAP). As the number of passages increased small, GFAP-positive cells became the predominant cell type. Flow cytometry demonstrated changes in GFAP expression between early and late passages; early passages showing greater fluorescence intensity than those at later passages. All cells, however, remained positive for GFAP. In addition, this line (IPSB-18) was both vimentin and glutamine synthetase positive and no change in the expression of these two proteins was detectable on continued sequential passaging. Fibronectin was not detectable at any stage. As the number of passages increased, the population doubling time was shortened from 72 h at passage 5, to 28 h at passage 30. This line is unusual in its retention of GFAP expression through 65 sequential passages; the majority of astrocytoma lines lose this capacity after a few passages in vitro. The monoclonal antibody, A2B5, which recognizes surface gangliosides, has been previously shown to identify sub-set type 2 astrocytes in normal neural tissue in vitro. The expression of A2B5 gangliosides by 5-20% of GFAP-positive IPSB-18 cells, and their co-expression with the ganglioside GD3 identified by the LB1 monoclonal antibody, shows that cells with a similar phenotype to the type 1 and type 2 astrocytes of optic nerve are present in cultured gliomas, even after long term sequential passaging.