Increased amounts of ganglioside GD3 [II3 (NeuAc)2-LacCer], associated with reactive gliosis, have been documented in a variety of neurodegenerative disorders. GD3 expression has also been reported in microglial cells, not only during development but also in reactive states, where the glial activation is considered to be part of the repair process. It is important to find markers in cerebrospinal fluid that will enable us to identify damage and register changes in pathological processes within the brain. A sensitive and practically applicable method for determination of GD3 ganglioside in cerebrospinal fluid has been developed. The procedure, which includes extraction, purification on silica gel and thin-layer enzyme-linked immunostaining, also allows determination of sulphatide, a marker of demyelinating processes, in the same portion of CSF. The method has been applied to CSF samples from 101 normal individuals aged 2-83 years. The GD3 concentration was found to be significantly correlated to age and reflecting the concentrations within the brain. GD3 ganglioside analysis by means of this method might be useful for studying glial changes during brain maturation as well as in brain disorders.