AbstractPlasmolipin is a plasma membrane proteolipid which has recently been described as a component of myelin (Cochary et al.:Journal of Neurochemistry55:602–610, 1990). The present study reports the expression and localization of plasmolipin in primary glial cultures and secondary oligooendrocyte cultures. Double‐label immunofluoresccnce showed that plasmolipin was expressed by galactoccrebroside (GC)‐positive oligodendrocytes, but was absent from astrocytes, characterized by their positive staining for glial fibrillary acidic protein (GFAP). At 1 week in culture plasmolipin staining was relatively weak in the cell body of some of the GC‐positive cells. During the following 3 weeks in culture plasmolipin staining of Oligodendrocytes gradually increased and was present in the cell body, its plasma membrane, and all the processes. However, the plasmclipin antibodies did not stain regions of the flat membrane sheets. Western blot analysis of homogenatcs from primary glial cultures showed that plasmolipin levels gradually increased during the first 5 weeks in culture. We conclude that the presence of plasmolipin in myelin is a result of its expression by Oligodend