A preparation enriched in Purkinje cells identified by morphological and immunocytochemical criteria

A preparation highly enriched in isolated Purkinje cell perikarya was obtained from the developing rat cerebellum. The technique, which also yields a fraction enriched in differentiated astrocytes, involved the removal of heterogeneously-sized replicating cells in vivo by treatment with hydroxyurea prior to dissociation of cells with mild trypsinization and fractionation by sedimentation at unit gravity. Fractions were monitored in terms of particle size and by light and electron microscopy. Pooled fractions containing at least 50% perikarya of > 14.5 micron diameter were enriched in cells resembling Purkinje neurons. Definitive identification of these cells was obtained using an antiserum specific to Purkinje cells. In immunocytochemical studies about 80% of the cells in the fraction enriched in large cells reacted with the antiserum vs about 4% in the unfractionated cell suspension, whilst most of the other cells were identified as astrocytes containing glial fibrillary acidic protein.