The neurological mouse mutant strainreelerdisplays abnormal laminar organization of several brain structures as a consequence of a defect in cell migration during neurodevelopment. This phenotype is a result of the disruption ofreelin, a gene encoding a protein that has several structural characteristics of extracellular matrix proteins. To understand the molecular basis of the action of Reelin on neuronal migration, we constructed a full-lengthreelinclone and used it to direct Reelin expression. Here, we demonstrate that Reelin is a secreted glycoprotein and that a highly charged C-terminal region is essential for secretion. In addition, we demonstrate that an amino acid sequence present in the N-terminal region of Reelin contains an epitope that is recognized by the CR-50 monoclonal antibody. CR-50 was raised against an antigen expressed in normal mouse brain that is absent inreelermice. The interaction of CR-50 with its epitope leads to the disruption of neural cell aggregationin vitro. Here, we used CR-50 to precipitate Reelin from reticulocyte extracts programmed withreelinmRNA, from cells transfected withreelinclones, and from cerebellar explants. Thereelingene product seems to function as an instructive signal in the regulation of neuronal migration.