AbstractAn epitope recognized by the monoclonal antibody I‐5G9 was expressed by all neural crest cells shortly after explantation into culture. At this time all neural crest cells actively migrated away from the neural tube. Immunoreactivity was localized intracellularly and organized into stress fiber‐like filaments. Often, immunofluorescence was particularly high in short fibers in the lamellipodia of the reading edge of migrating cells. Two‐week‐old cultures had a diameter of 8–10 mm. At that stage a ring of immunoreactive cells was present at the periphery of each culture, an area where cells were still migratory. An inner concentric circle had reduced and more granular staining. In this area cells had ceased to migrate. In the center of the culture cells were multilayered, nonmigratory, and did not bind I‐5G9. After creating a lesion in the nonreactive central region, some cells resumed migration into the lesioned area and reexpressed the epitope. I‐5G9 staining and phalloidin fluorescence colocalized partially in some cells and completely in others. It is concluded that the epitope recognized by I‐5G9 is expressed in a migration‐dependent manner. The partial colocalization of I‐5G9 and phalloidin fluorescence supports the notion that the epitope recognized by I‐5G9 is specifically expressed in stress fibers of migratory cells, possibly in one of the actin‐associated proteins or an F actin‐a