Replica-immunogold technique applied to studies on measles virus morphogenesis.

The replica technique was applied to studies on the dynamic process of measles virus budding on infected HeLa cells. Virus structures were identified by labeling with anti-measles antibodies and protein A-gold. The combination of these two methods enabled us to characterize the sequence of virus budding at the plasma membrane, to localize virus structures on cytoskeletons of infected cells, and to study the influence of Ca2+ ions on virus structures at the plasma membrane. Studies on platinum carbon surface replicas suggest that the process of virus budding is similar to the genesis of cellular microvilli. Replicas prepared from cytoskeletons of infected cells reveal a close association of budding virus with actin filaments composing the outer parts of the networks. Replicas of apical plasma membranes isolated from infected cells show the attachment of viral nucleocapsids to the protoplasmic membrane face of infected cells. These nucleocapsids are not present on membranes prepared from cells treated with calcium and the ionophore A23187. In addition viral cell surface antigens become randomly distributed on these cells. The data suggest that measles virus morphogenesis at the plasma membrane of cultured cells is dependent on the function of the cytoskeleton and may be influenced by Ca2+ ions.