Antibodies against vaccinia virus do not neutralize extracellular enveloped virus but prevent virus release from infected cells and comet formation.

Vaccinia virus (VV) produces two antigenically and structurally distinct infectious virions, intracellular mature virus (IMV) and extracellular enveloped virus (EEV). EEV is important for the efficient dissemination of virus both in vivo and in vitro where it causes formation of comet-shaped virus plaques. Here, we show that EEV, in contrast to IMV, is resistant to neutralization by antibodies bound to its surface. However, antibodies against EEV can prevent comet formation in cell culture. To explain this apparent paradox, we investigated the mechanism by which antibodies inhibit comet formation and demonstrated that antibodies prevent EEV release from infected cells, and consequently comet formation, by agglutination of the virus on the cell surface. Two complementary observations allow this conclusion: first, electron microscopy showed that infected cells incubated with medium containing anti-vaccinia virus antibodies have virus aggregates on their surface; second, culture medium from these cells contained a 4 log10 fold reduction in the physical particle/ml titre in comparison with control culture. A mechanism by which antibodies to EEV proteins provide immunological protection is thus restriction of EEV release rather than neutralization of free EEV particles.