We have examined the effects of heat and several chemical inactivating agents on the buoyant density, particle-associated polypeptides and ultrastructure of reovirus particles. Treatment at pH 11 removed polypeptide sigma 1 from the outer capsid of reovirus type 2 but not from type 1; resultant particles were unchanged in their buoyant density and morphology. Treatment of reovirus types 2 and 3 with 2.5 M-guanidine-HCl produced particles with unchanged polypeptide content but an increased buoyant density, and caused aggregation of type 3 but not type 2. Treatment with 1% SDS removed polypeptide sigma 3 from both types 1 and 2 and increased the buoyant density of the virus particles. The outer capsid of SDS-treated virions was greatly altered and often indistinct. Treatment of type 3 with either 1% phenol or 33% ethanol produced particles that had a full complement of polypeptides, were unaltered in buoyant density, but were greatly aggregated. Thus, these inactivating agents affect reovirus particles in specific and distinct ways. The differential effects of such treatments can thus be used to study the structure and function of the reovirus capsid components.