A significant reduction in the number of virus-induced myocardial lesions was effected by administration of murine interferon beta (IFN-β) or polyriboinosinic:polyribocytidylic acid copolymer (pI:pC) at -24, 0, or 24 h but not 72 h postinoculation (p.i.) of coxsackievirus B3 (CVB3) to adolescent CD-1 mice. Inoculation of interferon at any of the four times did not reduce virus titers in heart tissues at three or seven days p.i., but inoculation of pI:pC at -24, 0, or 24 h p.i. significantly reduced virus titers. Administration of antimurine IFN-β at 72 h p.i. significantly reduced myocarditic lesion numbers. The results suggest that there are two identifiable times after CVB3 inoculation in which interferon may play a role in CVB3-induced myocarditis: a very early time (±24 h of virus entry) in which the presence of interferon is beneficial to the animal and a later time (72 h p.i. of virus) in which absence of interferon is beneficial to the animal. In vitro studies on the effects of IFN-β or anti-IFN-β antiserum on replication of CVB3 in permissive primary cultures of murine neonatal skin fibroblasts show that this virus is sensitive to the antiviral action of interferon which is produced in infected cultures.