Enzyme treatment (protease or trypsin) was applied to formalin-fixed paraffin-embedded materials and virus-infected cultured cells to detect viral antigens by immunofluorescence. The viral antigens were demonstrated in several organs of autopsy or biopsy cases of which diagnoses had been established by immunofluorescence or virus isolation using frozen materials, or suspected on the basis of serology and/or histopathological findings. These included herpes simplex, varicella-zoster, cytomegalo, subacute sclerosing panencephalitis, progressive multifocal leukoencephalopathy, Japanese B encephalitis, measles, acute hemorrhagic conjunctivitis, Lassa and Korean hemorrhagic fever. Antigen could be recovered also in virus-infected cells (herpes simplex, measles, Lassa, Ebola, Marburg, Rift Valley, Congo and Korean Hemorrhagic fever) by enzyme treatment after periods of formalin fixation of four weeks and storage of three months. In herpes simplex virus-infected mouse brain, antigen was detected after fixation for three months in formalin.