From January 1973 through July 1981, 40 patients underwent brain biopsy for suspected herpes simplex encephalitis at the University of Alabama Medical Center. Biopsy was performed as a component of experimental antiviral studies to ensure correct diagnosis. A tentative diagnosis was made on clinical grounds, and the site of biopsy was chosen on the basis of localizing neurological signs, the electroencephalogram, and/or computed tomographic (CT) scan abnormalities. Patients ranged in age from 15 months to 73 years, with a median age of 19 years. Thirty patients were male, and 10 were female. Brain tissue from 17 of the 40 patients grew herpes simplex virus in tissue culture. There was one biopsy-related complication, a wound dehiscence requiring secondary closure. In no case was death attributable to the brain biopsy procedure. The outlook in patients with proven herpes encephalitis worsened directly with both an increasing interval between the onset of symptoms and the initiation of antiviral therapy and the age of the patient. The presence of a CT scan abnormality before the initiation of therapy was associated with a very poor prognosis. Three patients not included in this series were referred for brain biopsy to rule out herpes simplex encephalitis, but were found to have bacterial infections. Two had subdural empyemas and the third had a brain abscess with overlying subdural empyema. Current technique calls for biopsy of the anterior portion of the inferior temporal gyrus on the affected side.