IMPORTANCE: Brain biopsy specimens that exhibit encephalitis without specific histopathologic features pose a diagnostic challenge to neuropathologists and neurologists. Such cases are generally referred to pathologically as encephalitis, not otherwise specified (ENOS). A systematic approach to diagnostic evaluation in such patients is challenging, and currently there is no generally accepted algorithm. OBJECTIVE: To examine ultimate diagnostic outcomes in patients with ENOS diagnosed by brain biopsy. DESIGN, SETTING, AND PARTICIPANTS: This retrospective case series at the University of California, San Francisco, Medical Center, a tertiary care urban neurosciences center, studied patients with encephalitis diagnosed by brain biopsy from January 1, 1983, through December 31, 2011. EXPOSURES: Brain biopsy. MAIN OUTCOMES AND MEASURES: Clinical and neuropathologic diagnosis. RESULTS: Among 58 patients who met the inclusion criteria for the study, the original pathologic diagnosis was ENOS in 49 patients (84%). The median age was 40 years (interquartile range, 27-53 years), 35 patients were male, and 13 had known human immunodeficiency virus or AIDS. Median time from onset of symptoms to brain biopsy was 66 days (interquartile range, 18-135 days). For the 29 patients in whom material for pathologic analysis was still available, additional neuropathologic review led to a more specific categorization in 10 (34%). Clinical detail and follow-up information was available for 42 patients, and a specific diagnosis was reached with the help of ancillary testing and/or clinical follow-up in 12 patients. Despite a comprehensive neuropathologic review with additional studies and information, 27 patients still had to be classified in the ENOS category at the end of the study. CONCLUSIONS AND RELEVANCE: ENOS is the most common initial type of encephalitis diagnosed by brain biopsy. In such patients, it may be worth having the biopsy materials reviewed again in a comprehensive fashion by a neuropathologist because additional review led to a more specific categorization in one-third of our cases. Ancillary testing, clinical correlation, and clinical follow-up establish more specific diagnoses in some patients. ENOS still remains a diagnostic challenge after all these efforts in many cases. Current algorithms are of limited value. More advanced methods and better diagnostic algorithms are needed to characterize these patients.