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We have developed a simplified method for unambiguously typing herpes simplex virus. The method depends on the production of cell-associated virus at 34 degrees C and subsequently, on the separation of cellular DNA and viral DNA by Dounce homogenization and the removal of nuclei by centrifugation. Viral nucleic acid was prepared from the cytoplasmic fraction and analyzed after restriction endonuclease cleavage. The method obviates the use of radioactive isotopes, and the viral DNA is effectively free of interfering cellular DNA.