The distribution of the herpes simplex virus alkaline nuclease (HSV DNase) in rabbit fibroblast cells infected with HSV type 1 or 2 (HSV-1 or HSV-2) has been examined with the aid of immunocytochemical techniques using gold particles as markers. HSV DNase was found to be accumulated within infected nuclei as early as 2.5 hr post-infection. Labeled antibody to HSV DNase subsequently increased in all nuclei after 7 hr and 17 hr. At all times post-infection the virus induced nuclear dense bodies were always the most intensely labeled structures. The association of HSV DNase with nucleoprotein containing structures was readily dissociated by hypotonic shock and detergent treatment, but its association with the dense bodies was not disrupted. Nucleolar 100 kDa protein was found to be simultaneously present in the dense bodies. This suggests that HSV DNase might interfere with ribosomal RNA synthesis and play a role in the degradation of the host-cell metabolism.