The diagnosis of infection due to herpesviruses can be performed by several laboratory techniques, including virus culture, demonstration of virus-specific cellular antigens, detection of virus-induced serum antibodies, detection of viral nucleic acids (using radiolabeled probes), or visualization of histologic changes in infected tissue. Virus culture is the most sensitive technique for diagnosis of most infections due to herpes simplex virus (HSV) and cytomegalovirus (CMV), but is insensitive for the diagnosis of varicella-zoster (VZV) infection and not routinely available for Epstein-Barr virus (EBV). Identification of viral antigens on infected cells with monoclonal antibodies is useful for definitive identification of virus in cell culture and for rapid diagnosis of infection due to HSV, VZV, or CMV. Serologic studies are useful for the diagnosis of EBV infection, because clinical symptoms and an antibody response occur simultaneously, and other diagnostic tests are generally unavailable. Although serologic studies may be used retrospectively to diagnose infections due to HSV, VZV, or CMV, these results are seldom available in time to be useful in managing the acute infection or in guiding antiviral chemotherapy. Newer diagnostic techniques, such as detection of viral nucleic acids using radiolabeled probes, may become useful in the future for the rapid diagnosis of EBV or CMV infection.