Antibodies to human cytomegalovirus (CMV)-specific antigens were determined in sera serially collected from 10 infants with primary CMV infection. Antibodies to pre-early nuclear antigens (PENA), which are detectable in human embryonic lung cells within 3 h of CMV infection by anticomplement immunofluorescence staining, developed in all the patients. However, in contrast to the early response of anti-early antigens (EA), anti-late antigens (LA), and immunoglobulin M antimembrane antigens (MA), seroconversion or the maximum antibody response to PENA was usually observed 1 or more months later. Immunoglobulin M antibody to MA became undetectable soon after recovery from illness, followed by a decrease in anti-EA, anti-PENA, and then anti-LA titers. Results indicated analogy of the clinical significance of anti-PENA in CMV infection to that of anti-Epstein-Barr nuclear antigen in infectious mononucleosis and support the idea that parallel determinations of anti-PENA and IgM anti-MA antibodies can be useful for identifying the acute or chronic phase of primary CMV infection.