Results of polymerase chain reaction (PCR) and p24 antigen detection after immune complex dissociation (p24-ICD) were compared with antibody results after 18 months of age for human immunodeficiency virus (HIV) diagnosis in 345 prospectively followed, perinatally exposed infants. Of 59 infected and 286 uninfected infants tested at 1-6 months of age, sensitivity and specificity were, respectively, 100% and > 97% for PCR and 90% and > 97% for p24-ICD. Testing was done on > or = 2 occasions in the first 6 months of life in 43 infected infants; 77% had > or = 2 positive results with the same test. Of these infants, 68% had 2 positive p24-ICD tests. In uninfected infants, 96% had only negative tests; none had > 1 positive. By 6 months, all uninfected infants with > or = 2 PCR results could have been diagnosed. HIV status can be determined by PCR by age 6 months in most HIV-exposed infants. p24-ICD should not be used alone, because of its lower sensitivity, but may be useful in areas without advanced laboratory support.