Laboratory diagnostic resources for early detection of HIV perinatal infection in developing countries are limited. The initial signs and symptoms of HIV infection in children can be mistaken for prevalent diseases. The aim of this study was to compare the utility of signs and symptoms, immunologic abnormalities and polymerase chain reaction (PCR) for diagnosis of HIV infection in Brazilian infants younger than 6 months (mo) of age. A total of 58 infants born to HIV+ mothers were followed prospectively from birth. An in-house HIV-1 nested PCR was done at 0-1; 3±1 and 6±1 mo of age. Serum IgG, IgM, IgA and CD4 cell counts were measured by 6 mo. Infection was defined as the persistence of HIV-1 antibodies beyond 18 mo or by an AIDS-defining condition. Twenty children were infected (HIV+) and 38 were not infected (HIV-). Overall, during the first 6 mo of life one or more findings of lymphadenopathy (LY), splenomegaly (SPL), hepatomegaly (HEP), diarrhea(DIA), oral candidiasis (CAN), failure to thrive (FTT), upper respiratory infection (URI) or pneumonia (PN) was detected in 32/38 (84.2%) HIV- infants and in 18/20 (90%) HIV+ infants. The presence of any two signs such as LY, SPL and HEP detected 65% of HIV+ infants within 3 mo and 82% within 6 mo with false+ rates of 5.2% and 10.5%. CAN (76%) and PN (88%) were the most frequent findings among HIV+ infants within 6 mo with false+ rates of 18.4% and 10.5%. The combination of any two of DIA, CAN, FTT, URI, and PN had 100% sensitivity within 6 mo but 54% specificity. PCR was positive in 10/18 (55.5%) HIV+ infants by 1 mo and in all of them (100% sensitivity) within 3 mo of age (100% specificity). Hyper-IgG, M or A was detected in 13/16 (81.3%) HIV+ infants with a false+ rate of 31.3%. Low CD4 counts were found in 9/18 (50%) HIV+ infants with a false+ rate of 7.1%. PCR was the earliest and most reliable method for diagnosing HIV infection. Although having high sensitivity, the predictive value of a positive hyper-IG was low in this population. Diversely from developed countries, the presence of any two findings such as LY, SPL, HEP or the occurrence of PN or CAN can identify most of the truly infected infants within 6 mo. However, the presence of such findings in 10.5 to 18.4% of HIV-infants underscores the need of reliable laboratory tests for early confirmation of diagnosis.