Detection of HIV‐1 sequences in children using radioactive and colorimetric polymerase chain reactions

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Abstract
The detection of HIV‐1 proviral DMA in children born to seropositive mothers was studied using the polymerase chain reaction with either a radioactive electrophoretic method or a novel procedure that employs colorimetric microwell visualization.Peripheral blood mononuclear cell lysates from 18 HIV‐1 infected children and 28 uninfected subjects were assayed for a 142 bp fragment of DNA from the gag region of HIV‐1 using the primer pair SK145‐431. Detection of amplified DNA was carried out by hybridization with a radiolabeled SK102 probe, or with a tagged SK102 probe permitting colorimetric detection.The radioactive detection procedure demonstrated 100% specificity and correlated with the serological results. The assay was more sensitive than the p24 antigen test, but two false negative results were obtained. One was from a sample taken at 2 weeks, an age at which unde‐tectable provirus levels were reported in almost all HIV‐1 infected newborns. The second was probably due to a low copy number of proviral DNA, as positive results were obtained in all other (6) samples from this child.Comparative analysis in a limited number of specimens of radioactive and colorimetric detection following PCR revealed 100% specificity and comparable sensitivity with 4 discordant results.The results show that PCR is the best method for early diagnosis of HIV‐1 infection in pediatric subjects. The study also demonstrated the value of a colorimetric detection method for PCR products. This colorimetric microwell plate procedure may prove a useful technique in routine diagnosis of HIV‐1 infection in children.