title: Transmission of HIV-1 in infants born to seropositive mothers: PCR-amplified proviral DNA detected by flow cytometric analysis of immunoreactive beads.
abstract: The diagnosis of HIV infection in newborns is established by amplification of proviral DNA using the polymerase chain reaction (PCR). We developed a nonisotopic method for heminested PCR using a biotinylated primer among sets of three oligonucleotides, each selected from the HIV long terminal repeat (LTR) and gag sequences. An internal probe incorporating digoxigenin-dUTP was also synthesized by PCR. The PCR products, hybridized with LTR region or gag region probes, were captured with streptavidin-coated magnetic beads and detected by fluorescein isothiocyanate-labeled antidigoxigenin in flow cytometric analysis. This immunoreactive bead assay (PCR-IRB) detected about three copies of HIV proviral DNA. A panel of 50 coded DNA specimens of infants previously assayed by conventional PCR and with known clinical results revealed that the PCR-IRB findings using LTR, but not gag, were in agreement. A double-blind prospective study of blood samples from 14 mother-infant pairs using the PCR-IRB amplification of LTR gave results similar to the commercial Amplicor HIV-1 PCR test and were consistent with the clinical outcomes. PCR-IRB results were positive for 11 mothers and three infants, one at birth, one at 2 weeks after birth, and one at 8 weeks after birth. PCR-IRB is a simple, reliable, specific, and automatable assay useful in the early diagnosis of perinatal HIV infection in clinical practice and regional screening programs.
authors.affiliation: Department of Pediatrics, University of California San Francisco, 94143, USA.
authors.forename: A
authors.initials: A
authors.lastname: Dorenbaum
authors.forename: K S
authors.initials: KS
authors.lastname: Venkateswaran
authors.forename: G
authors.initials: G
authors.lastname: Yang
authors.forename: A M
authors.initials: AM
authors.lastname: Comeau
authors.forename: D
authors.initials: D
authors.lastname: Wara
authors.forename: G N
authors.initials: GN
authors.lastname: Vyas
mesh_terms: D000163:Acquired Immunodeficiency Syndrome / Q000175:diagnosis / Q000635:transmission*; D000293:Adolescent; D000328:Adult; D004279:DNA, Viral / Q000032:analysis*; D004311:Double-Blind Method; D005260:Female; D005434:Flow Cytometry; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D018445:Infectious Disease Transmission, Vertical; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011446:Prospective Studies; D011533:Proviruses / Q000235:genetics*
chemical_list: D004279:DNA, Viral