PCR detection of integrated HIV-1 DNA in peripheral blood lymphocytes (PBL) from infected individuals at various stages of disease.

Important note: Information in this article was accurate in 1992. The state of the art may have changed since the publication date.

PCR detection of integrated HIV-1 DNA in peripheral blood lymphocytes (PBL) from infected individuals at various stages of disease.

Abstr Annu Meet Am Soc Microbiol. 1991 May 5-9;91:334 (abstract no. T-5). Unique Identifier : AIDSLINE ASM91/0180292
Casareale D; Kung K; Dragon B; Giron J; Wolfe L; PCR Division, Roche Diagnostic Systems Inc, Fair Lawn, NJ.

Abstract: While available evidence indicates that viremia in HIV-infected individuals is not persistent and is influenced by neutralizing antibody levels, integrated HIV-1 DNA has been shown to be a more consistent marker for prior infection. Although most previous studies did not include terminal AIDS patients, in this study integrated DNA was evaluated by PCR in 50 HIV-1 seropositive individuals, enrolled at various stages of infection including asymptomatic, lymphadenopathy, ARC, AIDS, and terminal AIDS. Seronegative patients attending the same clinic served as controls. PBL, isolated from blood using Ficoll-Hypaque, LeucoPREP, or an RBC lysis procedure, were extracted using Proteinase K. PCR-based amplification of target DNA (1.25 x 10(5) PBL) was performed, using gag region biotinylated primer pairs Bio-SK38/39 and Bio-SK462/431 for 30 thermal cycles. Amplicons were evaluated by a non-isotopic microtiter plate assay with probes specific for each primer pair and by a standard isotopic oligomer hybridization (OH) assay. Assays incorporated the new UNG-restriction based sterilization to prevent carry-over mediated false positive reactions. The non-isotopic assay results agree well with the OH assay and show a direct correlation between PCR results and HIV-1 infection. The details of assay procedures, efficacy of sterilization, and correlation of PCR results to clinical staging will be presented.

Keywords: Acquired Immunodeficiency Syndrome/*BLOOD AIDS-Related Complex/*BLOOD DNA, Viral/*BLOOD Genes, gag Genes, Viral Human HIV Infections/*BLOOD HIV-1/GENETICS/*ISOLATION & PURIF Lymphocytes/*CHEMISTRY Polymerase Chain Reaction ABSTRACT
920830
M9281065

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