ABSTRACT
Experimental conditions affecting the successful propagation of HIV-1 from the plasma of seropositive individuals were examined. It was determined that whole blood samples collected with lithium heparin as the anticoagulant, immediate plasma separation, and immediate culturing were best suited for obtaining viable virus from plasma. Virus was isolated by infecting fresh phytohemagglutinin-stimulated normal donor peripheral blood mononuclear cells (PBMCs) with plasma followed by weekly cocultivation with new target cells. The plasma virus isolation rate was the greatest and HIV-1 titers were the highest for those individuals with less than 200 CD4+ cells/mm3 and decreased as the level of CD4+ cells approached normal values. We were able to obtain positive cultures from 29.5% of those patients with CD4+ counts greater than 500/mm3. HIV-1 titers in plasma also correlated with high serum p24 antigen levels when serum was treated with glycine to dissociate antigen-antibody complexes.
Subject(s)
HIV Seropositivity/microbiology , HIV-1/isolation & purification , Adult , Blood Coagulation Tests , CD4-Positive T-Lymphocytes/cytology , Cells, Cultured , HIV Core Protein p24/blood , HIV Seropositivity/blood , HIV-1/growth & development , Humans , Leukocyte Count , Leukocytes, Mononuclear/microbiology , Random AllocationABSTRACT
A branched DNA (bDNA)-based quantitation of plasma human immunodeficiency virus type 1 (HIV-1) RNA was used to monitor the virologic status of 102 patients (29-906 CD4 cells/mm3) enrolled in clinical trials of antiretroviral and immune-based therapies. Virion-associated RNA was measurable in plasma of 74% of patients tested (10,000-10,000,000 RNA equivalents/mL). Virus levels measured by the bDNA assay exceeded titers obtained by quantitative plasma culture and were inversely correlated (r = -.378; P < .05) with total CD4 cell counts. The assay was used to demonstrate a significant decline (mean, 5-fold; range, 0- to 30-fold), relative to pretreatment, in virus load after beginning antiviral therapy and a transient increase (mean, 15-fold; range, 2- to 50-fold) after treatment with interleukin-2. The decrease in RNA was more dramatic than changes in serum p24 antigen. The bDNA assay yields reproducible results, is relatively easy, and should be useful in measuring HIV-1 RNA in patients in clinical trials.