Document 0532
 DOCN  M9460532
 TI    Amplification and detection of a single molecule of human
       immunodeficiency virus RNA.
 DT    9404
 AU    Nakamura S; Katamine S; Yamamoto T; Foung SK; Kurata T; Hirabayashi Y;
       Shimada K; Hino S; Miyamoto T; Department of Bacteriology, Nagasaki
       University School of; Medicine, Japan.
 SO    Virus Genes. 1993 Dec;7(4):325-38. Unique Identifier : AIDSLINE
       MED/94167907
 AB    Detection of plasma viremia in human immunodeficiency virus type 1
       (HIV-1)-infected people is indispensable for the diagnosis of
       seronegative infection as well as for the evaluation of virus activities
       in vivo. The direct detection of HIV-1 RNA in circulation has been
       performed by means of reverse transcription followed by polymerase chain
       reaction (RT-PCR). As an attempt to establish a highly sensitive assay,
       we evaluated the effects of two-step amplification with nested primers
       and double priming of reverse transcription on the sensitivity of
       RT-PCR. The sensitivity of two-step amplification was 100 times higher
       than that of one-step amplification. The double priming of reverse
       transcription further increased the sensitivity of the following
       two-step amplification 100 times, which appeared to be enough to detect
       HIV-1 RNA from as little as a 2.2 x 10(-4) TCID50 unit equivalent of
       culture supernatant of HIV-1-infected cells and a single molecule of
       HIV-1 gag complementary RNA synthesized by in vitro transcription. By
       use of this most sensitive assay, we successfully detected HIV-1 RNA in
       serum or plasma from all 22 patients with acquired immune deficiency
       syndrome (AIDS) or AIDS-related complex (ARC) and 13 out of 14 untreated
       asymptomatic carriers. Of 43 asymptomatic carriers under the treatment
       with interferon-alpha or azidothymidine, 17 cases showed negative
       results, indicating that the virus activity was suppressed by the
       therapeutics. We also noted the inhibitory effect of heparin on RT-PCR.
 DE    Base Sequence  Cells, Cultured  DNA, Viral  Gene Products, gag/GENETICS
       Human  HIV Infections/DIAGNOSIS/*MICROBIOLOGY  HIV
       Seropositivity/DIAGNOSIS/MICROBIOLOGY  HIV-1/GENETICS/*ISOLATION & PURIF
       Molecular Sequence Data  Polymerase Chain Reaction/*METHODS  RNA,
       Viral/*ANALYSIS/BLOOD  Sensitivity and Specificity  Support, Non-U.S.
       Gov't  Viremia/DIAGNOSIS/*MICROBIOLOGY  JOURNAL ARTICLE

       SOURCE: National Library of Medicine.  NOTICE: This material may be
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