       Document 0244
 DOCN  M9640244
 TI    DNA probe hybridisation in microwells using a new bioluminescent system
       for the detection of PCR-amplified HIV-1 proviral DNA.
 DT    9604
 AU    Zammatteo N; Moris P; Alexandre I; Vaira D; Piette J; Remacle J;
       Laboratoire de Biochimie Cellulaire, Facultes Notre-Dame de la; Paix,
       Namur, Belgium.
 SO    J Virol Methods. 1995 Oct;55(2):185-97. Unique Identifier : AIDSLINE
       MED/96116854
 AB    A new bioluminescent detection system combined with a sandwich DNA
       hybridisation reaction in microwells has been developed for the
       detection of human immunodeficiency virus type 1 (HIV-1) provirus DNA
       amplified by the polymerase chain reaction (PCR). First, a fragment of
       the HIV-1 gag gene was amplified. The amplified DNA fragments were
       denatured and hybridised to a capture probe immobilised in microwells
       and to a biotinylated detection probe. A streptavidin-pyruvate kinase
       conjugate could then react on the biotinylated probe and the kinase
       activity detected by means of the luciferin-luciferase system, with
       production of a bioluminescent signal. This sandwich assay followed by a
       bioluminescent reaction detected as little as 7 amol of target DNA. The
       bioluminescent assay detected 5 HIV copies generated after one round of
       PCR, even if no band was seen on an agarose gel. The assay was applied
       to the detection of HIV-proviral DNA in peripheral blood mononuclear
       cells after one round of PCR and allowed to clearly identify a positive
       sample as compared to nested PCR.
 DE    Base Sequence  Chemiluminescence  DNA Primers  *DNA Probes  DNA,
       Viral/*ANALYSIS  Gene Products, gag/*GENETICS  Human
       HIV-1/GENETICS/*ISOLATION & PURIF  Leukocytes, Mononuclear/VIROLOGY
       Molecular Sequence Data  *Polymerase Chain Reaction
       Proviruses/GENETICS/ISOLATION & PURIF  Sensitivity and Specificity
       Support, Non-U.S. Gov't  JOURNAL ARTICLE

       SOURCE: National Library of Medicine.  NOTICE: This material may be
       protected by Copyright Law (Title 17, U.S.Code).

