A branched DNA signal amplification assay for quantification of nucleic acid targets below 100 molecules/ml.
The branched DNA hybridization assay has been improved by the inclusion of the novel nucleotides, isoC and isoG, in the amplification sequences to prevent non-specific hybridization. The novel isoC, isoG-containing amplification sequences have no detectable interaction with any natural DNA sequence. The control of non-specific hybridization in turn permits increased signal amplification. Addition of a 14 site preamplifier was found to increase the signal/noise ratio 8-fold. A set of 74 oligonucleotide probes was designed to the consensus HIV POL sequence. The detection limit of this new HIV branched DNA amplifier assay was approximately 50 molecules/ml. The assay was used to measure viral load in 87 plasma samples of HIV- infected patients on triple drug therapy whose RNA titers were <500 molecules/ml. In all 11 patients viral load eventually declined to below the detection limit with the new assay.
Collins
M L
ML
Chiron Diagnostics, 4560 Horton Street, Emeryville, CA 94608, USA.
Irvine
B
B
Tyner
D
D
Fine
E
E
Zayati
C
C
Chang
C
C
Horn
T
T
Ahle
D
D
Detmer
J
J
Shen
L P
LP
Kolberg
J
J
Bushnell
S
S
Urdea
M S
MS
Ho
D D
DD
D000241:Adenosine; D019380:Anti-HIV Agents / Q000627:therapeutic use; D003562:Cytidine / Q000737:chemistry; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004279:DNA, Viral / Q000032:analysis; D004359:Drug Therapy, Combination; D006151:Guanosine / Q000737:chemistry; D006678:HIV; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology; D006801:Humans; D007546:Isoquinolines / Q000627:therapeutic use; D019259:Lamivudine / Q000627:therapeutic use; D019888:Nelfinavir; D009693:Nucleic Acid Hybridization / Q000379:methods*; D012367:RNA, Viral / Q000032:analysis; D013451:Sulfonic Acids / Q000627:therapeutic use; D015215:Zidovudine / Q000627:therapeutic use
D019380:Anti-HIV Agents; D004279:DNA, Viral; D007546:Isoquinolines; D012367:RNA, Viral; D013451:Sulfonic Acids; D006151:Guanosine; C008184:isoguanosine; D019259:Lamivudine; C083576:isocytidine; D015215:Zidovudine; D003562:Cytidine; D004247:DNA; D019888:Nelfinavir; D000241:Adenosine