Performance characteristics for the quantitation of plasma HIV-1 RNA using branched DNA signal amplification technology.
Highly sensitive assays that quantitate human immunodeficiency virus type 1 (HIV-1) RNA may be valuable for clinical research and the treatment of HIV-1-infected patients. In this study we evaluated the reproducibility and accuracy of the first-generation branched DNA (bDNA-1.0) signal amplification assay under conditions that are relevant to routine use in a clinical context. We show that the bDNA-1.0 assay was able to discern two- to three-fold changes in plasma HIV-1 RNA levels as significant. Reverse transcription coupled to polymerase chain reaction (RT-PCR) was less reproducible and required a 3.7- to 5.8-fold change in plasma HIV-1 RNA levels to be statistically significant. The accuracy of the bDNA-1.0 assay in RNA quantitation was not affected by HIV-1 genotypic variation or by the presence of hemoglobin, bilirubin, lipemia, or any of a dozen therapeutic drugs. Using the bDNA-1.0 assay, we show that HIV-1 RNA levels in plasma specimens were stable when stored at -80 degrees C and were able to withstand at least three freeze-thaw cycles without significant loss. We also examined the performance of an ultrasensitive bDNA assay with improvements to the signal amplification technology. The ultrasensitive bDNA assay displayed a quantitation limit of approximately 500 RNA Eq/ml, yet maintained a dynamic quantitation range up to 1.6 x 10(6) RNA Eq/ml. Like the bDNA-1.0 assay, the ultrasensitive bDNA assay was not affected by HIV-1 genotype variability.
Todd
J
J
Chiron Corporation, Emeryville, CA 94608, USA.
Pachl
C
C
White
R
R
Yeghiazarian
T
T
Johnson
P
P
Taylor
B
B
Holodniy
M
M
Kern
D
D
Hamren
S
S
Chernoff
D
D
D000925:Anticoagulants / Q000592:standards; D000998:Antiviral Agents / Q000494:pharmacology; D001663:Bilirubin / Q000097:blood; D001793:Blood Preservation; D004279:DNA, Viral / Q000032:analysis; D005784:Gene Amplification; D014644:Genetic Variation; D005838:Genotype; D015497:HIV-1 / Q000235:genetics*; D006461:Hemolysis; D006801:Humans; D008055:Lipids / Q000097:blood; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000097:blood*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013048:Specimen Handling; D014158:Transcription, Genetic
D000925:Anticoagulants; D000998:Antiviral Agents; D004279:DNA, Viral; D008055:Lipids; D012367:RNA, Viral; D001663:Bilirubin