Performance characteristics of the QUANTIPLEX HIV-1 RNA 3.0 assay for detection and quantitation of human immunodeficiency virus type 1 RNA in plasma.
The QUANTIPLEX HIV-1 RNA assay, version 3.0 (a branched DNA, version 3.0, assay [bDNA 3.0 assay]), was evaluated by analyzing spiked and clinical plasma samples and was compared with the AMPLICOR HIV-1 MONITOR Ultrasensitive (ultrasensitive reverse transcription-PCR [US-RT-PCR]) method. A panel of spiked plasma samples that contained 0 to 750,000 copies of human immunodeficiency virus type 1 (HIV-1) RNA per ml was tested four times in each of four laboratories (1,344 assays). Negative results (<50 copies/ml) were obtained in 30 of 32 (94%) assays with seronegative samples, 66 of 128 (52%) assays with HIV-1 RNA at 50 copies/ml, and 5 of 128 (4%) assays with HIV-1 RNA at 100 copies/ml. The assay was linear from 100 to 500,000 copies/ml. The within-run standard deviation (SD) of the log(10) estimated HIV-1 RNA concentration was 0.08 at 1,000 to 500,000 copies/ml, increased below 1,000 copies/ml, and was 0.17 at 100 copies/ml. Between-run reproducibility at 100 to 500 copies/ml was <0.10 log(10) in most comparisons. Interlaboratory differences across runs were </=0.10 log(10) at all concentrations examined. A subset of the panel (25 to 500 copies/ml) was also analyzed by the US-RT-PCR assay. The within-run SD varied inversely with the log(10) HIV-1 RNA concentration but was higher than the SD for the bDNA 3.0 assay at all concentrations. Log-log regression analysis indicated that the two methods produced very similar estimates at 100 to 500 copies/ml. In parallel testing of clinical specimens with low HIV-1 RNA levels, 80 plasma samples with <50 copies/ml by the US-RT-PCR assay had <50 copies/ml when they were retested by the bDNA 3.0 assay. In contrast, 11 of 78 (14%) plasma samples with <50 copies/ml by the bDNA 3.0 assay had >/=50 copies/ml when they were retested by the US-RT-PCR assay (median, 86 copies/ml; range, 50 to 217 copies/ml). Estimation of bDNA 3.0 values of <50 copies/ml by extending the standard curve of the assay showed that these samples with discrepant results had higher HIV-1 RNA levels than the samples with concordant results (median, 34 versus 17 copies/ml; P = 0.0051 by the Wilcoxon two-sample test). The excellent reproducibility, broad linear range, and good sensitivity of the bDNA 3.0 assay make it a very attractive method for quantitation of HIV-1 RNA levels in plasma.
Department of Laboratory Medicine & Pathology Division of Infectious Diseases, University of Minnesota, Minneapolis, Minnesota 55455, USA. erice001@tc.umn.edu
A
A
Erice
D
D
Brambilla
J
J
Bremer
J B
JB
Jackson
R
R
Kokka
B
B
Yen-Lieberman
R W
RW
Coombs
D015342:DNA Probes; D005069:Evaluation Studies as Topic; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D006801:Humans; D007753:Laboratories / Q000592:standards; D009693:Nucleic Acid Hybridization / Q000379:methods*; D012367:RNA, Viral / Q000097:blood*; D011933:Reagent Kits, Diagnostic / Q000592:standards; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D014766:Viremia / Q000821:virology
D015342:DNA Probes; D012367:RNA, Viral; D011933:Reagent Kits, Diagnostic