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A sensitive branched DNA HIV-1 signal amplification viral load assay with single day turnaround.

ABSTRACT: Branched DNA (bDNA) is a signal amplification technology used in clinical and research laboratories to quantitatively detect nucleic acids. An overnight incubation is a significant drawback of highly sensitive bDNA assays. The VERSANT® HIV-1 RNA 3.0 Assay (bDNA) ("Versant Assay") currently used in clinical laboratories was modified to allow shorter target incubation, enabling the viral load assay to be run in a single day. To dramatically reduce the target incubation from 16-18 h to 2.5 h, composition of only the "Lysis Diluent" solution was modified. Nucleic acid probes in the assay were unchanged. Performance of the modified assay (assay in development; not commercially available) was evaluated and compared to the Versant Assay. Dilution series replicates (>950 results) were used to demonstrate that analytical sensitivity, linearity, accuracy, and precision for the shorter modified assay are comparable to the Versant Assay. HIV RNA-positive clinical specimens (n = 135) showed no significant difference in quantification between the modified assay and the Versant Assay. Equivalent relative quantification of samples of eight genotypes was demonstrated for the two assays. Elevated levels of several potentially interfering endogenous substances had no effect on quantification or specificity of the modified assay. The modified assay with drastically improved turnaround time demonstrates the viability of signal-amplifying technology, such as bDNA, as an alternative to the PCR-based assays dominating viral load monitoring in clinical laboratories. Highly sensitive bDNA assays with a single day turnaround may be ideal for laboratories with especially stringent cost, contamination, or reliability requirements.

SUBMITTER: Baumeister MA

PROVIDER: S-EPMC3314011 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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A sensitive branched DNA HIV-1 signal amplification viral load assay with single day turnaround.

Baumeister Mark A MA Zhang Nan N Beas Hilda H Brooks Jesse R JR Canchola Jesse A JA Cosenza Carlo C Kleshik Felix F Rampersad Vinod V Surtihadi Johan J Battersby Thomas R TR

PloS one 20120327 3

Branched DNA (bDNA) is a signal amplification technology used in clinical and research laboratories to quantitatively detect nucleic acids. An overnight incubation is a significant drawback of highly sensitive bDNA assays. The VERSANT® HIV-1 RNA 3.0 Assay (bDNA) ("Versant Assay") currently used in clinical laboratories was modified to allow shorter target incubation, enabling the viral load assay to be run in a single day. To dramatically reduce the target incubation from 16-18 h to 2.5 h, compo ...[more]

PMID: 22479381

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Project description:IntroductionViral load testing is essential to manage HIV disease, especially in infants and children. Early infant diagnosis is performed using nucleic-acid testing in children under 18 months. Resource-limited health systems face severe challenges to scale-up both viral load and early infant diagnosis to unprecedented levels. Streamlining laboratory systems would be beneficial to improve access to quality testing and to increase efficiency of antiretroviral treatment programmes. We evaluated the performance of viral load testing to serve as an early infant diagnosis assay in children younger than 18 months.MethodsThis study was an observational, prospective study, including children between one and 18 months of age who were born to HIV-positive mothers in 134 health facilities in Maputo City and Maputo Province, Mozambique. Dried blood spot specimens from heel or toe pricks were collected between January and April 2018, processed using SPEX buffer for both assays, and tested for routine EID and VL testing using the Roche CAP/CTM HIV-1 Qualitative v2 and Roche CAP/CTM HIV-1 Quantitative v2 assays respectively. The sensitivity, specificity and positive and negative predictive values were estimated using the EID results as the reference standard.ResultsA total of 1021 infants were included in the study, of which 47% were female. Over 95% of mothers and children were on antiretroviral treatment or received antiretroviral prophylaxis respectively. The sensitivity and specificity of using the viral load assay to detect infection were 100% (95% CI: 96.2 to 100%) and 99.9% (95% CI: 99.4 to 100%). The positive and negative predictive values were 99.0% (95% CI: 94.3 to 100%) and 100% (95% CI: 99.6 to 100%). The McNemar's test was 1.000 and Cohen's kappa was 0.994.ConclusionsThe comparable performance suggests that viral load assays can be used as an infant diagnostic assay. Infants with either low levels of viraemia or high cycle threshold values should be repeat tested to ensure the result is truly positive prior to treatment initiation, regardless of assay used. Viral load assays could replace traditional early infant diagnosis testing, substantially streamlining molecular laboratory services for children and lowering costs, with the additional advantage of providing baseline viral load results for antiretroviral treatment management.

Dataset Information

A sensitive branched DNA HIV-1 signal amplification viral load assay with single day turnaround.

Publications

A sensitive branched DNA HIV-1 signal amplification viral load assay with single day turnaround.

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