title: 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.
authors.affiliation: Siemens Healthcare Diagnostics Inc., Berkeley, California, United States of America.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Baumeister
authors.forename: Nan
authors.initials: N
authors.lastname: Zhang
authors.forename: Hilda
authors.initials: H
authors.lastname: Beas
authors.forename: Jesse R
authors.initials: JR
authors.lastname: Brooks
authors.forename: Jesse A
authors.initials: JA
authors.lastname: Canchola
authors.forename: Carlo
authors.initials: C
authors.lastname: Cosenza
authors.forename: Felix
authors.initials: F
authors.lastname: Kleshik
authors.forename: Vinod
authors.initials: V
authors.lastname: Rampersad
authors.forename: Johan
authors.initials: J
authors.lastname: Surtihadi
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Battersby
mesh_terms: D021121:Branched DNA Signal Amplification Assay / Q000379:methods*; D015658:HIV Infections / Q000175:diagnosis* / Q000821:virology*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods; D016133:Polymerase Chain Reaction; D011933:Reagent Kits, Diagnostic / Q000592:standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013997:Time Factors; D019562:Viral Load
chemical_list: D011933:Reagent Kits, Diagnostic