title: Detection of cytomegalovirus in whole blood using three different real-time PCR chemistries.
abstract: Several different primer-probe chemistries have been produced commercially for real-time PCR detection and quantification of cytomegalovirus, but there are few studies evaluating their relative performance. We assessed three such commercial reagents with respect to analytical and clinical operating characteristics. The samples included 149 clinical whole blood specimens that were de-identified and assayed in parallel with all primer-probe systems. Individual methods used TaqMan, dual fluorescence resonance energy transfer hybridization probes, and labeled primer chemistries. Method comparability was determined both qualitatively, based on pair-wise assessment of concordance, and quantitatively, based on pair-wise linear regression analysis. Analytical sensitivity and the lower end of the linear dynamic range reached 10 target copies per reaction for the TaqMan and labeled primer systems and 100 target copies per reaction for the dual fluorescence resonance energy transfer probe system. Quantitative linearity reached an upper limit of 10(5) copies per reaction for all methods. No assay cross-reactivity was seen with other common viral pathogens (100% analytical specificity). Pair-wise analysis of qualitative results from clinical samples showed no significant differences in sensitivity between the three sets of reagents, and linear regression analysis indicated that the quantitative values achieved were comparable in all positive specimens. The findings demonstrate that similar analytical and clinical performance characteristics can be demonstrated for quantitative detection of cytomegalovirus in clinical whole blood extracts using a wide variety of real-time PCR chemistries.
authors.affiliation: Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee.
authors.forename: Erica
authors.initials: E
authors.lastname: Vincent
authors.affiliation: Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee.
authors.forename: Zhengming
authors.initials: Z
authors.lastname: Gu
authors.affiliation: Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee.
authors.forename: Markus
authors.initials: M
authors.lastname: Morgenstern
authors.affiliation: Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee.
authors.forename: Candace
authors.initials: C
authors.lastname: Gibson
authors.affiliation: Department of Biostatistics, St. Jude Children's Research Hospital, Memphis, Tennessee.
authors.forename: Jianmin
authors.initials: J
authors.lastname: Pan
authors.affiliation: Department of Pathology, St. Jude Children's Research Hospital, Memphis, Tennessee. Electronic address: Randall.Hayden@stjude.org.
authors.forename: Randall T
authors.initials: RT
authors.lastname: Hayden
mesh_terms: D003587:Cytomegalovirus / Q000235:genetics / Q000302:isolation & purification*; D003586:Cytomegalovirus Infections / Q000097:blood / Q000175:diagnosis*; D004279:DNA, Viral / Q000097:blood; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D006801:Humans; D007202:Indicators and Reagents; D016133:Polymerase Chain Reaction / Q000379:methods*; D012044:Regression Analysis; D012680:Sensitivity and Specificity
chemical_list: D004279:DNA, Viral; D007202:Indicators and Reagents