title: Multi-Site PCR-based CMV viral load assessment-assays demonstrate linearity and precision, but lack numeric standardization: a report of the association for molecular pathology.
abstract: Viralload (VL) assessment of cytomegalovirus (CMV) by real-time PCR is an important tool for diagnosing and monitoring CMV viremia in patients with compromised immune systems. We report results from a sample exchange organized by members of the Association for Molecular Pathology that compared PCR results from 23 laboratories; 22 such laboratories used a laboratory-developed real-time PCR assay and one laboratory used a competitive PCR assay. The samples sent to each laboratory were comprised of a dilution panel of CMV virion-derived reference materials that ranged from 0 to 500,000 copies/ml. Accuracy, linearity, and intralaboratory precision were established for the different laboratory-developed assays. Overall, PCR results were linear for each laboratory (R(2) > 0.97 in all but two). While 13 laboratories showed no significant quantitative assay bias, 10 laboratories reported VLs that were significantly different compared with expected values (bias range, -0.82 to 1.4 logs). The intralaboratory precision [mean coefficient of variance of 2% to 5% (log-scale)] suggested that changes in VLs of less than 3- to fivefold may not be significantly different. There was no significant association between laboratory-specific technical variables (PCR platform, calibrator, extraction method) and assay linearity or accuracy. These data suggested that, within each laboratory, relative VL values were linear, but additional method standardization and a CMV DNA reference standard are needed to allow laboratories to achieve comparable numeric results.
authors.affiliation: CMV Working Group of the Association for Molecular Pathology Clinical Practice Committee, Medical University of South Carolina, USA. wolffd@musc.edu
authors.forename: Daynna J
authors.initials: DJ
authors.lastname: Wolff
authors.forename: Denise Lamarche
authors.initials: DL
authors.lastname: Heaney
authors.forename: Paul D
authors.initials: PD
authors.lastname: Neuwald
authors.forename: Kathleen A
authors.initials: KA
authors.lastname: Stellrecht
authors.forename: Richard D
authors.initials: RD
authors.lastname: Press
mesh_terms: D003587:Cytomegalovirus / Q000235:genetics*; D003586:Cytomegalovirus Infections / Q000175:diagnosis*; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D016014:Linear Models; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D012680:Sensitivity and Specificity; D019562:Viral Load / Q000379:methods* / Q000592:standards
chemical_list: D004247:DNA