Evaluation of the Epstein-Barr virus R-gene quantification kit in whole blood with different extraction methods and PCR platforms.
Reliable real-time quantitative PCR assays to measure Epstein-Barr virus (EBV) DNA load (EBV) are useful for monitoring EBV-associated diseases. We evaluated a new commercial kit, EBV R-gene Quantification kit (Argene, Varilhes, France) to quantify EBV DNA load in whole blood. Assay performance was assessed with two PCR platforms (LightCycler 2.0 and SmartCycler 2.0) and three commercial DNA extraction methods. The assay was compared with our in-house real-time EBV PCR using samples from the Quality Control for Molecular Diagnostics 2006 EBV proficiency program and using 167 whole-blood specimens from individuals with infectious mononucleosis, from transplanted or HIV-infected patients, and from EBV-seropositive healthy carriers. The EBV R-gene assay was sensitive to 500 copies of EBV DNA per milliliter of whole blood with the two PCR platforms and the three extraction methods and was linear across 4 orders of magnitude. Intra- and interassay coefficients of variations were less than 20%. Nine of 10 samples tested with the EBV R-gene were in agreement with the expected qualitative results of the Quality Control for Molecular Diagnostics 2006 EBV proficiency program, and 7 of 10 samples were within +/-0.5 log units of the expected quantitative values, with discrepant results mostly observed for low viral load (ie, <1000 copies/ml). In the clinical specimens, the correlation between the R-gene assay and the in-house PCR was high (r=0.92). In conclusion, the EBV R-gene assay accurately assesses the EBV DNA load in whole blood of patients with various forms of EBV infections.
Fafi-Kremer
Samira
S
Laboratoire de Virologie Médicale, Centre Hospitalier Universitaire, Université Joseph Fourier, Grenoble, France.
Morand
Patrice
P
Barranger
Come
C
Barguès
Gérard
G
Magro
Stéphane
S
Bés
Jérôme
J
Bourgeois
Philippe
P
Joannes
Martine
M
Seigneurin
Jean-Marie
JM
D004279:DNA, Viral / Q000097:blood* / Q000235:genetics / Q000302:isolation & purification*; D020031:Epstein-Barr Virus Infections / Q000175:diagnosis; D005814:Genes, Viral; D004854:Herpesvirus 4, Human / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D019562:Viral Load
D004279:DNA, Viral; D011933:Reagent Kits, Diagnostic