Detection of cytomegalovirus (CMV) DNA in EDTA whole-blood samples: evaluation of the quantitative artus CMV LightCycler PCR kit in conjunction with automated sample preparation.
Whole blood has been found to be a reliable matrix for the detection and quantitation of cytomegalovirus (CMV) DNA. In this study, the performance of the artus CMV LightCycler (LC) PCR kit in conjunction with automated sample preparation on a BioRobot EZ1 workstation was evaluated. The accuracy, linearity, analytical sensitivity, and inter- and intra-assay variations were determined. A total of 102 clinical EDTA whole-blood samples were investigated, and results were compared with those obtained with the in vitro diagnostics (IVD)/Conformité Européene (CE)-labeled CMV HHV6,7,8 R-gene quantification kit. When the accuracy of the new kit was tested, seven of eight results were found to be within +/-0.5 log(10) unit of the expected panel results. Determination of linearity resulted in a quasilinear curve over more than 5 log units. The lower limit of detection of the assay was determined to be 139 copies/ml in EDTA whole blood. The interassay variation ranged from 15 to 58%, and the intra-assay variation ranged from 7 to 35%. When clinical samples were tested and the results were compared with those of the routinely used IVD/CE-labeled assay, 53 samples tested positive and 13 samples tested negative by both of the assays. One sample was found to be positive with the artus CMV LC PCR kit only, and 35 samples tested positive with the routinely used assay only. The majority of discrepant results were found with low-titer samples. In conclusion, use of the artus CMV LC PCR kit in conjunction with automated sample preparation on the BioRobot EZ1 workstation may be suitable for the detection and quantitation of CMV DNA in EDTA whole blood in the routine low-throughput laboratory; however, low-positive results may be missed by this assay.
Michelin
Birgit D A
BD
Institute of Hygiene, Medical University of Graz, Universitaetsplatz 4, 8010 Graz, Austria.
Hadzisejdic
Ita
I
Bozic
Michael
M
Grahovac
Maja
M
Hess
Markus
M
Grahovac
Blazenka
B
Marth
Egon
E
Kessler
Harald H
HH
D000925:Anticoagulants; D001331:Automation; D003587:Cytomegalovirus / Q000235:genetics / Q000302:isolation & purification*; D003586:Cytomegalovirus Infections / Q000821:virology*; D004279:DNA, Viral / Q000097:blood*; D004492:Edetic Acid; D006801:Humans; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods*; D011933:Reagent Kits, Diagnostic; D015203:Reproducibility of Results
D000925:Anticoagulants; D004279:DNA, Viral; D011933:Reagent Kits, Diagnostic; D004492:Edetic Acid