International collaborative study on the second EUROHEP HCV-RNA reference panel.
Eighty-six laboratories participated in a collaborative study and tested the second EUROHEP HCV-RNA reference panel. The coded panel comprised 4 HCV-RNA positive plasma samples (one weak positive), 6 HCV-RNA negative plasma samples and two dilution series of HCV-RNA genotype 1 and 3 plasma standards. The 86 laboratories submitted 136 coded data forms for evaluation. Of these data sets 99 were tested using a PCR assay developed in-house, 28 using a commercially available HCV-PCR test (AMPLICOR, Roche Diagnostic Systems) and 9 using other amplification methods. Twenty-two data forms (16%) had faultless results, 39 (29%) missed the weak positive sample only and 75 data sets (55%) had false positive and/or false negative results. Participants using the commercial HCV-PCR test tended to reach a sufficient quality score more often than investigators using assays developed in-house (64% versus 45%, P = 0.11). The UNG system in the commercial HCV-PCR test did not prevent five laboratories generating false-positive results in the 6 HCV-RNA negative samples. Among the laboratories with satisfactory results, up to 10000-fold differences in sensitivity were observed in the dilution series. The 50% and 90% laboratories detection endpoints in the dilution series of the HCV genotype 1 plasma standard were approximately 600 genome equivalents per ml (geq/ml) and 7750 geq/ml according to a standard applied in a signal amplification assay (bDNA, Chiron). Our results suggest that the detection efficiency for genotype 3 by commercial HCV-RNA assays is lower than by the in-house assays. Internationally characterized HCV-RNA plasma standards should be made available for validation and standardization of HCV-RNA assays for HCV diagnosis and virological safety testing of blood products.
Damen
M
M
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, Viral Diagnostic Laboratory, Amsterdam, Netherlands.
Cuypers
H T
HT
Zaaijer
H L
HL
Reesink
H W
HW
Schaasberg
W P
WP
Gerlich
W H
WH
Niesters
H G
HG
Lelie
P N
PN
D016174:Hepacivirus / Q000235:genetics / Q000302:isolation & purification*; D006526:Hepatitis C / Q000097:blood / Q000821:virology*; D006801:Humans; D007391:International Cooperation; D016133:Polymerase Chain Reaction / Q000379:methods / Q000592:standards*; D012367:RNA, Viral / Q000032:analysis*; D012015:Reference Standards; D012680:Sensitivity and Specificity
D012367:RNA, Viral