World Health Organization collaborative study to calibrate the 3rd International Standard for Hepatitis C virus RNA nucleic acid amplification technology (NAT)-based assays

Background and Objectives A collaborative study was undertaken to evaluate a replacement World Health Organization International Standard for hepatitis C virus (HCV) RNA for nucleic acid amplification technology (NAT)-based assays. The candidate preparations were calibrated in International Units (IUs). Materials and Methods Three new candidate preparations were produced from a single bulk containing anti-HCV-negative, genotype 1a HCV RNA–positive plasma. Two samples were lyophilized (coded Sample 2 and Sample 3), whilst a third (Sample 4) contained liquid/frozen material. The samples were distributed together with the 2nd International Standard (Sample 1, NIBSC code 96/798) for evaluation by thirty-three laboratories, from fourteen countries. The panel of samples were assayed on four separate occasions. Stability studies were performed for the lyophilized samples by accelerated thermal degradation. Results Participants returned data from a wide range of commercial and in-house quantitative and qualitative assays. Twenty-five data sets were returned for quantitative assays and fourteen for qualitative assays. Excellent agreement was observed between laboratories and assay methods. The mean relative potencies of Samples 2–4 were 5·19, 5·41 and 5·70 log10 IU/ml, respectively, when compared against the 2nd International Standard. Samples 2 and 3 demonstrated stability of a similar order to the previous standards. Conclusions Based upon the results of the collaborative study, Sample 2 (code number 06/100) was established as the 3rd International Standard for HCV RNA with an assigned unitage of 5·19 log10 IU/ml. Each vial contains the equivalent of 0·5 ml of material; each vial contains 4·89 log10 IU of HCV RNA.