Verification of the Roche cobas® 6800 PCR 200 µl and 500 µl protocols for the quantification of HIV-1 RNA, HBV DNA and HCV RNA and evaluation with COBAS® Ampliprep/COBAS® TaqMan® assays.

Abstract

The analytical performance of the cobas 6800 HIV-1, HBV and HCV assays was verified and evaluated to the COBAS Ampliprep/COBAS TaqMan assays. The precision, limit of detection (LoD), limit of quantification (LoQ) and linearity were verified using pooled residual clinical samples. The analytical specificity was verified with negative plasma. HIV-1 analytical reactivity was verified with WHO reference preparations. Accuracy was verified using EQA plasma panels. Evaluation of the equipment was performed using prospectively collected clinical whole blood samples. Excellent precision was demonstrated using both testing protocols (coefficient of variation ≤15 %). The LoDs using the 500 and 200 µl protocols were 20 and 50 cp ml-1 for HIV-1, 10 and 20 IU ml-1 for HBV and 15 and 40 IU ml-1 for HCV, respectively. The LoQs were 40 and 100 cp ml-1 for HIV-1, 20 and 25 IU ml-1 for HBV and 30 and 80 IU ml-1 for HCV, respectively. Assays demonstrated good linearity (R2 ≥0.96). The analytical specificity of the assays was 100 %. There was excellent agreement between the cobas 6800 and CAP/CTM assays (kappa>0.94). The sensitivity, specificity, positive predictive value and negative predictive value for each of the assays were ≥99 %. The cobas HIV-1 and HCV mean quantifications were 0.03 log10 cp ml-1 and 0.17 log10 IU ml-1 higher than the CAP/CTM. The cobas HBV mean quantification was 0.17 log10 IU ml-1 lower than the CAP/CTM. Subtype/genotype specific differences were not observed. Cobas 6800 equipment and assays demonstrated excellent performance and correlated well with CAP/CTM assay results.