Verification and Validation of SARS-CoV-2 Assay Performance on the Abbott m 2000 and Alinity m Systems

Abstract

We verified the analytical performance of the Abbott RealTime SARS-CoV-2 assay on the m2000 system and compared its clinical performance to the CDC 2019-nCoV Real-Time PCR Diagnostic Panel and the ThermoFisher TaqPath RT-PCR COVID-19 kit. We also performed a bridging study comparing the RealTime SARS-CoV-2 assay with the new Abbott Alinity m SARS-CoV-2 assay. A number of standards, reference materials, and commercially available controls were used for the analytical verification to confirm the limit of detection, linearity, and reproducibility. We used nasopharyngeal (NP) swab specimens collected in saline for the clinical verification and bridging studies. Overall, we found 91.2% positive percent agreement (PPA) (95% CI 76.2 to 98.14%) and a 100% negative percent agreement (NPA) (95% CI 97.97 to 100%) between the results of the RealTime SARS-CoV-2 and CDC tests with 217 NP specimens (P=0.13). We found a PPA of 100% (95% CI 90.26 to 100%) and a NPA of 95.15% (95% CI 83.47 to 99.4%) between the results of the RealTime and TaqPath tests with 77 NP specimens (P=0.24). Finally, we tested 203 NP swab specimens for SARS-CoV-2 on the m2000 on the Alinity m systems. The PPA and NPA were 92.2% (95% CI, 85.3 to 96.59%) and 92% (95% PI, 84.8 to 96.5%), respectively (P=0.4). Although cycle number (Cn) values obtained for the concordant positive samples were highly correlated (R2, 0.95), the Cn values were on average 14.14 higher on the Alinity m system due to the unread cycles with the RealTime SARS-CoV-2 assay.

Real-Life Head-to-Head Comparison of Performance of Two High-Throughput Automated Assays for the Detection of SARS-CoV-2 RNA in Nasopharyngeal Swabs

“…In 18/37 individuals with Alinity m SARS-CoV-2 / cobas 6800 SARS-CoV-2 discrepant results, previous and/or follow-up SARS-CoV-2 PCR result(s) were identified: 10/18 had a recorded previous SARS-CoV-2 PCR positive result in samples collected 0, 6, 8, 9, 9, 10, 11, 27, 59, 90 days (mean: 22.9 days; median: 9.5 days; range 0-90 days) before the sample in this study was collected (for multiple previous samples identified, the first previously positive sample was considered for each individual) and 8/18 had a recorded follow-up SARS-CoV-2 PCR positive result in samples collected 0, 0, 0, 2, 2, 5, 6, 7 days (mean: 2.8 days; median: 2 days; range 0-7 days) after the sample in this study was collected (for multiple follow-up samples identified, the first positive follow-up sample was considered for each individual). virus DNA; 13,17,19,21 (ii) quantitative detection of hepatitis C virus RNA; 12,17,18,21 (iii) quantitative detection of HIV-1 RNA; [15][16][17]21 (iv) qualitative detection of SARS-CoV-2 RNA; 20 (v) qualitative detection of 14 high-risk human papillomaviruses coupled with extended genotyping; 14,22 (vi) qualitative detection and differentiation of Chlamydia trachomatis, Trichomonas vaginalis, Mycoplasma genitalium, and Neisseria gonorrhoeae 23 and (vii) qualitative detection and differentiation of SARS-CoV-2, influenza A and B viruses, and respiratory syncytial virus. Twelve evaluations of Alinity m assays are available in peerreviewed literature, [12][13][14][15][16][17][18][19][20][21][22][23] but only a single analytical and clinical evaluation of Alinity m SARS-CoV-2 assay on a limited number of samples is known to have been published to date.…”

“…virus DNA; 13,17,19,21 (ii) quantitative detection of hepatitis C virus RNA; 12,17,18,21 (iii) quantitative detection of HIV-1 RNA; [15][16][17]21 (iv) qualitative detection of SARS-CoV-2 RNA; 20 (v) qualitative detection of 14 high-risk human papillomaviruses coupled with extended genotyping; 14,22 (vi) qualitative detection and differentiation of Chlamydia trachomatis, Trichomonas vaginalis, Mycoplasma genitalium, and Neisseria gonorrhoeae 23 and (vii) qualitative detection and differentiation of SARS-CoV-2, influenza A and B viruses, and respiratory syncytial virus. Twelve evaluations of Alinity m assays are available in peerreviewed literature, [12][13][14][15][16][17][18][19][20][21][22][23] but only a single analytical and clinical evaluation of Alinity m SARS-CoV-2 assay on a limited number of samples is known to have been published to date. 20 In that study, analytical verification of Alinity m SARS-CoV-2 confirmed the manufacturer's limit of detection claim of 100 copies/mL, with clinical evaluation performed on 203 residual nasopharyngeal swabs from symptomatic and asymptomatic individuals with suspected SARS-CoV-2 infection.…”

“…Twelve evaluations of Alinity m assays are available in peerreviewed literature, [12][13][14][15][16][17][18][19][20][21][22][23] but only a single analytical and clinical evaluation of Alinity m SARS-CoV-2 assay on a limited number of samples is known to have been published to date. 20 In that study, analytical verification of Alinity m SARS-CoV-2 confirmed the manufacturer's limit of detection claim of 100 copies/mL, with clinical evaluation performed on 203 residual nasopharyngeal swabs from symptomatic and asymptomatic individuals with suspected SARS-CoV-2 infection. Samples were comparatively tested using Alinity m SARS-CoV-2 and RealTime SARS-CoV-2 assay (Abbott), and positive and negative percent agreement between both assays of 92.2% (95%CI: 85.3-96.6%) and 92.0% (95%CI: 84.8-96.5%), respectively, were observed.…”

“…Alinity m (Abbott Molecular, Des Plaines, IL, USA) is another recently launched fully integrated, automated sample-to-result molecular analyzer allowing continuous loading of samples and random-access testing. Seven molecular assays have been developed for use with Alinity m, [12][13][14][15][16][17][18][19][20][21][22][23] including an assay for SARS-CoV-2, which received U.S. FDA EUA on 11 May J o u r n a l P r e -p r o o f aliquots were prepared: 800 µl for Alinity m SARS-CoV-2 testing and 700 µl for cobas 6800 SARS-CoV-2 testing.…”

“…The Alinity m SARS-CoV-2 assay is a real-time reverse transcriptase (RT) PCR-based assay for the qualitative detection of SARS-CoV-2 RNA in nasal, nasopharyngeal, and oropharyngeal swabs. 20 By using target-specific fluorescent-labeled oligonucleotide probes, the assay allows simultaneous detection and amplification of two SARS-CoV-2-specific sequences targeting the RdRp gene and N gene reported as a combined signal in one channel and individually an internal control (IC) target sequence for evaluation of sample extraction and amplification efficiency reported separately in another channel. The assay is performed on Alinity m-a fully integrated, automated molecular analyzer that allows continuous loading of samples and performs sample preparation, RT-PCR assembly, amplification, detection, calculation, and reporting of the results.…”