key: cord-0703115-kcgfcnpu authors: Lieberman, J.; Pepper, G.; Naccache, S. N.; Huang, M.; Jerome, K. R.; Greninger, A. L. title: Comparison of Commercially Available and Laboratory Developed Assays for in vitro Detection of SARS-CoV-2 in Clinical Laboratories date: 2020-04-27 journal: nan DOI: 10.1101/2020.04.24.20074559 sha: 9982f404763c1e3174fb0bdf5fc261594fc7ed8e doc_id: 703115 cord_uid: kcgfcnpu Multiple laboratory developed tests and commercially available assays have emerged to meet diagnostic needs related to the SARS-CoV-2 pandemic. To date, there is limited comparison data for these different testing platforms. We compared the analytical performance of a laboratory developed test (LDT) developed in our clinical laboratory based on CDC primer sets and four commercially available, FDA emergency use authorized assays for SARS-CoV-2 (Cepheid, DiaSorin, Hologic Panther, and Roche Cobas) on a total of 169 nasopharyngeal swabs. The LDT and Cepheid Xpert Xpress SARS-CoV-2 assays were the most sensitive assays for SARS-CoV-2 with 100% agreement across specimens. The Hologic Panther Fusion, DiaSorin Simplexa, and Roche Cobas 6800 only failed to detect positive specimens near the limit of detection of our CDC-based LDT assay. All assays were 100% specific, using our CDC-based LDT as the gold standard. Our results provide initial test performance characteristics for SARS-CoV-2 RT-PCR and highlight the importance of having multiple viral detection testing platforms available in a public health emergency. Since the first infection with SARS-CoV-2 was detected in the United States in January 29 2020 (1), there has been an exponential growth in cases and deaths (2). At the time of this 30 writing, the US case count exceeds 600,000 with more than 30,000 deaths and considerable 31 geographic heterogeneity (2, 3). Despite social distancing policies, the outbreak of Covid-19, the 32 disease caused by SARS-CoV-2, continues to grow and threatens to overwhelm hospital systems 33 in multiple states (2). 34 The explosion of Covid-19 cases in the United States has highlighted the critical role 35 diagnostic testing plays in medical and public health decision making in containing and 36 mitigating the SARS-CoV-2 pandemic. Reliable test results enable appropriate utilization of 37 scarce hospital resources, including personal protective equipment (PPE) and negative pressure 38 isolation rooms, as well as public health resources for contact tracing or isolation decision-39 making (4). In rapid succession in March 2020, multiple assays have become available including 40 both FDA-EUA test platforms and laboratory developed tests (LDTs) for use in high-complexity 41 clinical laboratories. To solve supply chain difficulties, clinical laboratories have had to 42 implement multiple assays using scarce reagent resources, rendering thorough comparisons 43 challenging. A clear understanding of the analytical parameters of these options is important to 44 help guide assay selection by clinical laboratories when supply chain considerations subside (4). Finally, 28 specimens were used to compare the SARS-CoV-2 assay on the UW Panther Fusion 74 with the DiaSorin Simplexa assay. All same-sample comparisons were performed on specimens 75 stored at 4 o C for less than 72 hours with no freeze-thaws. Inconclusive results (one of two 76 targets detected) were considered positive due to the high specificity of all assays and limited 77 cross-reactivity seen for SARS-CoV-2 primer sets. This work was approved under a consent 78 waiver from the University of Washington Institutional Review Board. CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted April 27, 2020. . Panther Fusion RUO and EUA assays were slightly less sensitive than the CDC-based LDT, 94 missing one positive/inconclusive in each sample set (Table 1a/b). One additional specimen was 95 initially negative with the RUO reagents but was detected upon repeat with the Panther Fusion 96 EUA assay. Discordant specimens were either inconclusive (one target of two detected) or had 97 high average CTs (>37) by the CDC LDT test. All 29 negative specimens generated "Not 98 detected" results by the Hologic Panther Fusion SARS-CoV-2 assay. 99 100 DiaSorin Simplexa SARS-CoV-2. We next compared the DiaSorin Simplexa SARS-CoV-2 assay to 101 our CDC-based LDT. All 19 specimens (11 positives and 8 negatives) demonstrated complete 102 concordance between the two platforms (Table 2a) Simplexa generated one additional positive in the 12 specimens that were negative by the 107 Hologic Panther Fusion (Table 2b ). This discordant specimen was detected by the CDC-based 108 LDT with CTs of 36.8 (N1) and 35.8 (N2), confirming the DiaSorin Simplexa result. 109 110 Roche Cobas SARS-CoV-2. We next compared the Roche Cobas SARS-CoV-2 assay to our CDC 111 LDT. All 20 negatives demonstrated complete concordance between the two platforms (Table 112 3). One of the 20 positives was not detected by the Roche assay. This specimen had CTs of 38.0 113 (N1) and 37.4 (N2) in the LDT. Across the 20 positive specimens, CTs were only slightly higher 114 . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted April 27, 2020. After performing the above pairwise comparisons, we next compared twenty-six specimens (13 119 positive, 13 negative) from another high-complexity hospital laboratory (LabCorp Seattle). All 120 26 were also tested on the Cepheid Xpert Xpress SARS-CoV-2 assay ( Table 4) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted April 27, 2020. . https://doi.org/10. 1101 evaluation that demonstrated high sensitivity of the E-gene and N2 primer sets used by the 137 Cepheid assay (6). The Panther Fusion was somewhat less sensitive than either the LDT or the 138 DiaSorin; however, the Panther Fusion detected SARS-CoV-2 RNA in one specimen that was 139 inconclusive (1 of 2 targets detected, thus presumed positive) by the UW CDC LDT. The Roche 140 assay performed on the Cobas 6800 platform detected 28/30 positive samples; both of these 141 discordant specimens had low viral titers (UW CDC LDT C T >37) and one was the inconclusive 142 specimen. Therefore, we conclude that all the tested assays show good sensitivity for the 143 detection of SARS-CoV-2, with the UW CDC LDT and Cepheid Xpert Xpress SARS-CoV-2 assays 144 having the best and similar sensitivity, followed by the Roche Cobas 6800, DiaSorin Simplexa, 145 and Panther Fusion SARS-CoV-2 assays. 146 Our results are chiefly limited by the small sample sets used to compare these different 147 assays as well as asynchronous comparisons that only allowed for pairwise comparisons early in 148 the pandemic. For instance, these asynchronous panels most greatly affected our CDC LDT 149 versus Hologic Panther Fusion comparison, which had a greater proportion of high CT positive 150 specimens that resulted in a lower measured sensitivity for the Panther Fusion. . In clinical 151 practice, the minor differences in sensitivity are likely to have little effect on Hologic Panther 152 Fusion SARS-CoV-2 assay performance on VTM specimens, given the CT ranges we have 153 observed in our clinical populations. 154 Despite their limitations, these data provide a basis for differences in analytical 155 sensitivity at different CTs that may be seen between platforms. For instance, recent reports 156 have demonstrated a slightly higher analytical sensitivity of the Cepheid Xpert Xpress SARS-157 CoV-2 assay compared to the Roche Cobas SARS-CoV-2 test, and a slightly lower sensitivity of 158 . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted April 27, 2020 . . https://doi.org/10.1101 the DiaSorin Simplexa SARS-CoV-2 assay compared to a modified CDC assay, both of which are 159 concordant with our data (7, 8) . We also note that, while analytical sensitivity is of critical 160 importance, many other considerations factor into assay platform selection including assay 161 availability, cost, turnaround time, and throughput. 162 Our results provide an early assessment of performance characteristics of five separate 163 assays for the detection of SARS-CoV-2. During March 2020, reagent availability for SARS-COV-2 164 RT-PCR assays was heavily constrained, necessitating more limited assay comparisons. All 165 platforms examined here had acceptable performance criteria for testing during the early part 166 of this pandemic. As the supply chain for SARS-CoV-2 RT-PCR attempts to catch up with testing 167 demand, we look forward to additional assay comparison data. 168 169 . CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted April 27, 2020. CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted April 27, 2020 . . https://doi.org/10.1101 Tables Table 1a. is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint this version posted April 27, 2020. . is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted April 27, 2020 . . https://doi.org/10.1101 CC-BY 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. (which was not certified by peer review) The copyright holder for this preprint this version posted April 27, 2020. . First Case of 173 2019 Novel Coronavirus in the United States COVID-19 health service utilization forecasting team ICU-days, ventilator-days and deaths by US state in 176 the next 4 months An interactive web-based dashboard to track COVID-19 in 178 real time SARS-CoV-2 Testing Detection of 2019 novel coronavirus (2019-nCoV) by 184 real-time RT-PCR