Reliable quantification of the antibody response to SARS-CoV-2 is highly relevant, e.g., for identifying possible vaccine failure and estimating the time of protection. We compared the performance of five CE marked tests that quantify antibodies against the viral spike protein.

Background In the context of the COVID-19 pandemic, numerous new serological test systems for the detection of anti-SARS-CoV-2 antibodies rapidly have become available. However, the clinical performance of many of these is still insufficiently described. Therefore, we compared three commercial, CE-marked, SARS-CoV-2 antibody assays side by side. Methods We included a total of 1,154 specimens from pre-COVID-19 times and 65 samples from COVID-19 patients (≥14 days after symptom onset) to evaluate the test performance of SARS-CoV-2 serological assays by Abbott, Roche, and DiaSorin. Results All three assays presented with high specificities: 99.2% (98.6-99.7) for Abbott, 99.7% (99.2-100.0) for Roche, and 98.3% (97.3-98.9) for DiaSorin. In contrast to the manufacturers’ specifications, sensitivities only ranged from 83.1% to 89.2%. Although the three methods were in good agreement (Cohen’s Kappa 0.71-0.87), McNemar tests revealed significant differences between results obtained from Roche and DiaSorin. However, at low seroprevalences, the minor differences in specificity resulted in profound discrepancies of positive predictive values at 1% seroprevalence: 52.3% (36.2-67.9), 77.6% (52.8-91.5), and 32.6% (23.6-43.1) for Abbott, Roche, and DiaSorin, respectively. Conclusion We found diagnostically relevant differences in specificities for the anti-SARS-CoV-2 antibody assays by Abbott, Roche, and DiaSorin that have a significant impact on the positive predictive values of these tests.

Background: In the context of the COVID-19 pandemic, numerous new serological test systems for the detection of anti-SARS-CoV-2 antibodies have become available quickly. However, the clinical performance of many of them is still insufficiently described. Therefore we compared three commercial, CE-marked, SARS-CoV-2 antibody assays side by side. Methods: We included a total of 1,154 specimens from pre-COVID-19 times and 65 samples from COVID-19 patients (≥14 days after symptom onset) to evaluate the test performance of SARS-CoV-2 serological assays by Abbott, Roche, and DiaSorin. Results: All three assays presented with high specificities: 99.2% (98.6-99.7) for Abbott, 99.7% (99.2-100.0) for Roche, and 98.3% (97.3-98.9) for DiaSorin. In contrast to the manufacturers' specifications, sensitivities only ranged from 83.1% to 89.2%. Although the three methods were in good agreement (Cohen's Kappa 0.71-0.87), McNemar's test revealed significant differences between results obtained from Roche and DiaSorin. However, at low seroprevalences, the minor differences in specificity resulted in profound discrepancies of positive predictability at 1% seroprevalence: 52.3% (36.2-67.9), 77.6% (52.8-91.5), and 32.6% (23.6-43.1) for Abbott, Roche, and DiaSorin, respectively. Conclusion: We find diagnostically relevant differences in specificities for the anti-SARS-CoV-2 antibody assays by Abbott, Roche, and DiaSorin that have a significant impact on the positive predictability of these tests.

Background Reliable quantification of the antibody response to SARS-CoV-2 vaccination is highly relevant for identifying possible vaccine failure and estimating the time of protection. Therefore, we aimed to evaluate the performance of five different Anti-SARS-CoV-2 antibody assays regarding the quantification of anti-spike (S) antibodies induced after a single dose of BNT162b2. Methods Sera of n=69 SARS-CoV-2 naive individuals 21+/-1 days after vaccination with BNT162b2 (Pfizer/BioNTech) were tested using the following quantitative SARS-CoV-2 antibody assays: Roche S total antibody, DiaSorin trimeric spike IgG, DiaSorin S1/S2 IgG, Abbott II IgG, and Serion/Virion IgG. Test agreement was assessed by Passing-Bablok regression. Results were further compared to the percent inhibition calculated from a surrogate virus neutralization test (sVNT) by correlation and ROC (receiver-operating-characteristics) analysis. Results Individual values were distributed over several orders of magnitude for all assays evaluated. Although the assays were in good overall agreement (rho=0.80-0.94), Passing-Bablok regression revealed systematic and proportional differences, which could not be eliminated by converting the results to BAU/mL as suggested by the manufacturers. 7 (10%) individuals had a negative sVNT results (i.e. <30% inhibition). These samples were reliably identified by most assays and yielded low binding antibody levels (ROC-AUCs 0.84-0.93). Conclusions Although all assays evaluated showed good correlation, readings from different assays were not interchangeable, even when converted to BAU/mL using the WHO international standard for SARS-CoV-2 immunoglobulin. This highlights the need for further standardization of SARS-CoV-2 serology.

This work showed that the comparability of apparently standardized SARS-CoV-2 antibody assays (Roche, Abbott; both given in BAU/mL) after vaccination depends on the time of blood withdrawal. Initially (3 weeks after the first dose AZD1222), there were 3 times higher values in the Abbott assay, but this relationship inversed before boosting (11 weeks after the first dose) with Roche 2 times greater than Abbott.

Background SARS-CoV-2 antibody tests have undergone a remarkable improvement in performance. However, due to the low seroprevalence in several areas, very high demands are made on their specificity. Furthermore, the low antibody-response in some individuals requires high test sensitivity to avoid underestimating true seroprevalence. Optimization of testing has been reported through lowering manufacturer cut-offs to improve SARS-CoV-2 assay sensitivity or by combining two tests to improve specificity at the cost of sensitivity. However, these strategies have thus far been used in isolation of each other. Methods To increase sensitivity, cut-offs of three commercially available SARS-CoV-2 automated assays (Roche, Abbott, and DiaSorin) were reduced according to published values in a pre-pandemic specificity cohort (n=1117) and a SARS-CoV-2 positive cohort (n=64). All three testing systems were combined in an orthogonal approach with a confirmatory test, which was one of the remaining automated assays or one of two commercial ELISAs directed against the spike protein receptor binding-domain (RBD) or the nucleocapsid antigen (NP). Results The modified orthogonal test strategy resulted in an improved specificity of at least 99.8%, often even 100%, in all 12 tested combinations with no significant decline in sensitivity. In our cohort, regardless of whether the assays were used for screening or confirmation, combining Roche and Abbott delivered the best overall performance (+~10% sensitivity compared to the single tests and 100% specificity). Conclusion Here we propose a novel orthogonal assay strategy that approaches 100% specificity while maintaining or even significantly improving the screening test's sensitivity.

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Background There is preliminary evidence that individuals with previous SARS‐CoV‐2 infections exhibit a more pronounced antibody response. However, these assumptions have not yet been supported by data obtained through various CE‐marked tests. This study aimed to close this gap. Methods Sixty‐nine seronegatives and 12 individuals post‐SARS‐CoV‐2 infection (tested by CE‐labelled Roche NC immunoassay or PCR‐confirmed assay) were included 21 ± 1 days after receiving the first dose of the Pfizer/BioNTech BNT162b2 vaccine. Antibody response to viral spike protein (S) was assessed by CE‐labelled Roche S and DiaSorin S1/S2 assays and by a surrogate virus neutralization test (sVNT). Results After a single dose of BNT162b2, individuals after natural SARS‐CoV‐2 infection presented with markedly higher anti‐S levels than naïve individuals (Roche S: 9078.5 BAU/mL [5267.0‐24 298.5] vs 79.6 [24.7‐142.3]; and DiaSorin S1/S2: 1465.0 AU/mL [631.0‐5365.0] vs 63.7 [47.8‐87.5]) and showed all the maximum observed inhibition activity in the sVNT (98%), without overlaps between groups. There was a trend for higher responses in those with a more distant infection, although not statistically significant. The relative antibody increase after dose 2 was significantly higher among naïve individuals (25‐fold), but antibody levels remained below that of seropositives. Conclusions Compared with naïve individuals, seropositives after natural SARS‐CoV‐2 infection presented with a substantially higher antibody response already after dose 1 of BNT162b2, as measured by two CE‐marked in vitro diagnostic tests and a sVNT. These results should stimulate discussion and research on whether individuals after previous SARS‐CoV‐2 infection would benefit from a two‐part vaccination schedule or whether these currently much‐needed second doses could be saved.