Multi-center nationwide comparison of seven serology assays reveals a SARS-CoV-2 non-responding seronegative subpopulation

Abstract

Background: An Israeli national taskforce performed a multi-center clinical and analytical validation of seven serology assays to determine their utility and limitations for SARS-CoV-2 diagnosis.

Methods: Serology assays from Roche, Abbott, Diasorin, BioMerieux, Beckman-Coulter, Siemens, and an in-house RBD ELISA were included. Negative samples from 2391 individuals representative of the Israeli population, and 698 SARS-CoV-2 PCR positive patients, collected between March and May 2020, were analyzed.

Findings: Immunoassays sensitivities between 81.5%-89.4% and specificities between 97.7%-100% resulted in a profound impact on the expected Positive Predictive Value (PPV) in low (<15%) prevalence scenarios. No meaningful increase was detected in the false positive rate in children compared to adults. A positive correlation between disease severity and antibody titers, and no decrease in antibody titers in the first 8 weeks after PCR positivity was observed. We identified a subgroup of symptomatic SARS-CoV-2 positive patients (~5% of patients), who remained seronegative across a wide range of antigens, isotypes, and technologies.

Interpretation: The commercially available automated immunoassays exhibit significant differences in performance and expected PPV in low prevalence scenarios. The low false-positivity rate in under 20's suggests that cross-reactive immunity from previous CoV strains is unlikely to explain the milder disease course in children. Finding no decrease in antibody titers in the first 8 weeks is in contrast to some reports of short half-life for SARS-CoV-2 antibodies. The ~5% who were seronegative non-responders, using multiple assays in a population-wide manner, represents the proportion of patients that may be at risk for re-infection.

Funding: Israel Ministry of Health.

Keywords: Antibodies; COVID-19; IgG; SARS-CoV-2; Serology; Seronegative subpopulation; Validation study.

Fig. 1

Study selection process, total number of patients that were included and excluded, and details on the size of each tested group and kits based on the STARD guidelines. Out of 3098 available samples, 3089 samples were included in the study. Number of SARS-CoV-2 PCR positive (equal or above or below 14 days after first positive PCR) and negative samples tested with each of the serology assays are presented.

Fig. 2

Effect of SARS-CoV-2 prevalence on positive predictive values (PPV) of the different assays. The PPV of each assay was calculated and is depicted based on different SARS-CoV-2 prevalence in the population ranging between 0.5% and 15%..

Fig. 3

SARS-CoV-2 antibody levels during the first 8 weeks after a positive PCR result. Antibody titer of each sample was recorded for each serological assay. Restricted cubic spline regression line with five knots located at the 10th, 25th, 50th, 75th and 90th percentiles are presented for each immunoassay.