Vaccine-induced SARS-CoV-2 antibody response: the comparability of S1-specific binding assays depends on epitope and isotype discrimination

Abstract

Background: Quantification of the SARS-CoV-2-specific immune response by serological immunoassays is critical for the management of the COVID-19 pandemic. In particular, neutralizing antibody titers to the viral spike (S) protein have been proposed as a correlate of protection (CoP). The WHO established the First International Standard (WHO IS) for anti-SARS-CoV-2 immunoglobulin (Ig) (NIBSC 20/136) to harmonize binding assays with the same antigen specificity by assigning the same unitage in binding antibody units (BAU)/ml.

Method: In this study, we analyzed the S1-specific antibody response in a cohort of healthcare workers in Germany (n = 76) during a three-dose vaccination course over 8.5 months. Subjects received either heterologous or homologous prime-boost vaccination with ChAdOx1 nCoV-19 (AstraZeneca) and BNT162b2 (Pfizer-BioNTech) or three doses of BNT162b2. Antibodies were quantified using three anti-S1 binding assays (ELISA, ECLIA, and PETIA) harmonized to the WHO IS. Serum levels of neutralizing antibodies were determined using a surrogate virus neutralization test (sVNT). Binding assays were compared using Spearman's rank correlation and Passing-Bablok regression.

Findings: All assays showed good correlation and similar antibody kinetics correlating with neutralizing potential. However, the assays show large proportional differences in BAU/ml. ECLIA and PETIA, which detect total antibodies against the receptor- binding domain (RBD) within the S1 subunit, interact similarly with the convalescent plasma-derived WHO IS but differently with vaccine serum, indicating a high sensitivity to the IgG/IgM/IgA ratio.

Conclusion: All three binding assays allow monitoring of the antibody response in COVID-19-vaccinated individuals. However, the assay-specific differences hinder the definition of a common protective threshold in BAU/ml. Our results highlight the need for the thoughtful use of conversion factors and consideration of method-specific differences. To improve the management of future pandemics and harmonize total antibody assays, we should strive for reference material with a well-characterized Ig isotype composition.

Keywords: COVID-19 vaccines; SARS-CoV-2 antibody; WHO standard; correlate of protection; humoral immune response; neutralizing antibodies; serological testing; spike protein.

Figure 1

SARS-CoV-2 S1-specific antibody response in a heterogeneous vaccination cohort (n = 76) over 8.5 months. Serum samples were measured by three routine immunoassays: ECLIA (Roche; black line), ELISA (Euroimmun; blue line), and PETIA (Diasys; red dotted line). Mean binding antibody units per milliliter (BAU/ml) for each time point (TP1–TP11) are plotted in logarithmic scale.

Figure 2

Comparison of immunoassays. Passing–Bablok regression analysis (TP6; n = 63). (A) Scatter diagram. Regression line (blue line), 95% CI of the regression line (dotted red lines), and identity line (thin red line). (B) Residual plot. Distribution of differences from the regression line (blue line). The red square indicates an outlier.

Figure 3

Neutralizing potential over time. S1-specific antibody titers (BAU/ml) for five selected time points are plotted against percentage inhibition (IH%) measured by a surrogate virus neutralization test (sVNT; SARS-CoV-2-NeutraLISA; Euroimmun). Negative cut-off, <20 IH%; positive cut-off, ≥35 IH% (green line).

Figure 4

(A) Proportion of samples (%) in six different sVNT categories (<20; 20–45; ≥35, 35–59; 60–89; and ≥90 IH%) and above three potential BAU/ml thresholds (≥100; ≥200; and ≥400 BAU/ml) for five selected time points. (B) Proportion of samples (%) in four different categories (<35 IH%/<100 BAU/ml; <35 IH%/≥100 BAU/ml; ≥35 IH%/<100 BAU/ml; ≥35 IH%/≥100 BAU/ml) for three selected time points (TP3, TP5, and TP9).