Quantitative Measurement of IgG to Severe Acute Respiratory Syndrome Coronavirus-2 Proteins Using ImmunoCAP

Abstract

Background: Detailed understanding of the immune response to severe acute respiratory syndrome coronavirus (SARS-CoV)-2, the cause of coronavirus disease 2019 (CO­VID-19) has been hampered by a lack of quantitative antibody assays. Objective: The objective was to develop a quantitative assay for IgG to SARS-CoV-2 proteins that could be implemented in clinical and research laboratories. Methods: The biotin-streptavidin technique was used to conjugate SARS-CoV-2 spike receptor-binding domain (RBD) or nucleocapsid protein to the solid phase of the ImmunoCAP. Plasma and serum samples from patients hospitalized with COVID-19 (n = 60) and samples from donors banked before the emergence of COVID-19 (n = 109) were used in the assay. SARS-CoV-2 IgG levels were followed longitudinally in a subset of samples and were related to total IgG and IgG to reference antigens using an ImmunoCAP 250 platform. Results: At a cutoff of 2.5 μg/mL, the assay demonstrated sensitivity and specificity exceeding 95% for IgG to both SARS-CoV-2 proteins. Among 36 patients evaluated in a post-hospital follow-up clinic, median levels of IgG to spike-RBD and nucleocapsid were 34.7 μg/mL (IQR 18–52) and 24.5 μg/mL (IQR 9–59), respectively. Among 17 patients with longitudinal samples, there was a wide variation in the magnitude of IgG responses, but generally the response to spike-RBD and to nucleocapsid occurred in parallel, with peak levels approaching 100 μg/mL, or 1% of total IgG. Conclusions: We have described a quantitative assay to measure IgG to SARS-CoV-2 that could be used in clinical and research laboratories and implemented at scale. The assay can easily be adapted to measure IgG to mutated COVID-19 proteins, has good performance characteristics, and has a readout in standardized units.

Comparison of SARS-CoV-2 Antibody Response by Age Among Recipients of the BNT162b2 vs the mRNA-1273 Vaccine

“…Serum was isolated and IgG to SARS-CoV-2 spike RBD and nucleocapsid were measured with a quantitative ImmunoCAP-based system using a Phadia 250 (Thermo-Fisher/Phadia), as previously described. 3 Statistical analysis was performed with GraphPad Prism 8 (GraphPad Software). Antibody levels were expressed by geometric mean with 95% CIs and comparisons were made between groups with the Mann-Whitney U test.…”

“…Serum was isolated and IgG to SARS-CoV-2 spike RBD and nucleocapsid were measured with a quantitative ImmunoCAP-based system using a Phadia 250 (Thermo-Fisher/Phadia), as previously described. 3 Statistical analysis was performed with GraphPad Prism 8 (GraphPad Software).…”

“…The dotted lines denotes cutoff of assay, as previously described. 3 Accounting for multiple comparisons, P < .0125 was considered as significant. a P < .001.…”

Lipid bilayer coatings for rapid enzyme-linked immunosorbent assay

“…Using the three ELISA protocols and various blocking options, we proceeded to evaluate anti-N antibody detection in human serum. A normal human serum (NHS) specimen was spiked with 20 µg/mL anti-N antibody, which is within the range of clinically feasible conditions [23] for an individual infected with COVID-19 and was then diluted 1:100 fold and two-fold dilutions thereof for ELISA measurement ( Fig. 5 ).…”

Reply to: The antibody response to the glycan α‐Gal correlates with COVID‐19 symptoms

“…Both the IgG and IgE assays used α-Gal-HSA as the assay solid-phase, as previously described. 7 (B) IgG to tetanus toxoid was measured by ImmunoCAP using the commercial assay (Thermo-Fisher/Phadia)…”