Quantitative measurement of IgG to SARS‐CoV‐2 antigens using monoclonal antibody‐based enzyme‐linked immunosorbent assays

Abstract

Objective Standardised quantitative analysis of the humoral immune response to SARS‐CoV‐2 antigens may be useful for estimating the extent and duration of immunity. The aim was to develop enzyme‐linked immunosorbent assays (ELISAs) for the quantification of human IgG antibodies against SARS‐CoV‐2 antigens. Methods Enzyme‐linked immunosorbent assays were developed based on monoclonal antibodies against human IgG and recombinant SARS‐CoV‐2 antigens (Spike‐S1 and Nucleocapsid). The WHO 67/086 immunoglobulin and WHO 20/136 SARS‐CoV‐2 references were used for standardisation. Sera of a study group of COVID‐19‐positive subjects (n = 144), pre‐pandemic controls (n = 135) and individuals vaccinated with BioNTech–Pfizer BNT162b2 vaccine (n = 48) were analysed. The study group sera were also tested using EuroImmun SARS‐CoV‐2‐ELISAs and a quantitative S1‐specific fluorescence enzyme immunoassay (FEIA) from Thermo Fisher. Results The ELISA results were repeatable and traceable to international units because of their parallelism to both WHO references. In the study group, median anti‐S1‐IgG concentrations were 102 BAU mL−1, compared to 100 and 1457 BAU mL−1 in the vaccination group after first and second vaccination, respectively. The ELISAs achieved an area under the curve (AUC) of 0.965 (S1) and 0.955 (Nucleocapsid) in receiver operating characteristic (ROC) analysis, and a specificity of 1 (S1) and 0.963 (Nucleocapsid) and sensitivity of 0.903 (S1) and 0.833 (Nucleocapsid) at the maximum Youden index. In comparison, the commercial assays (S1‐FEIA, S1 and Nucleocapsid ELISA EuroImmun) achieved sensitivities of 0.764, 0.875 and 0.882 in the study group, respectively. Conclusions The quantitative ELISAs to measure IgG binding to SARS‐CoV‐2 antigens have good analytical and clinical performance characteristics and units traceable to international standards.

Single‐Response Duplexing of Electrochemical Label‐Free Biosensor from the Same Tag

“…[94] Nonetheless, one should emphasize that the linear range of our devices is within the clinical range of COVID-19 IgG S (0 to 40 μg mL −1 ). [38,96] Further, using peptide as a recognition element typically adds advantages over big-sized biomolecules in terms of cost and stability, which play a crucial role in the commercial translation of diagnostic technologies. [6,7,16] In practice, the peptide could be chemically synthesized, and it is not amenable to denaturation.…”

Comparison of nucleocapsid and spike antibody ELISAs for determining SARS‐CoV‐2 seropositivity in Kenyan women and infants

“… 9 , 10 , 11 , 12 Heterogeneity in levels and specificity of spike and nucleocapsid antibodies from natural infection with SARS‐CoV‐2 could impact the sensitivity and specificity of serological assays in diverse populations. 7 , 13 Importantly, current vaccine‐induced immunity is limited to spike antibodies; thus, detection of antibodies against nucleocapsid antigen permits specific detection of natural infections in vaccinated populations.…”

Duplex Electrochemical Microfluidic Sensor for COVID‐19 Antibody Detection: Natural versus Vaccine‐Induced Humoral Response

“…The detection of S‐IgG showed a linear correlation in the concentration range of 20–1000 ng mL −1 , expressed by I = (0.028 ± 0.002) C + (1.3 ± 0.8), with an R 2 of 0.949 and the LOD was equal to 15 ng mL −1 . The ng mL −1 range of LOD from our electrochemical sensors has the sensitivity well within clinical range of the COVID‐19 antibody concentration (20–100 µg mL −1 [ 77,78 ] and 0–40 µg mL −1 [ 59,77 ] for N‐IgG and S‐IgG antibodies, respectively).…”

“…To assess the accuracy and the utility of our duplex electrochemical immunosensor for antibody detection in real samples, we employed human serum to conduct the recovery studies. [81] Human serum (Sigma-Aldrich, USA), was first diluted 500-fold in PBS (containing 1% BSA), in order to reach the linear range of the developed sensors, as anti-COVID antibodies have been reported in the range of 20-100 µg mL −1 [77,78] and 0-40 µg mL −1 [59,77] for N-IgG and S-IgG, respectively. Recovery studies were performed in diluted serum samples spiked with different concentrations of S-IgG and N-IgG.…”