Related Experiment Video
Updated: Jul 29, 2025
Detection of SARS-CoV-2 Neutralizing Antibodies using High-Throughput Fluorescent Imaging of Pseudovirus Infection
Published on: June 5, 2021
Detection of SARS-CoV-2 Antibodies: Comparison of Enzyme Immunoassay, Surrogate Neutralization and Virus
Tatjana Vilibic-Cavlek1,2, Maja Bogdanic1, Ema Borko1
1Department of Virology, Croatian Institute of Public Health, 10000 Zagreb, Croatia.
Background:
Since sensitivity and specificity vary widely between tests, SARS-CoV-2 serology results should be interpreted with caution.
Methods:
The study included serum samples from patients who had recovered from COVID-19 (n = 71), individuals vaccinated against SARS-CoV-2 (n = 84), and asymptomatic individuals (n = 33). All samples were tested for the presence of binding antibodies (enzyme immunoassay; EIA), neutralizing (NT) antibodies (virus neutralization test; VNT), and surrogate NT (sNT) antibodies (surrogate virus neutralization test; sVNT) of SARS-CoV-2.
Results:
SARS-CoV-2-binding antibodies were detected in 71 (100%) COVID-19 patients, 77 (91.6%) vaccinated individuals, and 4 (12.1%) control subjects. Among EIA-positive samples, VNT was positive (titer ≥ 8) in 100% of COVID-19 patients and 63 (75.0%) of the vaccinated individuals, while sVNT was positive (>30% inhibition) in 62 (87.3%) patients and 59 (70.2%) vaccinated individuals. The analysis of antibody levels showed a significant moderate positive correlation between EIA and VNT, a moderate positive correlation between EIA and sVNT, and a strong positive correlation between VNT and sVNT. The proportion of positive sVNT detection rate was associated with VNT titer. The lowest positivity (72.4%/70.8%) was detected in samples with low NT titers (8/16) and increased progressively from 88.2% in samples with titer 32 to 100% in samples with titer 256.
Conclusions:
sVNT appeared to be a reliable method for the assessment COVID-19 serology in patients with high antibody levels, while false-negative results were frequently observed in patients with low NT titers.
