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Optimization and evaluation of a live virus SARS-CoV-2 neutralization assay
Anders Frische1, Patrick Terrence Brooks2, Mikkel Gybel-Brask2
1Department of Virus & Microbiological Special Diagnostics, Statens Serum Institut, Copenhagen, Denmark.
Plos One
|July 28, 2022
Summary
This study details a new live virus microneutralization assay for SARS-CoV-2. The optimized assay accurately quantifies functional antibody responses against the virus, crucial for vaccine and therapeutic development.
Area of Science:
- Virology
- Immunology
- Assay Development
Background:
- Virus neutralization assays are essential for measuring functional antibody responses against viral infections.
- These assays are critical for evaluating vaccine efficacy, therapeutic antibody potency, viral immune evasion, and natural immunity.
- Standardized methods are needed for reliable SARS-CoV-2 neutralization testing.
Purpose of the Study:
- To develop, optimize, and evaluate a live virus microneutralization assay for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).
- To establish a standardized assay for comparing neutralization investigations and assessing antibody potency against SARS-CoV-2.
Main Methods:
- Development of a live virus microneutralization assay using SARS-CoV-2 clinical isolates and Vero E6 cells.
- Quantification of replicating virus via ELISA targeting the SARS-CoV-2 nucleocapsid protein.
- Evaluation of critical parameters including virus titration, cell number, viral dose, incubation time, and normalization methods.
Main Results:
- Optimal virus titration was determined at 96 hours to accommodate varying clinical isolate growth kinetics.
- Nucleocapsid protein levels showed a strong correlation with virus inoculum at 24 hours post-inoculation.
- Two normalization models effectively reduced the coefficient of variance to below 16.5%, enhancing assay precision.
Conclusions:
- The developed SARS-CoV-2 microneutralization assay, based on WHO influenza protocols, is robust and reproducible.
- This assay provides a standardized method for quantifying functional antibody responses against SARS-CoV-2.
- The assay is suitable for evaluating vaccine and therapeutic antibody potency and understanding post-infection immunity.
