2021
DOI: 10.1101/2021.02.16.21251653
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IMMUNO-COV™ v2.0: Development and Validation of a High-Throughput Clinical Assay for Measuring SARS-CoV-2-Neutralizing Antibody Titers

Abstract: Neutralizing antibodies are key determinants of protection from future infection, yet well-validated high-throughput assays for measuring titers of SARS-CoV-2-neutralizing antibodies are not generally available. Here we describe the development and validation of IMMUNO-COV™ v2.0 a scalable surrogate virus assay, which titrates antibodies that block infection of Vero-ACE2 cells by a luciferase-encoding vesicular stomatitis virus displaying SARS-CoV-2 spike glycoproteins (VSV-SARS2-Fluc). Antibody titers, calcul… Show more

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“…Vero-DSP1 and Vero-DSP2 co-monolayers, which produce functional Renilla luciferase following virus-mediated cell fusion, were seeded at 6×10 4 cells per well in 96-well black-walled plates the day before assay. Neutralizing antibody cocktails were prepared in OptiMEM as follows: α-SARS-CoV-2 spike cocktail (mAb10914 at 6 µg/mL combined with mAb10922 at 2 µg/mL, Genscript) [4], α-VSV-G cocktail (mAbVSV-G 8G5F11, Absolute Antibody #Ab01401-2.3) at 2 µg/mL combined with mAb VSV-G IE9F9 (Absolute Antibody #Ab01402-2.0) at 10 µg/mL [27], and α-SARS-CoV-2 spike and α-VSV-G cocktail (combination of all four antibodies). Three-fold serial dilutions of the antibody cocktails were prepared and incubated with an equal volume of VSV-SARS2 or VSV-SARS2(+G).…”
Section: Supplementary Materialsmentioning
confidence: 99%
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“…Vero-DSP1 and Vero-DSP2 co-monolayers, which produce functional Renilla luciferase following virus-mediated cell fusion, were seeded at 6×10 4 cells per well in 96-well black-walled plates the day before assay. Neutralizing antibody cocktails were prepared in OptiMEM as follows: α-SARS-CoV-2 spike cocktail (mAb10914 at 6 µg/mL combined with mAb10922 at 2 µg/mL, Genscript) [4], α-VSV-G cocktail (mAbVSV-G 8G5F11, Absolute Antibody #Ab01401-2.3) at 2 µg/mL combined with mAb VSV-G IE9F9 (Absolute Antibody #Ab01402-2.0) at 10 µg/mL [27], and α-SARS-CoV-2 spike and α-VSV-G cocktail (combination of all four antibodies). Three-fold serial dilutions of the antibody cocktails were prepared and incubated with an equal volume of VSV-SARS2 or VSV-SARS2(+G).…”
Section: Supplementary Materialsmentioning
confidence: 99%
“…However, all currently approved SARS-CoV-2 vaccines are administered via intramuscular injection which increases the cost and complexity of vaccination programs, contributes to vaccine hesitancy and fails to stimulate mucosal immunity [2,3]. Also, protective antibody titers are known to wane after successful vaccination (or after natural infection), such that periodic booster immunizations will be needed to sustain immunity, particularly in vulnerable individuals, so long as the virus remains endemic in the human population [4][5][6].…”
Section: Introductionmentioning
confidence: 99%
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