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Author Notes:

Filipp Frank, Department of Biochemistry, Emory University School of Medicine, Emory University, Atlanta, GA 30322, USA. Email: ffrank2@emory.edu

William H. Hudson, Emory Vaccine Center, Emory University School of Medicine, Atlanta, GA 30322, USA. Email: william.hudson@bcm.edu

Eric A. Ortlund, Department of Biochemistry, Emory University School of Medicine, Emory University, Atlanta, GA 30322, USA. Email: eortlun@emory.edu

F.F., W.H.H., and E.A.O. designed mammalian surface-display and mutational scanning experiments and analyzed data. M.M.K. and F.F. performed flow cytometry titration and DMS experiments. J.A.S., M.G., L.B., A.R., H.B.B., and W.A.L. designed testing experiments. A.R., L.B., and H.B.B. performed preparation of pools, dilution, quality control of test panels containing VOC/I, testing of tests, unblinding test results, analyzing data, and reporting results. X.L. performed protein expression, purification, and BLI experiments and analysis. A.B.P. performed negative stain electron microscopy experiments and analysis. M.L.C. performed mutagenesis and sequence conservation analyses. A.P. analyzed patient sequencing data.

Research reported in this publication was supported by the National Institute of Biomedical Imaging and Bioengineering of the National Institutes of Health (under award numbers 75N92019P00328, U54EB015408, and U54EB027690) as part of the Rapid Acceleration of Diagnostics (RADx) initiative, launched to speed innovation in the development, commercialization, and implementation of technologies for COVID-19 testing. The funders had no role in the decision to submit the work for publication and the views expressed herein are the authors’ and do not necessarily represent the views of the National Institutes of Health or the United States Department of Health and Human Services. E.A.O. was supported by NIDDK award 5R01DK115213. W.H.H. was supported by the NIH under award number K99AI153736. X.L. was supported by an American Heart Association career development award number 848388. The following reagent was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, NIAID, and NIH: SARS-related coronavirus 2, isolate USA (WA1/2020), and heat-inactivated (NR-52286). We thank the laboratories: Helix OpCo LLC (Jimmy Ramirez), LabCorp (Susan de Los Rios), and the University of Washington (Alex Greninger and Pavitra Roychoudhury) for providing the remnant clinical samples. We thank Kimberly Pachura for her contributions to quality control of samples used in testing the tests. We would also like to thank Mimi Le and the Children’s Clinical and Translational Discovery Core for help with organizing and selecting the variant samples needed for generating pools. We thank Hans Verkerke and Andrew Neish for help with BLI measurements. Next generation sequencing services were provided by the Yerkes NHP Genomics Core, which is supported in part by NIH P51 OD011132. Sequencing data were acquired on an Illumina NovaSeq6000 funded by NIH S10 OD026799. FACS experiments were carried out in the Flow Cytometry Core Facility of the Emory University School of Medicine.

The authors declare no competing interests.

Subjects:

Keywords:

  • Science & Technology
  • Life Sciences & Biomedicine
  • Biochemistry & Molecular Biology
  • Cell Biology
  • RECEPTOR-BINDING DOMAIN
  • PROTEIN
  • CORONAVIRUS

Deep mutational scanning identifies SARS-CoV-2 Nucleocapsid escape mutations of currently available rapid antigen tests

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Journal Title:

CELL

Volume:

Volume 185, Number 19

Publisher:

, Pages 3603-+

Type of Work:

Article | Final Publisher PDF

Abstract:

The effects of mutations in continuously emerging variants of SARS-CoV-2 are a major concern for the performance of rapid antigen tests. To evaluate the impact of mutations on 17 antibodies used in 11 commercially available antigen tests with emergency use authorization, we measured antibody binding for all possible Nucleocapsid point mutations using a mammalian surface-display platform and deep mutational scanning. The results provide a complete map of the antibodies’ epitopes and their susceptibility to mutational escape. Our data predict no vulnerabilities for detection of mutations found in variants of concern. We confirm this using the commercial tests and sequence-confirmed COVID-19 patient samples. The antibody escape mutational profiles generated here serve as a valuable resource for predicting the performance of rapid antigen tests against past, current, as well as any possible future variants of SARS-CoV-2, establishing the direct clinical and public health utility of our system.
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