Yue Zheng1, Erin T Larragoite1, Elizabeth S C P Williams1, Juan Lama2, Isabel Cisneros2, Julio C Delgado1,3, Patricia Slev1,3, Jenna Rychert3, Emily A Innis1, Mayte Coiras4, Matthew T Rondina5, Adam M Spivak6, Vicente Planelles7. 1. Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT, USA. 2. RetroVirox, Inc., San Diego, CA, USA. 3. Associated Regional and University Pathologists (ARUP) Laboratories, Salt Lake City, UT, USA. 4. AIDS Immunopathology Unit, National Center of Microbiology (CNM), Instituto de Salud Carlos III, Madrid, Spain. 5. Department of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT, USA. 6. Department of Medicine, University of Utah School of Medicine, Salt Lake City, UT, USA. 7. Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT, USA. vicente.planelles@path.utah.edu.
Abstract
BACKGROUND: Virus neutralization by antibodies is an important prognostic factor in many viral diseases. To easily and rapidly measure titers of neutralizing antibodies in serum or plasma, we developed pseudovirion particles composed of the spike glycoprotein of SARS-CoV-2 incorporated onto murine leukemia virus capsids and a modified minimal murine leukemia virus genome encoding firefly luciferase. This assay design is intended for use in laboratories with biocontainment level 2 and therefore circumvents the need for the biocontainment level 3 that would be required for replication-competent SARS-CoV-2 virus. To validate the pseudovirion assay, we set up comparisons with other available antibody tests including those from Abbott, Euroimmun and Siemens, using archived, known samples. RESULTS: 11 out of 12 SARS-CoV-2-infected patient serum samples showed neutralizing activity against SARS-CoV-2-spike pseudotyped MLV viruses, with neutralizing titers-50 (NT50) that ranged from 1:25 to 1:1,417. Five historical samples from patients hospitalized for severe influenza infection in 2016 tested negative in the neutralization assay (NT50 < 25). Three serum samples with high neutralizing activity against SARS-CoV-2/MLV pseudoviruses showed no detectable neutralizing activity (NT50 < 25) against SARS-CoV-1/MLV pseudovirions. We also compared the semiquantitative Siemens SARS-CoV-2 IgG test, which measures binding of IgG to recombinantly expressed receptor binding domain of SARS-CoV-2 spike glycoprotein with the neutralization titers obtained in the pseudovirion assay and the results show high concordance between the two tests (R2 = 0.9344). CONCLUSIONS: SARS-CoV-2 spike/MLV pseudovirions provide a practical means of assessing neutralizing activity of antibodies in serum or plasma from infected patients under laboratory conditions consistent with biocontainment level 2. This assay offers promise also in evaluating immunogenicity of spike glycoprotein-based candidate vaccines in the near future.
BACKGROUND: Virus neutralization by antibodies is an important prognostic factor in many viral diseases. To easily and rapidly measure titers of neutralizing antibodies in serum or plasma, we developed pseudovirion particles composed of the spike glycoprotein of SARS-CoV-2 incorporated onto murine leukemia virus capsids and a modified minimal murine leukemia virus genome encoding firefly luciferase. This assay design is intended for use in laboratories with biocontainment level 2 and therefore circumvents the need for the biocontainment level 3 that would be required for replication-competent SARS-CoV-2 virus. To validate the pseudovirion assay, we set up comparisons with other available antibody tests including those from Abbott, Euroimmun and Siemens, using archived, known samples. RESULTS: 11 out of 12 SARS-CoV-2-infectedpatient serum samples showed neutralizing activity against SARS-CoV-2-spike pseudotyped MLV viruses, with neutralizing titers-50 (NT50) that ranged from 1:25 to 1:1,417. Five historical samples from patients hospitalized for severe influenza infection in 2016 tested negative in the neutralization assay (NT50 < 25). Three serum samples with high neutralizing activity against SARS-CoV-2/MLV pseudoviruses showed no detectable neutralizing activity (NT50 < 25) against SARS-CoV-1/MLV pseudovirions. We also compared the semiquantitative Siemens SARS-CoV-2 IgG test, which measures binding of IgG to recombinantly expressed receptor binding domain of SARS-CoV-2spike glycoprotein with the neutralization titers obtained in the pseudovirion assay and the results show high concordance between the two tests (R2 = 0.9344). CONCLUSIONS:SARS-CoV-2spike/MLV pseudovirions provide a practical means of assessing neutralizing activity of antibodies in serum or plasma from infectedpatients under laboratory conditions consistent with biocontainment level 2. This assay offers promise also in evaluating immunogenicity of spike glycoprotein-based candidate vaccines in the near future.
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