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Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2
Pseudoviruses are useful surrogates for highly pathogenic viruses because of their safety, genetic stability, and scalability for screening assays. Many different pseudovirus platforms exist, each with different advantages and limitations. Here we report our efforts to optimize and characterize an H...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7946320/ https://www.ncbi.nlm.nih.gov/pubmed/33690649 http://dx.doi.org/10.1371/journal.pone.0248348 |
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author | Neerukonda, Sabari Nath Vassell, Russell Herrup, Rachel Liu, Shufeng Wang, Tony Takeda, Kazuyo Yang, Ye Lin, Tsai-Lien Wang, Wei Weiss, Carol D. |
author_facet | Neerukonda, Sabari Nath Vassell, Russell Herrup, Rachel Liu, Shufeng Wang, Tony Takeda, Kazuyo Yang, Ye Lin, Tsai-Lien Wang, Wei Weiss, Carol D. |
author_sort | Neerukonda, Sabari Nath |
collection | PubMed |
description | Pseudoviruses are useful surrogates for highly pathogenic viruses because of their safety, genetic stability, and scalability for screening assays. Many different pseudovirus platforms exist, each with different advantages and limitations. Here we report our efforts to optimize and characterize an HIV-based lentiviral pseudovirus assay for screening neutralizing antibodies for SARS-CoV-2 using a stable 293T cell line expressing human angiotensin converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2). We assessed different target cells, established conditions that generate readouts over at least a two-log range, and confirmed consistent neutralization titers over a range of pseudovirus input. Using reference sera and plasma panels, we evaluated assay precision and showed that our neutralization titers correlate well with results reported in other assays. Overall, our lentiviral assay is relatively simple, scalable, and suitable for a variety of SARS-CoV-2 entry and neutralization screening assays. |
format | Online Article Text |
id | pubmed-7946320 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-79463202021-03-19 Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2 Neerukonda, Sabari Nath Vassell, Russell Herrup, Rachel Liu, Shufeng Wang, Tony Takeda, Kazuyo Yang, Ye Lin, Tsai-Lien Wang, Wei Weiss, Carol D. PLoS One Research Article Pseudoviruses are useful surrogates for highly pathogenic viruses because of their safety, genetic stability, and scalability for screening assays. Many different pseudovirus platforms exist, each with different advantages and limitations. Here we report our efforts to optimize and characterize an HIV-based lentiviral pseudovirus assay for screening neutralizing antibodies for SARS-CoV-2 using a stable 293T cell line expressing human angiotensin converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2). We assessed different target cells, established conditions that generate readouts over at least a two-log range, and confirmed consistent neutralization titers over a range of pseudovirus input. Using reference sera and plasma panels, we evaluated assay precision and showed that our neutralization titers correlate well with results reported in other assays. Overall, our lentiviral assay is relatively simple, scalable, and suitable for a variety of SARS-CoV-2 entry and neutralization screening assays. Public Library of Science 2021-03-10 /pmc/articles/PMC7946320/ /pubmed/33690649 http://dx.doi.org/10.1371/journal.pone.0248348 Text en https://creativecommons.org/publicdomain/zero/1.0/ This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 (https://creativecommons.org/publicdomain/zero/1.0/) public domain dedication. |
spellingShingle | Research Article Neerukonda, Sabari Nath Vassell, Russell Herrup, Rachel Liu, Shufeng Wang, Tony Takeda, Kazuyo Yang, Ye Lin, Tsai-Lien Wang, Wei Weiss, Carol D. Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2 |
title | Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2 |
title_full | Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2 |
title_fullStr | Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2 |
title_full_unstemmed | Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2 |
title_short | Establishment of a well-characterized SARS-CoV-2 lentiviral pseudovirus neutralization assay using 293T cells with stable expression of ACE2 and TMPRSS2 |
title_sort | establishment of a well-characterized sars-cov-2 lentiviral pseudovirus neutralization assay using 293t cells with stable expression of ace2 and tmprss2 |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7946320/ https://www.ncbi.nlm.nih.gov/pubmed/33690649 http://dx.doi.org/10.1371/journal.pone.0248348 |
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