Dataset Information

Establishment and validation of a pseudovirus neutralization assay for SARS-CoV-2.

ABSTRACT: Pseudoviruses are useful virological tools because of their safety and versatility, especially for emerging and re-emerging viruses. Due to its high pathogenicity and infectivity and the lack of effective vaccines and therapeutics, live SARS-CoV-2 has to be handled under biosafety level 3 conditions, which has hindered the development of vaccines and therapeutics. Based on a VSV pseudovirus production system, a pseudovirus-based neutralization assay has been developed for evaluating neutralizing antibodies against SARS-CoV-2 in biosafety level 2 facilities. The key parameters for this assay were optimized, including cell types, cell numbers, virus inoculum. When tested against the SARS-CoV-2 pseudovirus, SARS-CoV-2 convalescent patient sera showed high neutralizing potency, which underscore its potential as therapeutics. The limit of detection for this assay was determined as 22.1 and 43.2 for human and mouse serum samples respectively using a panel of 120 negative samples. The cutoff values were set as 30 and 50 for human and mouse serum samples, respectively. This assay showed relatively low coefficient of variations with 15.9% and 16.2% for the intra- and inter-assay analyses respectively. Taken together, we established a robust pseudovirus-based neutralization assay for SARS-CoV-2 and are glad to share pseudoviruses and related protocols with the developers of vaccines or therapeutics to fight against this lethal virus.

SUBMITTER: Nie J

PROVIDER: S-EPMC7144318 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Establishment and validation of a pseudovirus neutralization assay for SARS-CoV-2.

Nie Jianhui J Li Qianqian Q Wu Jiajing J Zhao Chenyan C Hao Huan H Liu Huan H Zhang Li L Nie Lingling L Qin Haiyang H Wang Meng M Lu Qiong Q Li Xiaoyu X Sun Qiyu Q Liu Junkai J Fan Changfa C Huang Weijin W Xu Miao M Wang Youchun Y

Emerging microbes & infections 20201201 1

Pseudoviruses are useful virological tools because of their safety and versatility, especially for emerging and re-emerging viruses. Due to its high pathogenicity and infectivity and the lack of effective vaccines and therapeutics, live SARS-CoV-2 has to be handled under biosafety level 3 conditions, which has hindered the development of vaccines and therapeutics. Based on a VSV pseudovirus production system, a pseudovirus-based neutralization assay has been developed for evaluating neutralizing ...[more]

PMID: 32207377

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Project description:Neutralization assays are important for understanding and quantifying neutralizing antibody responses toward severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The SARS-CoV-2 lentivirus surrogate neutralization assay (SCLSNA) can be used in biosafety level 2 (BSL-2) laboratories and has been shown to be a reliable alternative approach to the plaque reduction neutralization test (PRNT). In this study, we optimized and validated the SCLSNA to assess its ability as a comparator and prescreening method to support the PRNT. Comparability between the PRNT and SCLSNA was determined through clinical sensitivity and specificity evaluations. Clinical sensitivity and specificity assays produced acceptable results, with 100% (95% confidence interval [CI], 94% to 100%) specificity and 100% (95% CI, 94% to 100%) sensitivity against ancestral Wuhan spike-pseudotyped lentivirus. The sensitivity and specificity against B.1.1.7 spike-pseudotyped lentivirus were 88.3% (95% CI, 77.8% to 94.2%) and 100% (95% CI, 94% to 100%), respectively. Assay precision measuring intra-assay variability produced acceptable results for high (50% PRNT [PRNT50], 1:≥640), mid (PRNT50, 1:160), and low (PRNT50, 1:40) antibody titer concentration ranges based on the PRNT50, with coefficients of variation (CVs) of 14.21%, 12.47%, and 13.28%, respectively. Intermediate precision indicated acceptable ranges for the high and mid concentrations, with CVs of 15.52% and 16.09%, respectively. However, the low concentration did not meet the acceptance criteria, with a CV of 26.42%. Acceptable ranges were found in the robustness evaluation for both intra-assay and interassay variability. In summary, the validation parameters tested met the acceptance criteria, making the SCLSNA method fit for its intended purpose, which can be used to support the PRNT. IMPORTANCE Neutralization studies play an important role in providing guidance and justification for vaccine administration and helping prevent the spread of diseases. The neutralization data generated in our laboratory have been included in the decision-making process of the National Advisory Committee on Immunization (NACI) in Canada. During the coronavirus 2019 (COVID-19) pandemic, the plaque reduction neutralization test (PRNT) has been the gold standard for determining neutralization of SARS-CoV-2. We validated a SARS-CoV-2 lentivirus surrogate neutralization assay (SCLSNA) as an alternative method to help support the PRNT. The advantages of using the SCLSNA is that it can process more samples, is less tedious to perform, and can be used in laboratories with a lower biosafety level. The use of the SCLSNA can further expand our capabilities to help fulfill the requirements for NACI and other important collaborations.

Dataset Information

Establishment and validation of a pseudovirus neutralization assay for SARS-CoV-2.

Publications

Establishment and validation of a pseudovirus neutralization assay for SARS-CoV-2.

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