We have validated a method for quantifying neutralization of the SARS-CoV-2 spike-ACE2 interaction in a single drop of blood, collected on filter paper following a simple finger stick. The protocol is based on a commercially available immunoassay platform, and can be scaled up for large-scale testing of samples collected in centralized or remote locations.
We document dose-dependent reduction in neutralization in dilutions of samples with high initial neutralization, as well as close correspondence in results from DBS in comparison with matched serum samples. The method also produces results that are precise and reliable, which is particularly important for studies with large numbers of samples assayed across multiple plates.
Furthermore, the method can be implemented in ubiquitous BSL2 laboratories, without the specialized facilities and biohazard risks associated with live virus methods. Finger stick DBS sampling is relatively low-cost and minimally-invasive, and samples can be self-collected in the home 14 , In addition, antibodies remain stable in DBS for several weeks at room temperature, and can be sent in the mail as nonregulated, exempt materials The possibility of remote serological testing, with large numbers of people, may be particularly advantageous when clinical resources are stretched thin and people are being encouraged to stay home.
A disadvantage of DBS includes the small volume of blood in comparison with venipuncture. Other potential issues, depending on the application, are variable quality of self-collected samples, and the possibility that samples may be delayed or lost during shipment by mail.
Recent studies have documented neutralization antibody responses to vaccination and natural infection with SARS-CoV-2 8 , 25 , 26 , and prior research with other viruses suggests that neutralizing antibodies are excellent indicators of protective immunity 9.
More widespread, community-based testing is important for determining the level of neutralizing antibodies that provide protection against COVID, and for estimating the level and duration of immunity in the general population—and subpopulations of particular interest—following multiple waves of community transmission Testing for neutralization antibodies can also play a role in evaluating the effectiveness of vaccines, and heterogeneity in vaccine responses across individuals 8.
Our quantitative DBS-based assay for neutralizing antibodies can help advance research and surveillance in these important areas.
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Widge, A. Crawford, K. Viruses 12 , Download references. Elizabeth M. McNally, Matthew P. You can also search for this author in PubMed Google Scholar. All authors reviewed and approved the final version of the manuscript. Correspondence to Thomas W. D'Aquila reports personal fees from Abbvie, outside the submitted work.
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In most pseudovirus neutralization assays, cell lines of human and animal origin have shown high sensitivity to pseudovirus binding and entry, 94 especially Vero and Huh7 cell lines. The conformational structure of the pseudoviral spike proteins is highly similar to that of the natural viral proteins, which can effectively mediate viral entry into host cells. Pseudoviruses are only suitable for studying virus entry and the role of antibodies against the S protein.
The Pearson correlation coefficient r values range from. Replicating chimeric viruses can be used to measure the ability of NAbs to reduce the growth of virus particles or eliminate viruses. If conditions permit, a live virus neutralization assay is finally required as a strong verification.
Moreover, the results of assays across laboratories are heterogeneous due to various culture conditions, virus strains and cell lines, so it is difficult to standardize these results without global protocols and standards for assays. Although there is a certain relationship with NAbs, it does not specifically refer to NAbs. At present, only a few of laboratories have verified the relevance with the live virus neutralization assay.
The correlation R 2 between sVNT and pseudovirus neutralization assays ranged from. However, Meyer et al. The sensitivity drops to This may be due to the different sample titres between the two assays, and the sensitivity of sVNT relies on antibody titres in the sample, which is positively correlated. In the experiment of Sholukh et al, the correlation coefficient r was. In summary, the sVNTs do not require the use of live virus or pseudovirus, and the detection can be completed in a short time under ordinary experimental conditions.
However, they still have some limitations. Firstly, the virus gene needs to be modified. NAbs titre is a key parameter for predicting immunity. Nowadays, a variety of mature methods for the detection of NAbs have been applied in practice and have provided assistance to the development of vaccines and antivirals.
However, most of these methods have their limitations Table 3 and need to be continuously improved. For instance, authentic virus assay requires exposure to viruses, with safety issues. It still remains unclear whether and how the dynamic changes in the NAbs titres are correlated with clinical outcomes. The authors have no conflicts of interest to declare that are relevant to the content of this article.
Zeliang Chen had the idea for the article. Yuying Lu wrote the first draft of the manuscript. Jin Wang critically revised the workhand. Qianlin Li and Huan Hu searched for references and drew pictures. Jiahai Lu provided some revision opinions for the first draft. All authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. Scand J Immunol. Yuying Lu and Jin Wang contributed equally to the work. National Center for Biotechnology Information , U.
Author information Article notes Copyright and License information Disclaimer. Zeliang Chen, Email: nc. Corresponding author. Email: nc. It can be used for unrestricted research re-use and analysis in any form or by any means with acknowledgement of the original source, for the duration of the public health emergency. Associated Data Data Availability Statement Data sharing is not applicable to this article as no new data were created or analyzed in this study.
Open in a separate window. Evaluation of convalescent plasma CP therapy, mAbs and vaccines Before effective drugs and vaccines are developed, convalescent plasma could represent an immediate stopgap treatment against viruses. Live virus neutralization assay Virus replication requires host cells to supply raw materials, energy and replication sites. Pseudovirus neutralization assay Pseudovirus or pseudotyped virus is a recombinant virus particle whose core skeleton and surface protein derive from a variety of viruses.
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