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Defeating COVID-19: The Science Behind a New ELISA for COVID-19 Seroconversion Detection

Contributed by Aliyah Weinstein, Ph.D.

The recent and rapid spread of the novel coronavirus, SARS-CoV-2, has necessitated the development of new assays that are capable of detecting the presence of this virus in patient samples or evidence of recent infection. One strategy being developed are assays to detect the presence of anti-SARS-CoV-2 antibodies in patient sera.  

Defeating COVID-19: The Science Behind a New ELISA for COVID-19 Seroconversion Detection

Defeating COVID-19: The Science Behind a New ELISA for COVID-19 Seroconversion Detection

Bethyl’s SARS-CoV-2 ELISA detects antibodies against the receptor binding domain (RBD) of the Spike protein of SARS-CoV-2 (S1-RBD) in serum of patients who have recently been infected with the virus. The S1-RBD is responsible for the entry of the virus into human cells via the ACE2 receptor. While SARS-CoV-2 is in some ways similar to the SARS-CoV virus that caused the SARS outbreak in 2003, the spike protein of SARS-CoV-2 is evolutionarily distinct1. Instead, the spike protein of SARS-CoV-2 shares a 97.7% identity with the spike protein of the bat coronavirus from which it originated. The receptor binding domain S1-RBD differs from the receptor binding domain of the spike protein of SARS-CoV and several bat coronaviruses, with many amino acid substitutions and insertions. The uniqueness of S1-RBD makes it a useful target for this ELISA as antibodies against other viruses and more common coronaviruses are unlikely to crossreact with this antigen.  Given the relative sequence homology between SARS-CoV-2 and SARS-CoV immune crossreactivity could be expected. 

It is hypothesized that the Spike (S) protein of SARS-CoV-2 is one of the most immunogenic of its proteins, making it a good candidate for an ELISA to detect an immune response to SARS-CoV-2. This hypothesis is based on studies of patients infected with the related virus, SARS-CoV, showing that antibodies against portions of the S protein could be detected in all patients in the study2. These antibodies were present for at least 30 weeks following the onset of disease. In the case of SARS-CoV-2, antibodies against S1-RBD are neutralizing antibodies against viral entry into human cells, as these antibodies compete for ACE2 binding to the virus3.

The S protein of SARS-CoV-2 is also a better choice of antigen than the nucleocapsid (N) protein for ELISAs. In a head to head comparison of ELISAs using either the S or N protein as capture, the ELISA using the S protein was more sensitive than the ELISA that used the N protein, when detecting IgM antibodies4. No significant difference was observed between using either protein for the detection of IgG.

Using an ELISA to detect SARS-CoV-2 antibodies is a better choice than a lateral flow immunoassay, which is commonly used for clinical diagnostics due to the speed at which they can be performed. ELISA is the more sensitive of the two assays. A recent study to evaluate commercial lateral flow immunoassays vs. ELISAs against the Spike protein of SARS-CoV-2 indicated that lateral flow assays cannot be used as diagnostic tools due to the low sensitivity of all nine lateral flow assays tested for IgM, IgG, and total antibodies. Conversely, ELISA could reliably detect anti-S IgM and IgG antibodies from patient samples, for at least 60 days following onset of COVID-19 symptoms5.

Serologic assays, namely ELISAs against S1-RBD, have a role in the future of tracking the spread of SARS-CoV-2 and gaining a better understanding of immunity in the fight against COVID-19.


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1. Ortega JT, Serrano ML, Pujol FH, Rangel HR (2020) Role of changes in SARS-CoV-2 spike protein in the interaction with the human ACE2 receptor: An in silico analysis. EXCLI Journal; 19:Doc410; ISSN 1611-2156.

2. Qiu M, Shi Y, Guo Z, Chen Z, He R, Chen R, Zhou D, Dai E, Wang X, Si B, Song Y, Li J, Yang L, Wang J, Wang H, Pang X, Zhai J, Du Z, Liu Y, Zhang Y, Li L, Wang J, Sun B, Yang R (2005) Antibody responses to individual proteins of SARS coronavirus and their neutralization activities. Microbes and Infection 7:882–889 .

3. Ju, B, Zhang, Q, Ge, X, Wang, R, Yu, J, Shan, S, Zhou, B, Song, S, Tang, X, Yu, J, Ge, J, Lan, J, Yuan, J, Wang, H, Zhao, J, Zhang, S, Wang, Y, Shi, X, Liu, L, … Zhang, L (2020). Potent human neutralizing antibodies elicited by SARS-CoV-2 infection. Cold Spring Harbor Laboratory.

4. Liu W, Liu L, Kou G, Zheng Y, Ding Y, Ni W, Wang Q, Tan L, Wu W, Tang S, Xiong Z, Zheng S (2020) Evaluation of Nucleocapsid and Spike Protein-based ELISAs for detecting antibodies against SARS-CoV-2. Journal of Clinical Microbiology.

5. Adams ER, Ainsworth M, Anand R, Andersson MI, Auckland K, Baillie JK, … Whitehouse J (2020) Antibody testing for COVID-19: A report from the National COVID Scientific Advisory Panel. medRxiv.