Developing Antibodies and Antigens for COVID-19 Diagnostics
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A month after the SARS-CoV-2 genome was published, The Native Antigen Company became one of the first recognized suppliers to announce the commercial release of the virus’ S1 and S2 antigens. In addition, the company is continually working to develop a comprehensive range of other coronavirus antigens and antibodies.
Technology Networks recently spoke to Dr Andy Lane, Commercial Director, The Native Antigen Company, to learn how these proteins are produced, the role Spike proteins play in mediating entry of SARS-CoV-2 into the cell, and how they can be used to drive the development of COVID-19 diagnostics.
Anna MacDonald (AM): In what ways can viral infections such as COVID-19 be diagnosed?
Andy Lane (AL): The two major methods for diagnosing viral infection are the polymerase chain reaction (PCR) and immunoassays (antibody and antigen kits).
PCR assays are rapid and easy to design, which has made them fundamental to COVID-19 diagnosis. However, as PCR relies on the amplification of viral RNA samples from the respiratory tract, it is limited to use during the acute phase of infection when the virus is still present. Due to the need for amplification to detect RNA, PCR tests are also difficult to miniaturize for use at the point-of-care (PoC).
Like RNA, antigens are also present in the respiratory tract of infected individuals and can be used to diagnose acute-phase infection. However, the added advantage of antigen testing over PCR, is that antigens can be detected with inexpensive, simple, PoC formats, such as the lateral-flow assay (LFA), which does not rely on sample amplification.
Basic design of LFAs, akin to home pregnancy tests. LFAs typically comprise a plastic cassette, containing a strip of paper that is able to absorb and transport analyte deposited in blood or sputum. Antigens migrate and bind to conjugate and test antibodies to confirm infection, indicated by a solid, visible line. Secondary control lines ensure that the test is working correctly.
Thanks to their simple design, LFAs can be mass-produced and made widely available for the community, or door-to-door testing, providing rapid results to determine whether individuals should self-isolate and/or seek medical care. This could be a game-changer for COVID-19, as much larger proportions of the population could be tested, without requiring individuals to travel to already overwhelmed medical facilities.
Contrary to antigens and RNA, antibodies that are raised against SARS-CoV-2 are slow to present (typically 9–10 days for IgM (Lei et al., 2020; Li et al., 2020)), but may persist in the bloodstream for many years. Therefore, while antibodies may not be good markers of acute infection, they are ideal for detecting historic infections.
Antibody testing could be invaluable in determining the true transmission and case fatality rates of COVID-19, enabling public health bodies to better model its spread and direct healthcare policy. Antibody tests could also be used to diagnose healthcare workers who have been exposed to the virus to ensure that they have developed natural immunity before returning to work, as well as measuring patient immune responses to support the rapid development of an urgently-needed COVID-19 vaccine.
AM: Where do reagents fit into this process?
AL: The foundation of any effective immunoassay are the critical reagents used to design it, namely antigens and antibodies. These proteins drive the fundamental reactions of serological tests and ultimately determine their sensitivity and specificity.
Due to the biosafety implications of handling live viruses, many labs must work with antigens that are expressed from recombinant systems. However, not all protein-expression systems are the same and this can result in significant discrepancies between the recombinant antigen and its native counterpart. For example, simpler systems like E. coli lack the necessary glycosylation machinery to produce proteins with all the post-translational modifications of human cells, which can alter epitopes and protein conformation. Consequently, this can lead to poor sensitivity and specificity when designing diagnostic assays.
The Native Antigen Company, uses a proprietary, mammalian, VirtuE expression system to produce recombinant proteins with complete glycosylation and proper folding—both of which are essential for full biological and antigenic activity. By using antigens that very closely resemble those found in nature, assay developers are then able to design kits with high sensitivity and specificity.
AM: What is the WHO R&D Blueprint? How does the work of The Native Antigen Company support this?
AL: In 2015, at the request of 194 member states, the World Health Organization brought together a broad coalition of experts to develop the Research and Development Blueprint for priority emerging infectious diseases. The Blueprint comprises an evolving list of diseases that pose a severe public health risk with epidemic potential, and for which insufficient, or no preventive and curative solutions currently exist. The Blueprint is paramount in guiding our efforts to support ongoing research into the prevention and diagnosis of emerging diseases.
One of the diseases included on the list is “Disease X”, a yet-unknown pathogen with pandemic potential, or in other words, the next “big thing”. It has become quite apparent over recent months that COVID-19 is a new “Disease X”, and has since been added to the Blueprint’s list.
Following the guidance outlined in the Blueprint, The Native Antigen Company has developed a comprehensive range of coronavirus antigens and antibodies to support in vitro diagnostics and pharmaceutical researchers to help drive the development of serological assays and vaccines that will be vital in stemming the spread of this disease.
AM: The Native Antigen Company released SARS-CoV-2 antigens after just one month of its genome being published. How are these antigens being used to help stem the spread of COVID-19?
AL: In February, The Native Antigen Company was one of the world’s first recognized suppliers to release commercially available S1 and S2 antigens for SARS-CoV-2. These antigens were produced to support the urgently needed development of SARS-CoV-2 countermeasures and are now being used by world-leading organizations to develop diagnostics and vaccines for COVID-19. Our scientists are still hard at work in lab, developing new antigens and antibodies to support coronavirus research. To keep up-to-date with our latest product developments, please see our coronavirus pipeline.
AM: What is the role of the coronavirus Spike protein? Why were antigens for this protein in particular so important to produce?
AL: Spike proteins are the visible protrusions on the surface of SARS-CoV-2, which give the virus its characteristic, crown-like appearance. These homotrimeric proteins are heavily glycosylated, with each comprising two distinct subunits: S1 and S2.
Transmission electron micrograph of SARS-CoV-2. This image is the work of the Australian Animal Health Laboratory (AAHL), CSIRO.
The role of Spike is to act as a molecular key, which it achieves by recognizing and binding to specific ACE2 cell-surface receptors (the locks) present on the surface of our cells, through the S1 receptor-binding domain. When S1 binds to ACE2, Spike undergoes dramatic structural changes to alter the conformation of ACE2 and mediate entry of the virus into the cell.
However, as Spike proteins must project into the external environment to effectively bind cell-surface receptors, they are exposed to recognition by the immune system. This makes Spike the immunodominant coronavirus antigen, causing it to elicit a strong neutralizing antibody response (Ju et al., 2020) that has made it the focal target of diagnostic and vaccine development.
Dr Andy Lane was speaking to Anna MacDonald, Science Writer, Technology Networks.