The benefits of globalization have ushered in the reality of fast-spreading infectious diseases. This has made the quick development of preventive vaccines and immune therapies imperative. Spread of pathogens through rapid and constant travel is compounded by the multipronged challenges ranging from immune surveillance to viral evolution. 

Iswariya Venkataraman, PhD, associate director, scientific affairs, EUROIMMUN, a PerkinElmer company, talked to GEN Protocols about the most prominent advances in immunodiagnostic protocols in infectious disease research and development.

Serology testing for immunodiagnostics has come to the forefront due to COVID-19. Researchers and clinicians are more curious now than possibly ever before about measuring antibody levels post-infection or post-vaccination,” says Venkataraman.

EUROIMMUN has developed various ELISA tests for this purpose, targeting the S1 domain of the spike protein of SARS-CoV-2, including the receptor binding domain (RBD). These include ELISAs that can specifically detect IgA antibodies and measure the levels of IgG antibodies in serum or plasma (B-cell response), a surrogate virus neutralization test (sVNT) called NeutraLISA that detects neutralizing antibodies that inhibit the binding of S1’s receptor binding domain to host cell ACE2 receptor, and a SARS-CoV-2 Interferon Gamma Release Assay (IGRA) that helps researchers measure T cell response to the pathogen.

In vitro diagnostics, marketed directly to consumers without the involvement of a health care provider are called direct-to-consumer tests or DTC, which are facilitated through easy-to-collect samples matrixes such as dried blood spots (DBS), saliva or swabs that allow the safe and stable transport of samples via mail.

“At EUROIMMUN, most autoimmunity and infectious portfolio tests including those for COVID-19, have been adapted to accommodate DBS technology as an alternative sample matrix in addition to serum and plasma,” says Venkataraman.

Over the past few hundred years the world has been ravaged by the plague, flu, polio, AIDS, H1N1, Ebola, zika, and other viral infections. However, prior to COVID-19, technology, knowledge, and communication had never been as advanced nor was the global population as educated about infection and pathogens. Due to COVID-19, terms such as “antibody,” “antigen,” “PCR,” and “neutralization,” that were previously relegated to scientific circles, have become household terms. The scientific advances in the last few decades have made it possible to develop vaccines to the COVID-19 virus in record time, when until 2019 only 28 human diseases had effective vaccines.

Measuring serum antibody levels following vaccination has not been common practice. However, COVID-19 has resulted in researchers, clinicians, and even the general population seeking to determine their B cell and T cell immune response following vaccination or a natural infection.

Venktataraman says, “To meet the needs of such a rapidly evolving environment, diagnostic and pharmaceutical companies must market rapid diagnostic tests and therapeutic solutions.”

The plaque reduction neutralization test, or PRNT, is one of the technologies that has significantly advanced measuring the titer of neutralizing antibody for a virus.

“PRNT is the gold standard for measuring virus-neutralizing antibodies in many infectious diseases. It has also been used for determining protective neutralizing antibody response following vaccination for Japanese-encephalitis,” says Venkataraman. 

PRNT assays are generally cumbersome. They require many culture plates and five to seven days to obtain results. This affects turnaround times and outcomes. 

According to Venkataraman, “The assays also need live-virus strains, and complicated incubation and culture procedures that can be conducted only in a BSL-3 facility or higher. Moreover, there is large assay variability due limited standardization of the neutralization evaluation point. The titers can vary significantly based on the testing conditions, viral strains, passage, and cell type.”

To overcome these challenges, surrogate virus neutralization tests have been developed during COVID-19. These assays detect neutralizing antibodies against the spike protein without the need for level-3 biosafety facilities. These assays can be completed in one to three hours and do not need live virus or cells, making them accessible both for research and clinical applications. 

“Currently, the FDA has granted emergency use authorization for two of the surrogate virus neutralization tests for in vitro diagnosis of SARS-CoV-2. This is a simple ELISA test, mimicking the virus–host interaction in an ELISA plate. The plate immobilizes the ACE2 receptor. In the presence of neutralizing antibodies from the patient sample, the binding of ACE2 to the soluble Spike/RBD domain is inhibited,” says Venkataraman.

Alternatively, assays can have the specific region, S1, of the spike protein immobilized and the reagents contain the tagged-ACE2 receptor. If the sample contains neutralizing antibodies, these bind to the ACE2 receptor which prevents the binding of ACE2 to the immobilized S1 protein, and no signal is produced. Such tests help support vaccine efficacy evaluation, herd immunity estimation, and potential protective immunity assessments.

“The possibility of measuring neutralizing antibody against a virus through assays that are accessible to all scientists and researchers without the need for complex laboratory facilities, resources and equipment is a significant advancement in testing for infectious diseases during the COVID-19 era,” says Venkataraman.

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