Background and Current Events

Foreign invaders gain entry to the human body. They launch an offensive and spawn infection. The human immune system conjures a defense, launching innate and adaptive immune responses. The battle wages on until the defeated is vanquished and the victor looks on – ready for the next invasion.

This simple analogy of the immune response to infection is a basic intro to the complex problem the world is currently facing. Detecting invasion by the SARS-COV-2 virus (the causative agent of COVID-19) is an important first step in identifying pathogen exposure. Measuring the human immune response is the critical next stage to quantify true infection rates and to decipher the development of immunity. The current COVID-19 outbreak has demanded unprecedented action and the advent of innovative testing technologies in the fight towards immunity.


Testing for Virus

First line measures during an outbreak include testing for detection of the pathogen. In the case of COVID-19, the majority of these tests involved detection of the viral RNA by use of the real-time PCR method. Real-time PCR can detect minute amounts of virus, and therefore can excel at sensitivity, and it can also detect unique markers within samples, and therefore be highly specific. Since SARS-CoV-2 is a retrovirus or RNA virus, the test is actually termed real-time reverse transcriptase PCR.

The test samples require careful, clean, and contaminant-free preparation. This stringency, coupled with the length of test turnaround (several hours) and the specialized equipment and methods involved, created initial challenges to the deployment and accessibility of these tests. Other technologies have emerged and a concerted effort (presently over 60 tests have been approved by the FDA for emergency use) has pushed virus testing towards its potential on the front lines of the COVID-19 battle.


What is a Serology Test?

A second type of test focuses on the host response, as opposed to direct detection of the virus itself. This level of surveillance can be highly informative and a test for immune response, otherwise known as a serology test, can confirm or refute exposure.

Description of how a serology test works first requires definition of key terms.

Serology is the scientific study of blood serum and other body fluids. The term is most often associated with the detection of antibodies in serum of patients who are exposed to microorganisms, although other conditions including autoimmune diseases can result in antibody responses.

The innate immune response is a first line of protection which involves physical barriers such as mucous membranes and immune cells including neutrophils, macrophages, cytokines, and others.

The adaptive immune response is more specific and involves targeting towards invading antigens using dendritic cells, T- and B-cells, and antibodies. Although this response takes longer to activate, the components involved interact directly with target antigens and form a strong defense against invaders.

Antibodies (also called immunoglobulins) are proteins that are secreted by immune activated B cells and plasma cells which recognize distinct sections of the invader proteins (antigens). Antibodies are Y-shaped proteins with an antigen-binding variable region (Fab) at each tip and an Fc region at the stem which allows communication with other immune factors. There are five isotypes based on five different types of Fc regions: IgM, IgA, IgG, IgE, and IgD

The antibody response is initiated as part of the adaptive immune response and involves activation of B cells and plasma cells which secrete antibodies specific to invading antigens. There are two antibody types, those that freely circulate in blood plasma and those that are membrane-bound on the surface of B cells and involved in further amplifying antibody production in response to antigen. Antigens, after being bound by antibodies, can no longer function as needed during pathogen invasion. Antibodies also bind and present antigen-bearing cells to immune cells such as phagocytes and natural killer cells for clearance and activation of downstream immune processes.


How does a Serology Test Work?

Serologic tests measure the presence and abundance of certain types of antibodies, including: IgM or IgG alone, IgM and IgG together, or IgM, IgG, and IgA antibodies also called complete antibody tests.

  • IgM antibodies, also known as surveillance antibodies, appear within 10 days after exposure to the invader.
  • After identifying foreign antigens, IgM antibodies mature into IgG, or attack antibodies, waging war by attaching to the receptors and surface proteins of the invading virus.
  • IgG antigen binding prevents the virus from gaining entry to host cells and staging further infection.
  • In time, IgA antibodies arrive to further attack and push back the virus towards exit of the body.

Serology tests work on blood samples rather than nasal or throat swabs as with virus detection tests. Traditional test methods include enzyme-linked immunosorbent assay (ELISA), immunoprecipitation, fluorescent or chemiluminescent detection or a number of other detection techniques.

The COVID-19 pandemic has spurred development of several new testing techniques and modalities, as evidenced by the growing list of FDA Emergency Use Approved tests. ELISA-types tests have been designed with immobilized antigen, such as the outer spike protein from the virus coat, embedded on a binding surface. Blood serum is incubated with the immobilized antigen and any free antibodies bind with high specificity, after which a fluorescent substrate is applied and samples are detected for positive (or negative) binding activity.

A number of tests have been built based upon microfluidics, rapid-detection by lateral flow, and other technologies. Although the overall response to test development has been prolific, some of these tests have shown deficiencies in sensitivity and specificity in the field, and as a result, the FDA has begun to vet tests using internal validation measures.


Why is a Serology Testing Important?

Serology testing is important to identify an initial immune response to virus exposure. Perhaps a person did not incur symptoms and therefore did not undergo virus testing, a serology test would verify if exposure had indeed taken place. A negative serology test, when performed correctly, would indicate the lack of any exposure, and susceptibility of that person to future exposure.

Although antibody response can be an accurate method to identify virus exposure, the appearance of virus antigen-specific antibodies alone does not necessary equate to immunity. Subsequent or sustained exposure to the virus, and the generation of strong neutralizing antibody and adaptive immune responses, primes the system to recognize subsequent encounters with the foreign invader. In general, this is the basis of long-term immunity.

This is also the main limitation of serology testing. Is the test measuring short-term response or rather long-term immunity? In order to better clarify that question, a more comprehensive picture of what long-term immunity looks like, and how it is achieved, must be gained.

A second, and equally important question with the current COVID-19 outbreak is whether long-term immunity can be achieved at all, and if so, how long will last before the adaptive response wanes. A growing body of evidence suggests that reinfection is not possible, at least in the near term – indicating immunity is indeed a possibility.


Outlook

Serological testing is important in determining the prevalence of the virus in the community. As such, seroprevalence testing will help in determining the true rates of exposure and perhaps the progression of the population towards herd immunity. Overinterpretation is a real danger however, and the details behind long-term protective immunity and other factors must be seriously considered before planting the victory flag. To fortify the fight, the FDA has very recently created an umbrella emergency pathway for serology tests, to enable only those that attain independent validation to be used for approved testing. These and other measures may further increase the odds in the battle against COVID-19.


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