Testing is one of the key factors in combating the spread of COVID-19 infection worldwide. The World Health Organization has also emphasised its importance, calling for as many people as possible to be tested.

When COVID-19 first appeared, testing was essential for rapidly detecting and isolating affected individuals and calculating accurate infection and survival rates, which are crucial for appropriate epidemiological measures. As time passed and knowledge about the infection grew, the basic principles of testing were joined by an increasing need to identify asymptomatic people as well as those who had recovered and developed immunity.

At the beginning of the illness, the virus is present in the respiratory system and can be detected in a nasopharyngeal (the nasal part of the throat) and/or throat swab using direct molecular virological methods, including RT-PCR (real-time polymerase chain reaction). At the start of the epidemic, most countries relied on RT-PCR tests because these were the fastest to develop.

RT-PCR is a highly sensitive and specific method that can reliably diagnose the presence of viral RNA within just three hours. However, RT-PCR testing requires a specialised laboratory, trained personnel and time. Although the testing itself takes only a few hours, sample collection, transport and processing must also be considered, so several days may pass before a result is available.

The second group comprises serological tests, which detect antibodies to the SARS-CoV-2 virus and help determine its spread in the population.

The importance of serological tests lies in monitoring the development of immunity both in asymptomatic patients (infected people showing no signs of illness) and those with a clear clinical picture of COVID-19 infection.

The body produces different antibodies against many viral components. Some antibodies are very useful, killing the virus or stopping infection, while others are less useful because they bind to shared viral components and do not help defend the body.

A key target of human antibodies against SARS-CoV-2 is the so-called S protein, or “spike”, on the viral surface. Serological tests measure IgM, IgG and IgA antibodies. IgM antibodies are the first to appear in response to viral infection. They usually emerge towards the end of the first week after illness begins, during the acute phase, and then disappear. IgG antibodies appear shortly after IgM, reach high levels during the acute phase and fall to lower levels after recovery, remaining present for a longer period. IgA antibodies show a similar pattern to IgM.

This makes it clear that the timing of serological testing matters: testing too early may produce false-negative serological results even though a person is infected.

Serological tests must also be checked on a range of different samples, not only to establish accuracy but also to determine when a sample becomes positive after infection begins and how long it remains positive after recovery, because each person produces a unique set of antibodies.

Serological testing uses two methods: rapid immunochromatographic tests and conventional laboratory enzyme immunoassays. Rapid tests use capillary blood and results can be read within minutes, whereas conventional serological enzyme immunoassays (ELISA) use venous blood and results are read after 3-4 hours.

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Although immunochromatographic tests can be performed at home and results read very quickly (similarly to pregnancy tests), they require considerable caution. Unverified and unapproved tests may produce a positive result without SARS-CoV-2 infection because the person may have antibodies to a similar viral component; a negative result can also be read simply because the lines are faint.

Other methods include an immunochromatographic test detecting viral proteins in a nasopharyngeal swab within 10-15 minutes. The test is not yet widely available and does not help monitor immunity development. The urgent current need for tests has encouraged the development of a range of new, innovative microbiological and laboratory tests.

Whether molecular or serological tests are used, all tests must be validated. Validation is a set of procedures used to check a test’s accuracy and reliability.

Most importantly, none of the currently used tests should be viewed in isolation, and no test excludes the other. While molecular techniques help identify newly infected individuals, serological tests help monitor people who either show no signs of illness or have recovered.

As time passes, the importance of tests will naturally change, and serological tests will become increasingly significant because they will show trends towards a slowing pandemic and the development and duration of immunity. This underlies vaccine development and the need to vaccinate the population, all with the aim of returning to normal life.

Sources:

1. Corman VM et al. Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR. Euro Surveill 2020; 25(3): 2000045.
2. Xiao SY et al. Evolving status of the 2019 novel coronavirus infection: Proposal of conventional serologic assays for disease diagnosis and infection monitoring. Journal of Medical Virology 2020; 92(5): 464-7.