It is common for health authorities to request a PCR test to confirm the positive result of a diagnostic test. Why?
The PCR test amplifies DNA fragments of any origin: viral, bacterial, etc. to levels that are detectable. When reverse transcription of RNA into DNA is performed prior to PCR, it is known as RT-PCR (Reverse Transcription- Polymerase Chain Reaction).
RT-PCR is the most widely used NAAT test to determine if some genes of the SARS-CoV-2 virus are present in a clinical sample. The previous phase of DNA synthesis from RNA is necessary, since the genome of this virus is RNA. Commonly, since the start of the pandemic, RT-PCR has been referred to as PCR.
RT-PCR of nasopharyngeal, nasal, or oropharyngeal swabs is the gold standard test for the molecular diagnosis of SARS-CoV-2 infection.
It can also be done in saliva, but in this sample the viral load is lower, so the test may have less sensitivity. However, some studies suggest that in cases of omicron infection, saliva tests could have better performance. This test can also detect the virus in other types of samples such as lower respiratory tract, blood, urine, and stool.
How does each screening test work?
TAAN tests detect genetic sequences of two or more of the structural proteins, S (spicule), E (envelope) and the N (nucleocapsid) protein, and other non-structural proteins of SARS-CoV-2.
For their part, antigen tests are immunoassays that are designed, for the most part, to detect the presence of the N protein. To achieve this, these tests carry antibodies that are developed in animals such as mice, rabbits, and even in humans. These identify the protein if it is present and when interacting with it they give a positive reaction. In antigen tests, the sample must be previously treated with a solution that releases the N protein from the rest of the viral structure, so that it can react with the antibodies that are present in the test.
Most of the antigen tests authorized in Europe detect viral antigen in nasal, nasopharyngeal and oropharyngeal swab samples, and some in saliva.
Are there false positives in antigen tests?
We speak of a false positive when the test result indicates the presence of the virus without the person being infected.
The RT-PCR technique is highly specific (99.5%), so the probability of a false positive is very low. It is also very sensitive, so much so that it allows the virus to be detected between 3 and 5 days after acquiring the infection and even weeks later. Although this viral RNA test does not always indicate current infection, it does not differentiate between active infection and resolved infection.
False positives in the RT-PCR test are usually due to sample contamination. Low viral load, especially at the beginning of the infection or during its resolution, can also give a doubtful positive result.
Antigen tests, like NAATs, are highly specific. They are comparable to the RT-PCR technique in the detection of SARS-CoV-2 infection in the first week after infection. In the nasopharynx, the highest viral load is observed mainly in the first 5 to 7 days and then decreases until it disappears. After this period of time, RT-PCR has a higher diagnostic sensitivity than the antigen test.
False positives in antigen tests are more frequent in mass screening in populations with a low prevalence of infection. They can also be due to sample contamination.
The factors that minimize the possibility of false positives in diagnostic tests are a high prevalence of infection in the community, the presence of symptoms suggestive of covid-19 in the person, and a high specificity of the test.
Confirm the result of an antigen test
The result of the antigen test must be confirmed by NAAT depending on the type of case and the probability of infection. It is considered that there is a high probability of infection if there is contact with a confirmed or suspected positive case and the person is not fully vaccinated or has not suffered from SARS-CoV-2 infection in the last 3 months.
In the community setting, in a person with symptoms suggestive of covid-19 of 5 days or less of evolution, a professional positive antigen test confirms the infection.
In general, the positive antigen test may need confirmation by RT-PCR in a situation of low probability of infection, whether or not the person is symptomatic. Also in the event of an outbreak in a social health center to confirm the first cases and in people who are hospitalized. Likewise, a positive result from a self-diagnostic antigen test should be confirmed to ensure that infection is present.
However, health authorities may consider the results of self-diagnosis tests to establish isolation and control measures.
In the case of a negative antigen test, if the person is symptomatic, it should be confirmed by RT-PCR, especially if the symptoms began more than 5 days earlier, due to the decreased sensitivity of the antigen test. Also in the case of a negative antigen test in an asymptomatic person if there is a high probability of infection.
The diagnosis of infected people is crucial for the control and reduction of transmission. It usually marks the start of contact tracing.
Despite the great efficacy of RT-PCR as a diagnostic confirmation test, sometimes a negative result may not completely rule out SARS-CoV-2 infection. If there is suspicion of infection, the test results should be evaluated in the clinical-epidemiological context of the patient and together with other diagnostic tests.
