Antibodies, genetic material, cell cultures – there are many methods for testing for Sars-CoV-2. They reveal who is sick, but maybe in the future who will be immune.
“Test, test, test” is the best cure against Covid-19, WHO Secretary General Tedros Adhanom Ghebreyesus said on March 16 at a press conference. But with which procedure at all? And what should you measure? Because there is actually more than just one test for the Sars-CoV-2. In fact, the further the pandemic progresses, the more test methods and strategies experts develop.
As a result, some of these tests make statements other than just that the virus is present in the sample. Some are faster than previous tests , with others you could theoretically test many more people than before – and some tests could even answer the question of how high the mysterious undisclosed number of undetected infected people is. However, probably only afterwards. An overview.
Rapid tests with antibodies
The greatest hopes for faster and more effective tests for Sars-CoV-2 are currently based on antibody tests . These result in a short time, which is usually very reliable – for example, this is how the classic pregnancy test works. Special antibodies are used for this purpose, which dock in a highly specific and targeted manner on surface proteins of the virus; under certain circumstances, they do this as deliberately as the DNA genetic chains. A color reaction shows whether a virus is present within a few minutes. There are similar antibody color tests for flu.
However, such tests are difficult to develop. In contrast to the short, precisely fitting genetic strands in RT-PCR, antibodies are complex molecules that cannot be easily ordered online. They are also at greater risk of binding to the antigens of related coronaviruses and also producing positive results with them. Nevertheless, many experts rely on that type of rapid test – which may be available in a few weeks and months.
Blood tests for immunity
Another type of rapid test, on the other hand, is to provide an overview of who already had the disease. Antibodies also play an important role here, but those that the immune system forms during and after the disease. Experts want to detect these immune molecules in the blood.
Using these molecules , experts want to get an overview of how far the virus has actually spread in the general population . This could, among other things, clarify the open question of the previously controversial undisclosed number of undetected infections with Sars-CoV-2. Many mathematical models suggest that it could be relatively high compared to the number of infections detected.
Different types of antibodies develop during the infection: During the infection, the immune system produces immunoglobulin M (IgM) and immunoglobulin A (IgA) as a direct reaction to the pathogen. Such an antibody test is unsuitable for reliably diagnosing the infection in acutely ill people.
The immune reaction only starts a while after the symptoms appear, so that a negative test has no meaningfulness. A positive test shows, however, that the virus was already here. Another antibody, immunoglobulin G (IgG), which only develops after infection, is interesting for another reason: it reveals permanent immunity to a pathogen.
So far, such blood tests have also had to be carried out in the laboratory. For this purpose, the biotechnologically produced surface protein of the virus is used, which binds the desired antibodies – and another antibody, which then triggers a color reaction. There are already such rapid tests that – like the pregnancy test – indicate within a few minutes with a color reaction whether the corresponding antibodies are in the blood.
Such rapid tests in particular could reveal who is already immune to the virus and who is not yet – valuable information in hospitals and other facilities where contact with infected people is inevitable. The idea of systematically testing the entire population and making those who are already immune return to normal goes even further. At the current stage, however, it is questionable whether an antibody test is sufficiently precise for such a screening. So far, none of these tests has been validated, i.e. officially checked for accuracy.
The standard: RT-PCR
The best validated and most accurate tests against the virus are the RT-PCR tests that are currently used to test for Covid-19. These tests, which were developed on the basis of the Sars-CoV-2 gene sequences, including that of a team led by Christian Drosten at the Berlin Charité, show the genetic makeup of the virus. The abbreviation RT-PCR describes the procedure for this procedure.
In the first step, reverse transcription (RT), the virus RNA is translated into DNA. This is necessary because the second step, the polymerase chain reaction (PCR), only works with DNA. Short strands of DNA, called probes, that bind precisely to individual areas of the virus genome, together with a protein called polymerase, ensure that only the genetic material of the pathogen is multiplied exponentially. When that happens, another component releases a beacon.
Which areas of the virus genome the DNA probes recognize differs from test to test; that of the Charité recognizes regions of the E gene typical of all coronaviruses as well as RdRp, the code for an enzyme that reproduces the virus genome. It is important that the selected areas are so specific for Sars-CoV-2 that the test responds to the virus searched for, but not to the viruses related to it, such as Sars or the human corona viruses.
The RT-PCR is the most sensitive of the coronavirus tests. Thanks to the exponential replication, the test responds to very small amounts of virus. This method only requires about five to ten copies of the virus RNA for positive detection. The process is based on a well-known and widely used technique that delivers the result within a few hours. If one finds the RNA of the virus in the throat of a patient with cough and fever, one can assume that it is Covid-19.
The disadvantages of RT-PCR
However, there are some pitfalls – the smear must be done correctly, which is not necessarily the case with a self-test at home . In addition, the amount of virus in the throat can fluctuate greatly during the infection – especially later, when the virus is mainly in the lungs, the cotton swab may no longer detect viruses. Conversely, the proof can still be positive after the end of the disease and pretend that the patient can infect others. But the PCR test says nothing about how contagious someone is.
It is also not possible to determine whether the genome is undamaged and the sample contains active viruses. This is also a problem for how long the virus survives on surfaces. The message that Sars-CoV-2 was still detected on the Diamond Princess after 17 days was based on the PCR method. It is therefore unclear what the finding actually means.
For the polymerase chain reaction (PCR), the samples have to be heated up and cooled down regularly.
The picture shows a thermal cycler, a device that allows this process to run automatically.
A major disadvantage of the PCR method for diagnosing the infection is that it requires several time-consuming work steps in the laboratory. Before duplication can begin, the virus RNA must be removed from the smear and cleaned. For the RT-PCR itself, the sample has to be heated and cooled several times.
Rapid tests for virus RNA
To accelerate this process, there are now several simplified variants of the RT-PCR, the cartridge tests. This also includes the rapid test presented by Bosch, which determines Sars-CoV-2 together with other viruses such as influenza. The test strip is placed in a cartridge that contains all the necessary ingredients. These cartridges are automatically processed by a separate analysis device. Although this is faster than the “manual” process in the laboratory, the capacities of the devices are usually limited.
You also still need a specially equipped laboratory. That’s why experts are looking for a real rapid test that will produce results in a doctor’s office or hospital within minutes. The U.S. Food and Drug Administration (FDA) has now approved such a test by Abbott.According to the company, the system also uses the genetic makeup of the virus . However, the RNA is not duplicated according to the RT-PCR pattern and accordingly does not have to be heated and cooled several times. Instead, in the case of a double strand of a given DNA and the matching virus RNA, one of the two strands is repeatedly cut through and replaced by a newly produced strand. This only happens if the virus has genetic material in the sample. The test is based on an existing system that has been used to detect influenza and streptococci.
Pooling increases test capacity
Not only the speed, but also the number of simultaneous PCR tests can be significantly increased under certain circumstances. To do this, combine a larger number of samples into a single sample called a pool ; if the test for the pool is negative, none of the samples involved contains the virus. So you get more tests for the effort of one. If the test of a pool is positive, all people in the pool are tested individually.
This procedure enables the same number of tests to be performed many times over the normally possible tests. However, the method is only effective if the proportion of infected people in the samples does not become too large. According to an analysis by two researchers at the Medical University of Vienna , with an infection rate of 0.1 percent, the same number of samples can be analyzed with a fifteenth of the tests.
If, on the other hand, more than ten percent of those tested are infected, the procedure becomes inefficient. However, this means that in many countries the previous samples could not be pooled effectively. Especially in severely affected countries, a high percentage of the samples are positive. In Germany, too, the proportion of positive tests has so far been between five and ten percent, so it is not entirely clear to what extent the technology can be implemented here.
Detection with cell culture
None of the tests can tell whether you are dealing with contagious Sars-CoV-2 or only with the destroyed remains of virus particles or infected cells. For healthy people, for example, where the usual tests still work several days afterwards. Does this mean that patients are still contagious? The same applies to viruses that are discovered on surfaces. Are you still infectious? To know that, you have to try it.
However, professionals do not lick the door handles , but infect a suitable cell culture with the sample. The virus destroys the cells – and you can see that in the microscope. In addition, the amount of virus RNA is determined in the liquid above the cell culture by means of PCR. With this you can clearly identify the virus. The proteins of the virus can also be detected with labeled antibodies on the surface of infected cells – under the microscope you can see not only the destroyed cells, but also the “zombie infected” who are still alive, but for whom the virus has already taken control Has.
The detection of Sars-CoV-2 in cell culture is the most accurate, but also the slowest and most complex test. For other applications, such as rapid tests on Covid-19 in hospitals, tests must above all deliver quick results on site. When in doubt, it is not so tragic when a patient is tested too positively as long as no infected people slip through. Accuracy is extremely important for screening the population, because even small errors are inflated to large inaccuracies by the high number of tests. That’s why there will never be one ultimate Covid 19 test that solves all problems.
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