Are you still in shock about what happened to this World in the last 10 months? First, there was a terrifying, mysterious virus, then came PCR, swab, antibody, antigen, rapid tests, saliva, home collection kits, and finally the vaccine. The whole world revolves around the word COVID-19 and every single word related to it. There are lots of questions in mind, and most of them stem from a lack of knowledge. Do we know what these words that we use/hear hundreds of times every day mean? Are you wondering how we came from swabs that go up to your brain to collecting your own saliva at home to be tested in a lab? Well, I am here to answer all your questions about the COVID-19 infection and the testing methods. To be able to understand the testing methods better, let’s first take a look at how COVID-19 infects your body.
The general consensus is that the coronavirus spreads by inhaling contaminated droplets through person-to-person interaction, or by touching a contaminated surface before contacting with facial mucous membranes, just like common cold or influenza. Once the virus finds the way to your body, it enters through your respiratory tract and penetrates your cells with the help of a certain peptide that is recognized by the other peptides on our cell membranes. (Remember the little bulges on the surface of the coronavirus? Those are the spike glycoproteins that allow the virus to enter your cells.) Once the coronavirus membrane and the cellular membrane merge, the viral RNA enters the cell and finds the cell’s own machinery to synthesize its essential proteins and replicate itself. Viral RNA is super sneaky; it fools your ribosomes (the organelles that produce peptides) and RNA polymerases (the enzymes that synthesize RNA) to mistake it for your own RNA. Finally, when a large number of viral proteins and RNA are produced, they are packed in a particular compartment within the cell to produce new viruses, and the new viruses are released via a cellular process called exocytosis. In short, all the virus wants is to make more copies of itself, using your own cells’ machinery by manipulating it with particular signals. Below is a chart of this process for you to visualize what’s going on in the cells during a viral infection.
Figure 1: The infection process & life cycle of COVID-19.
Now that we know what COVID-19 is made of and how it infects the cells, it will be a lot easier to understand the basics of the tests. There are three types of corona tests: Molecular test (RT-PCR), antigen test, and antibody test. But what do they test? What do these tests look for?
As shown above, COVID-19 has only 3 compartment: Viral RNA, spike glycoproteins, and cell membrane. Thus, you can either look for the viral RNA with the help of RT-PCR or viral peptide using an antigen test to diagnose whether you have an active coronavirus or not, or you can search for the antibodies that your body produces to defend itself to the coronavirus.
PCR (Polymerase Chain Reaction) is a type of NAAT (Nucleic Acid Amplification Test) that looks for the genetic material, being RNA in this case. It is so far the most sensitive and accurate method to detect viral infection. With a very successful sample collection and PCR application, the presence of even 1 viral RNA can be detected. To detect the presence of the viral RNA, RT-PCR (Reverse-Transcriptase Polymerase Chain Reaction), which is a specific type of PCR that detects the presence of particular RNA sequences, is used. Here is a brief summary of the whole process: First, the sample is collected either with a nasopharyngeal swab (don’t mind the fancy name, it’s just that horrible swab) and dissolved in a viral transport medium, or directly as the saliva itself (finally, a substitution for the swab). Then, a DNA molecule is produced using the information coded in viral RNA, because the PCR cycles require a DNA molecule, and SARS-CoV-2 does not have DNA. Then, the DNA molecule is duplicated over and over with multiple cycles of DNA amplification, until millions to billions of DNA molecules are produced. Here comes the tricky part: The building blocks of DNA that are present in the solution are tagged with a fluorescence probe. The probe emits light only if it is used to produce a DNA molecule. Lastly, the sample is examined under a certain wavelength: if it emits light, then it’s positive, and if no fluorescence is detected, then it is negative. In other words, if the sample contains viral RNA, the reaction happens, and the fluorescence is detected. If no light is detected, it means that the reaction did not happen, because the sample does not contain any viral RNA. You can follow all these steps on the chart below:
Figure 2: Basics of an RT-PCR reaction
Antigen tests look for the presence of certain viral peptides. Anything that is foreign to the cell and triggers an immune response is an antigen, and can be detected by using antibodies that bind to the particular antigen exclusively. This is the basic principle of antigen tests. They are much faster and cheaper than PCR, but less sensitive. Although they might be quite specific, depending on the peptide they detect, false-positives resulting from the interactions with highly conserved peptides of other coronaviruses occur often. The biggest advantage of antigen tests is that the results come in less than an hour, unlike PCR.
Antibody tests do not detect the virus, but they detect the antibodies (defense molecules) that your body produces against the virus. If there is an antibody, there is or was the virus. There are different types of antibodies, and the presence or absence of each tells us a story about the timeline of the infection, if it exists. The details of each case are explained in Figure 3.
Figure 3: Immunoglobulin M and Immunoglobulin G & reading of the antibody test results
After almost a year of experience, we all know that having no symptoms does not mean that you do not carry the virus. We generally believe that the symptoms are caused by the virus itself, but they are actually caused by the defense mechanism of our own immune system. Thus, you might not show any symptoms at all, if the virus is having a hard time reproducing in your body, or you might show severe symptoms if your immune system is having a hard time stopping your cells to produce viral nucleic acids.
According to CDC, the best and foremost precaution to stop coronavirus from spreading is not to be exposed to the virus. But life continues, people go to work, to grocery stores, to hospitals, to malls… Since it is not possible to keep us social animals in our homes for another year, the best way to prevent the virus from spreading is to detect those who are contagious and do isolation procedures immediately. This is why PCR is extremely important in this battle against COVID-19, especially now that it is much more accessible, pain-free, and convenient with home-collection kits that do not require a swab application, and collect the person’s saliva instead.
PCR used to be performed only with nasopharyngeal swab, as it used to be believed that the sample should be collected from the back of the upper respiratory tract to obtain the most accurate results. However, a group of scientists in Tokyo, Japan reported that saliva is also a reliable alternative to nasopharyngeal swabs, after RT-PCR was run on both samples for 27 patients, and it was found that both samples have similar rates of viral positivity. The ground of this idea is that just as the upper respiratory tract, the tongue also contains high levels of receptors on its cells that spike glycoproteins of COVID-19 interact with, and the virus can be detected even before it goes down to the lungs and causes symptoms. This is much more important and promising than it sounds: Not only it promises a faster, easier, and earlier detection of the virus, but also lowers the risk of the exposure of healthcare workers and the common public to the virus.
Let’s get one thing clear: Home collection kits are not rapid home tests. They are still the same PCR test, run by the laboratories, having the same accuracy. The only difference between these kits and the regular swab tests is that you collect your own sample without leaving your home, then send the sample in a sealed envelope to the lab with a pre-prepared return label to be tested. Lab runs an RT-PCR test on your sample, just as the regular system. To obtain the same accuracy, it is crucial not to drink or eat anything 30 mins before taking the sample, to collect the sample on weekdays, while the shipping companies are working, and send it to the lab immediately. The proper collection of the sample is as important as the test itself.
Keep social distance, follow basic hygiene rules, wear a mask and get tested regularly by legitimate laboratories without even leaving your home. This way, we will get through this together!