PCR test

During the last couple of years PCR tests have become quite well-known, mostly in relation to the COVID-19. In reality, using PCR tests to diagnose diseases has been around for some time and it’s becoming increasingly more popular and diverse, allowing for better pathogen detection not just in the field of medicine, but also in other areas such as agriculture. 
 
How does a PCR test work?
 
The whole PCR test includes 4 steps: collecting a sample, extracting genetic material from the rest of the material in the sample, PCR and analysing results.
 
In case of human diseases, first a sample is taken from the patient. Sample can be taken from nose, mouth, blood, urine etc. depending on the test. For example COVID-19 tests usually use samples from the nose, HIV tests use blood samples. The idea is to collect pathogens or potentially infected cells from the patient’s body and then extract the genetic material of the pathogen, for which there are many different methods. If there is RNA (usually with viral diseases), then it’s reverse transcribed into DNA. Pathogen’s DNA is then amplified, using PCR (polymerase chain reaction), into detectable amounts. (If you are interested in getting a more detailed description of how PCR works, then read our previous blog posts.)
 
If the pathogen’s genetic material (RNA or DNA) is in the sample, then it will be multiplied by the PCR and the results can be seen. If there is no pathogen’s genetic material in the sample, then there is nothing to multiply and nothing will be seen in the results.
 
What diseases are diagnosed with a PCR test?
 
Basically every infectious disease can be diagnosed using a powerful technique called PCR (whether it is viral, bacterial, fungal or something else). With minor problems doctors usually don’t resort to PCR testing, they will just write out some medicine or antibiotics based on their experience. Pathogens that can cause severe consequences to their host body, like SARS-COV-19, HPV or HIV, are properly diagnosed using tests that also include the PCR step. Unfortunately, unlike bacterial or fungal diseases, there isn’t much that can be done in case of viral diseases, even after they have been diagnosed, except to alleviate the symptoms. So this is a great research problem for scientists that needs solving.
 
In addition to human diseases, PCR tests are just as effective when it comes to diagnosing animal or plant pathogens. Agriculture is a vital part of our lives and it’s important to identify (new) pathogens to be able to get rid of them or make plants immune to them, so that our food would have a good quality.
 
The myths about PCR test
 
In the past few years many myths have come with PCR tests. We have brought out 5 most common ones and the truth behind them.
 
  • Myth: PCR test isn’t meant to detect infectious diseases.
A myth, that has been talked about a lot, is that the inventor of PCR, Kary Mullis, never intended it to be used for detecting infectious diseases. There is no evidence though that he ever said this and it was probably just a misunderstanding or taken out of context. What Kary Mullis did think was that there are limitations in detecting the specific levels of a virus from a sample using PCR testing, but that was in 1996 and science has come a long way since then.
You can read more about it from here: https://fullfact.org/online/pcr-test-mullis/

  • Myth: it’s not accurate.
PCR tests are generally very reliable and the ones used to test pathogens are highly standardised and approved by professionals. The myth may come from the fact that the sensitivity of the PCR test is not always extremely high (depends a lot on the company producing the test), which means that a negative test doesn’t always rule out the possibility of being infected. At the same time the specificity of the PCR test is very high (close to 100%), so a positive test is almost always correct.
 
If there is a mistake, then oftentimes it’s a result of human error, not because there is something wrong with the test itself. Maybe the sample wasn’t taken correctly or something went wrong during transportation of the sample.
 
Another reason for a mistake can be the fact that PCR test doesn’t differentiate between live and dead or dying pathogen, so even if the person is technically not ill anymore, the test can still give a positive result. This ironically shows very high sensitivity and specificity of the test, because even very small parts of the pathogen are detected.
 
Of course there are also people who don’t show any symptoms while still carrying the pathogen.
 
  • Myth: PCR test can’t recognise the pathogen after it has mutated. 
In most cases the PCR test will take into account multiple genes of the pathogen, so even if part of the pathogen has mutated, it can still be recognised by the test.
 
  • Myth: PCR tests can’t tell the difference between two different types of pathogens.
Like with the previous myth, the PCR test will take into account multiple genes that are specific to the pathogen in question, so it can’t be mistaken for something else. This is also how new variants of a pathogen are detected – a change in a gene.
  • Myth: CT values used are too high.
The cycle threshold (Ct) is reached when enough double-stranded DNA has accumulated to have higher fluorescence than the background. The higher the cycle threshold (CT), the smaller amount of pathogen particles are in the sample. That being said, it doesn’t matter how high or low the CT values are, since the idea of the PCR test is to say if a pathogen is present or not. How much of it is present doesn’t really matter. Just because a person has a small amount of pathogen particles present at the moment the sample was taken, doesn’t mean that the amount of particles can’t grow in the body or that the person is not infectious to others.
 
Solis BioDyne is also keen on getting the most accurate results with PCR testing as possible and breaking all the myths. That is why you can find sensitive and highly specific 1-step RT-qPCR reagents optimised for SARS-CoV-2 detection from our product list, which have been approved by many users [1][2].
 
The SOLIScript® SARS-CoV-2 RT-qPCR Multiplex Assay Kit 2.0 is optimised for one-step RT-qPCR detection of SARS-CoV-2 viral RNA in RNA samples extracted from nasopharyngeal swabs, bronchoalveolar lavage, sputum, or any other respiratory sample. The kit is designed for multiplex detection of three distinct regions in the SARS-CoV-2 genome - the Nucleocapsid (N), Envelope (E) and RNA-dependent RNA polymerase (RdRP) genes; and the internal control - the human RNase P transcript (RPP30), all in a single reaction.
 
There is also proof from our partner diagnostic laboratory that it can detect Omicron (B.1.1.529) from clinical samples with 100% detection rate. Furthermore, the kit is 100% in agreement with LDT and Seegene CE-IVD kit. 
 
Contact the experts at sales@solisbiodyne.com.

Read more about our products to improve your SARS-CoV-2 testing: 
 
 
 
Citations
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[1] Michel J, Neumann M, Krause E, Rinner T, Muzeniek T, Grossegesse M, Hille G, Schwarz F, Puyskens A, Förster S, Biere B, Bourquain D, Domingo C, Brinkmann A, Schaade L, Schrick L, Nitsche A. Resource-efficient internally controlled in-house real-time PCR detection of SARS-CoV-2. Virol J. 2021 Jun 2;18(1):110. doi: 10.1186/s12985-021-01559-3. PMID: 34078394; PMCID: PMC8170437.
 
[2] Zrelovs N, Ustinova M, Silamikelis I, Birzniece L, Megnis K, Rovite V, Freimane L, Silamikele L, Ansone L, Pjalkovskis J, Fridmanis D, Vilne B, Priedite M, Caica A, Gavars M, Perminov D, Storozenko J, Savicka O, Dimina E, Dumpis U, Klovins J. First Report on the Latvian SARS-CoV-2 Isolate Genetic Diversity. Front Med (Lausanne). 2021 Apr 6;8:626000. doi: 10.3389/fmed.2021.626000. PMID: 33889583; PMCID: PMC8055824.