What is the meaning of PCR nucleic acid detection, methods and steps
Nucleic acid testing is currently an effective way to determine whether the virus is carrying the new coronavirus. At present, people mainly use nasal swabs, throat swabs and antibody serum lgm for reference. But according to the latest Singapore entry requirements, PCR nucleic acid testing is performed, so what does PCR nucleic acid testing mean?
What is the meaning of PCR nucleic acid detection
PCR means polymerase chain reaction, which can be used to amplify DNA, so as to amplify a small amount of DNA to the extent that it can be detected. The nucleic acid of some viruses is DNA, and this method needs to be used for detection. Therefore, PCR is not a check performed, but a method of detecting DNA. For example, hepatitis B virus DNA quantification uses this method. The purpose of DNA inspection is to diagnose the disease on the one hand. If even the DNA of the pathogen can be detected, it means that the pathogen is infected; on the other hand, it is to judge how much the virus has replicated, so as to judge the severity of the disease or the effect of treatment.
PCR nucleic acid detection methods and steps
Nucleic acid testing requires five steps: sampling, sample retention, retention, nucleic acid extraction, and computer testing.
The first step in nucleic acid detection is to collect human secretions, and wipe the nasal cavity or back wall of the pharynx and bilateral pharyngeal tonsils with a nasal test or a pharyngeal test.
In the second step, the medical staff kept the sample, immersed the test piece head in the cell preservation solution, and screwed the tube cap immediately after breaking the tail.
In the third part, put the sample in an airtight bag, keep it and send it for inspection in time.
The fourth step is to send the sample to the laboratory to extract nucleic acid.
In the fifth step, the extract is subjected to a fluorescent PCR amplification reaction.
What is nucleic acid
Nucleic acid (nucleic acid) is a type of biological macromolecule composed of nucleotides (nucleotide), divided into ribonucleic acid (RNA) and deoxyribonucleic acid (DNA), in which RNA is mostly single Chain structure, DNA is mostly double-stranded structure. In addition to prion, nucleic acid is a biological macromolecule necessary for life. Its main function is to constitute the genetic information carrier of life. In addition, some nucleic acids can be used as and participate in the formation of biologically active enzymes. Molecules or other molecular machines.
What is nucleic acid testing
Nucleic acid detection, as the name implies, is to detect nucleic acids.
What are the unique advantages of nucleic acid testing? Why can it be used as the "gold standard" for the diagnosis of new coronavirus?
In fact, every detection method has its own application scenarios. Taking virus detection as an example, the detection object of the usual immunological detection methods (colloidal gold, ELISA, chemiluminescence, etc.) is the antigen or antibody of the virus, which is equivalent to "side drawing". Since there is a time window from infection to detectability, immunological detection methods are generally not suitable for early diagnosis.
What is the meaning of PCR nucleic acid detection, methods and steps
False positives and false negatives
1. False positive
There are usually fewer false positives. Generally, improper operation of laboratory personnel can lead to cross-contamination between samples or residual contamination of amplified products. In this case, false positives may be caused.
However, false positives can be effectively circumvented by strictly controlling the detection process and implementing a strategy of randomly participating in the detection of several negative samples.
2. False negative
What needs to be explained here is that false negatives in PCR detection and clinical false negatives are actually not the same concept.
Taking the diagnosis of the new coronavirus, which is discussed intensely on the Internet, as an example, clinical false negatives refer to cases where the clinical symptoms and imaging studies are highly suspected of being infected, but the PCR test is repeatedly or always negative; and the PCR test false negative refers to the collected There is a sufficient amount of the new coronavirus in the sample but it has not been detected.
To avoid false negatives in nucleic acid testing, the main needs to be solved are: (1) there is sufficient virus in the cells of the infected person, (2) cells containing the virus can be collected in the collected sample, and (3) the virus in the sample is detected These three links.
Among them, the technical parameters of PCR kits mainly affect the third detection link.
Only for the third test link, if the number of viruses in the sample is below a certain level (below the minimum detection limit), the PCR kit will not be able to detect it. From this perspective, false negatives in PCR detection cannot be completely avoided. This is why it is necessary to supplement the development of virus-specific antibody testing.
