Polymerase chain reaction (PCR) test plays a key role in the detection and diagnosis of various infectious diseases, including the global fight against the COVID-19 pandemic. However, it is not foolproof and false positives/negatives can occur. The problem of false positives in PCR is highly valued, but the problem of false negatives in PCR is more serious. In fact, the problem of false positives in PCR is relatively simple, and generally only involves pollution and non-specificity of primers. Pollution can be basically solved through strict laboratory management, reasonable environment settings, and adding UNG enzyme anti-pollution, while the non-specificity of primers basically belongs to the quality of the manufacturer's reagents. However, the false negative of PCR is different. It involves almost all personnel and technical links related to PCR experiment, which is very complicated.
False Negatives in PCR Tests
False negatives occur when a PCR test fails to detect the presence of a pathogen or fails to amplify the target genetic material sufficiently. The repercussions of false negatives are profound, as they can lead to delayed or incorrect diagnoses, inadequate treatment, and inadvertently spreading the infection further. The multifaceted nature of false negatives makes it imperative to identify and address the contributing factors comprehensively.
Factors Contributing to False Negatives
First of all, the development of the kit requires more than a dozen key chemical raw materials such as enzymes and synthetic DNA. Different companies choose different suppliers of raw materials, and the activity of enzymes and the purity of DNA will vary, which will affect the accuracy of nucleic acid detection reagents. In the process of kit product development, the preparation of raw materials, the selection of production process, the ability of R&D personnel, and the use of technical means may cause deviations in each link, and the accumulation of errors will also lead to differences in the sensitivity of kit products.
Secondly, the irregular sampling process will also affect the subsequent separation and determination results, resulting in false negative test results. Taking the most commonly used and easy-to-operate throat swab collection as an example, due to the low content of the new coronavirus in the throat, it may cause missed detection. Although it is only a few strokes with a cotton swab on the pharynx, if there are no good professional practical skills, the sample collection site and method may not meet the specifications, which will directly affect the test results.
Third, the detected viral RNA is very easily degraded by the environment, making the detection false negative, and the newly infected patient may also have a negative test result due to the limited amount of virus in the body.
In addition, the test samples must be transported in a cold chain. If the samples are damaged during the transport, the test results will also be affected. Moreover, nucleic acid testing has high requirements in the testing environment. Once the virus testing laboratory is contaminated, it will also lead to false negative test results.
How to Reduce False Negatives?
Although optimizing the timing and frequency of testing, improving the skills of the sampler, increasing the sensitivity and specificity of the test and integrating complementary diagnostic methods can reduce false negatives in PCR testing, the possibility of false negatives cannot be completely eliminated. Therefore, diagnosis should be made in conjunction with clinical symptoms, epidemiological factors and other diagnostic modalities when necessary.
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