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Reagents that claim to achieve rapid detection within 30 minutes or even 15 minutes are mostly sacrificing some performance in exchange for speed improvements.

Since the outbreak of the new crown, due to factors such as the number of kits and laboratories, a large number of suspected patients in Wuhan cannot be diagnosed.

Recently, Wuhan has performed a “zero” nucleic acid test on all suspected patients. The test capacity has been increased from 200 copies per day to 8,000 copies per day. At the moment of the epidemic, a large number of suspected patients and asymptomatic infections are waiting to be diagnosed, which makes urgent demand for related testing products

On February 3, 2020, a biotechnology company in Jiangsu Province, the production staff packed the finished reagent. Published by China News Agency, photo by Tang Dehong

On February 8th, the Ministry of Science and Technology released “Research on New Coronavirus (2019-nCoV) (“Guide”) for the purpose of issuing emergency project declaration guidelines, aiming to break through the limitations of existing testing technologies on personnel, testing sites, and other conditions To improve the efficiency and convenience of accelerated detection, it is conducive to the rapid diagnosis of suspected patients and the on-site screening of close contact with the crowd.

The emergency quick inspection products collected by the Ministry of Science and Technology all require product development to be completed within one to two weeks, obtain a clinical certificate at the latest one and a half months, and list relevant assessment indicators.

A pharynx swab sample is placed in a small POS-sized nucleic acid tester. Press the start button and the test result will be available in half an hour. On-site rapid inspection (POCT) is also called “bedside detection” , which relies on portable instruments and reagents, customs Epidemic detection personnel, grass-roots epidemic prevention and control personnel, and some community workers can quickly screen suspected cases at the sampling site.

Compared to the complicated process of real-time fluorescence PCR (Polymerase Chain Reaction) , POCT can be separated from the laboratory and does not require wearing Professionals in the third-level protective clothing operate carefully, as long as they follow the instructions, a short-trained general inspector or nurse can do it. The entire testing process can save at least 3-4 hours. Successful rapid initial screening will lay an important foundation for hierarchical diagnosis and treatment across the country.

There are three main types of new on-site rapid detection of coronaviruses collected by the Ministry of Science and Technology, namely Nucleic acid on-site rapid detection equipment and reagents, rapid antigen detection reagents and Antibody rapid detection reagents < / strong>.

A researcher from the State Key Laboratory specializing in nucleic acid detection told us that the technical principles of rapid nucleic acid on-site detection and real-time PCR are the same, that is, Gene amplification . A piece of RNA is reverse transcribed into double-stranded DNA, and then amplified by DNA polymerase. The gene fragment carrying the virus is continuously amplified and then captured by a fluorescent probe. When the amplified virus concentration reaches a critical value Will produce a fluorescent signal, meaning that the sample was detected to carry the new crown virus.

ResearchThe investigator pointed out that compared with the traditional PCR detection in the laboratory, the integrated on-site rapid detection, although portable and simple to operate, , if its emphasis is placed on its “fast” characteristics, its sensitivity may be affected.

For example, in the assessment indicators for rapid nucleic acid detection products listed in this Guide, the detection rate is required compared to traditional real-time fluorescent PCR reagents. (Sensitivity) is not less than 95%. “This is equivalent to artificially reducing the sensitivity requirements for it. In theory, the rapid detection of nucleic acids should be the same as the sensitivity of real-time fluorescent PCR.” The researcher said.

In addition, a rapid on-site detection of nucleic acids The sample size that can be detected is very limited , the number of samples may be as few as 7-8, and as many as a dozen. The PCR instrument used in the laboratory can generally detect 96 samples at a time, and up to 384 samples.

Unlike on-site rapid nucleic acid detection, both rapid antigen detection reagents and antibody rapid detection reagents belong to immunoassay . The principle of combining viral antigens and antibodies is collected by collecting serum from patients.

Antibody detection is the use of antibodies to detect the presence of viral antigens in patients. Antigen detection Viral antigens are used to detect whether specific antibodies have been elicited in patients.

An expert in disease control systems specializing in nucleic acid detection pointed out to China News Weekly that immune detection is faster than nucleic acid detection. The former usually yields results in 15-30 minutes, but the sensitivity is not as good as nucleic acid. Detection .

For example, the assessment criteria in the Guide states that the requirement for rapid antigen detection reagents is that the detection rate can reach more than 80% of nucleic acid detection. Equivalent to nucleic acid detection can detect 100 positive samples, antigen detection requires only 80 detection. “In fact, this goal is not easy to achieve,” the expert said with emotion.

The aforementioned researchers also said that, on the technical level, it is also the amplification of viral gene fragments, and nucleic acids can be amplified more than 1 million times, which is far greater than the amplification effect of immunity. This means that although immunoassay is fast, the detection rate of samples with low concentration of virus is low, which may cause missed detection.

In addition, compared with real-time fluorescent PCR The specificity of antigen detection is poor, which can cause false positives.

Previously, a medical device company in Zhejiang Province used the “colloidal gold method” to develop a test strip that can quickly screen for coronavirus, which can produce results in ten minutes. The research and development staff of the product pointed out that this test strip is a broad-spectrum preliminary screening for coronavirus, not just for new coronavirus.

“Any detection of a gene fragment is the most direct, and the detection of immunity is indirect,” said the aforementioned expert.

A number of experts told China News Weekly that, whether it is a rapid on-site test or a real-time PCR test that requires instrumentation in the laboratory, Under the existing technology, the time reduction is generally to a certain extent Sacrifice sensitivity. In other words, false negatives are more likely to occur.

Some medical device companies in Shandong, Shanghai, Tianjin and other provinces have recently launched new coronavirus detection kits that can be detected within half an hour. It is understood that the relevant kit has entered the emergency approval channel of the State Drug Administration.

Researchers from the aforementioned State Key Laboratory specializing in nucleic acid detection told China Newsweek that at present, real-time fluorescent PCR detection takes as fast as 1 to 1.5 hours. Because the fastest time to extract nucleic acids is 30 to 40 minutes, and the amplification time is at least 40 to 60 minutes.

A technical expert from another kit manufacturer also said frankly to “China News Weekly” that claims that reagents that can be quickly detected within 30 minutes or even 15 minutes are mostly sacrificing some performance in exchange for speed Promotion.

The aforementioned disease control experts point out that for molecular genetic diagnostic technology, specificity and sensitivity always need to be balanced. The stronger the specificity, the lower the sensitivity; if the sensitivity is good, the specificity will be weakened. The choice of different target sites, primers and probes, and even the design of the reaction system are to better balance these two characteristics.

From his point of view, from a technical point of view, in terms of the balance of sensitivity and specificity, compared with some rapid detection methods, The best balance is still the real-time fluorescent PCR detection kit.

From the past major epidemics, whether it is West Africa Ebola virus, influenza A or Middle East respiratory syndrome (MERS) , fluorescence Quantitative