Description
A new type of coronavirus pneumonia or disease (COVID-19) caused by a new type of coronavirus (SARS-CoV-2 or CoV-19) has broken out at the end of 2019 and has spread rapidly. As of now, it has caused nearly 200 cases worldwide Over 40 million people in the world have been infected, which has caused great harm to public health security.
The COVID-19 is a positive chain single strand RNA virus, with a genome about 30kd, and is a large RNA virus. Coronaviruses belong to the order nidovales, coronaviridae, and can be divided into 4 genera (α, β, γ and δ). The incubation period of 2019-ncov was about 1-14 days, and the longest was 24 days. The clinical symptoms caused by 2019-ncov were similar to those of SARS-CoV and mers-cov, mainly fever and dry cough. Most of the patients had mild fever and no symptoms. Most of the patients with severe diseases are elderly patients or patients with severe underlying diseases. Dyspnea usually occurs one week after infection, and then develops into acute respiratory distress syndrome and sepsis. Computed tomography (CT) shows multiple ground glass opacity and infiltration in the lung, and even lung consolidation.
At the beginning of the COVID-19, the COVID-19 detection method has not yet been developed and applied, and a large number of "patients" have poured into the hospital, and the hospital lacks sufficient infrastructure and equipment, which has caused great pressure on the medical system.
In the mid-stage of the epidemic, the COVID-19 detection process has certain limitations. Most of the detection methods used in existing research technologies at home and abroad require high laboratory conditions and personnel requirements, or are prone to false negatives. It can be used in conjunction with other methods or can only be used as an auxiliary method by itself. Therefore, it is very important to develop a testing device with high accuracy, simple portability and low cost.
Existing detection technologies at home and abroad include gene sequencing, real-time fluorescent RT-PCR nucleic acid detection, digital PCR, isothermal amplification, combined probe anchoring polymerization sequencing and CRISPR detection technology. Each technique has advantages over other methods, but at the same time it also has unique shortcomings. It needs to be combined with other methods for testing to ensure sensitivity and accuracy. The problems of existing virus detection platforms include: greater pressure on the medical system, lack of medical resources deployment; cumbersome operation, larger body size, and need for supporting professionals to operate; high detection cost and lengthy time; limitations The accuracy cannot be guaranteed.
At present, in the face of such a large number of patients, in the absence of specific drugs, good detection can be timely controlled, thus cutting off the route of infection. At present, there are two directions for virus detection, one is the detection of virus specific nucleic acid, such as fluorescent RPA, This is also the main means currently used; for example, another path on the market is to do immune detection for specific structural proteins of viruses, such as immunochromatography. Due to the limitation of window period and reaction conditions, the detection steps of virus nucleic acid signal are cumbersome and false positive occurs.
At the same time, in the current situation, most of the testing pressure is concentrated on the central laboratory. If there is a large-scale testing demand, it will cause tremendous pressure on the central laboratory and medical staff, and it may also cause time delays. Similar to community hospitals and rural hospitals, there is no way to perform on-site testing and immediate testing. Central laboratories still need to perform testing. Therefore, in order to further improve efficiency and make primary medical systems such as community hospitals and rural hospitals always more powerful in detection capabilities, we designed this year's project to strive for high accuracy, portability, and ease of use.
In the bio part, we constructed a new process for detection.
Based on the COVID-19 pneumonia structure, we chose N protein as a marker for our detection. N protein sequence has high conservatism and plays an important role in the process of virus replication. N and N proteins are often used as diagnostic tools for coronavirus detection. N is a new diagnostic tool for coronavirus. N gene pseudovirus is widely used as positive control or quality control material for RNA extraction and QRT PCR detection, and it is also a powerful tool for simulation test and performance evaluation of various related kits. After obtaining the N protein, we use a nucleic acid aptamer to convert it into a nucleic acid signal, and then perform HCR or RCA amplification, and the final result is tested by colloidal gold test strips.
In order to ensure that our biological part can play a better role, we also match a special microfluidic chip and smart phone support module.
In our project, a microfluidic chip driven by a fidget spinner is used as a reaction carrier to realize the biological part. The manufacturing material is polymethyl methacrylate (PMMA), and the chip is assembled by hot-key bonding method. The overall frame of our microfluidic chip can be approximately regarded as a rectangle, the turning radius is slightly smaller than the average length of the adult hand gap, and there is a rotating shaft composed of a roller bearing and a gasket in the center of the chip. By holding the gasket in the center and rotating the chip with the other hand, the centrifugal force generated by the rotation can be used to drive the liquid path of the microfluidic chip.
According to the biological part of the detection process, this project decided to perform sample pre-processing steps such as cell lysis, release of contents, and protein signal conversion outside the chip. On the basis of the microfluidic chip, we successively realize the isothermal amplification of nucleic acid in the isothermal amplification area, and realize the visual signal conversion based on the lateral chromatography test strip in the signal detection area. Finally, the mobile phone support module optically detects the signal on the test strip, and performs subsequent signal processing. Specifically, on the test strip, our project adopts a signal conversion method based on colloidal gold technology, and compares the colloidal gold degree with the standard positive result through the color reaction to obtain the reaction result.
The temperature control module is mainly composed of a single-chip microcomputer, a ceramic heater and a thermistor. The combination of these three can meet our temperature requirements for HCR and RCA reactions.
In the optical signal visual detection part, we use the MINA framework to build a WeChat mini program to take pictures and upload the colloidal gold test strips, as well as reading the colloidal gold image with the software, and fitting the relationship between the image characteristics and the concentration of the test object, and finally get the virus test results.
Through the use of nucleic acid signaldetection, coupled with the necessary hardware and smart phone functional modules, we hope to be able to initially try to achieve our goal, and further improve afterwards, so that the final production is more accurate, portable and easy to operate. It can be applied to detection devices in communities, villages and other places.
Created By Beijing Institute of Technology