Respiratory infections caused by viruses appears mainly in children, the elderly and immuno compromised patients. The major respiratory infection viruses are known as Influenza virus, Parainfluenza virus (PIV), Respiratory syncytial virus (RSVs), Entero virus, Adeno virus, etc.

In recent years, respiratory infections of Rhinovirus, Coronavirus and Metapneumo virus (MPV) have been increasing, and Boca virus has been included in the major respiratory virus tests. Corona virus is a virus that can infect animals and humans. There are six known corona viruses that can infect humans. Four of them are known as viruses that cause diseases such as the common cold, and the other two are MERS-CoV(Middle East Respiratory Syndrome Coronavirus) and SARS-CoV(Severe Acute Respiratory Syndrome Coronavirus), which have been fatal to humans.

Novel Coronavirus (2019-nCoV), which originated in Wuhan, China in 2019, has also been infected and transmitted in humans, with a 14-day incubation period, which has been reported to have a lower mortality rate and a higher incidence than SARS-CoV or MERS CoV. Sequencing of the virus revealed that the new coronavirus was 89.1% similar to bat-derived SARS (bat-derived severe acute respiratory syndrome (SARS)-like coronaviruses, bat-SL-CoVZC45, bat-SL-CoVZXC21), and 79% similar to SARS-CoV (SARs, severe Acute Respiratory Syndrome).

It is also very important to accurately diagnose Novel Coronavirus (2019-nCoV), since it has sequences similar to those of the same genus. Respiratory virus testing requires the selection of appropriate test methods depending on the characteristics of the hospital’s patient population and laboratory conditions. Antigen testing, virus culture, and molecular biological methods have been used to detect viruses. Among them, the molecular biological method using Reverse Transcriptase is highly sensitive and is recognized as a standard method for detecting viruses that cannot be cultured or exist in low concentrations. Recently, one-step RT-PCR reaction, in which reverse transcriptase reaction and polymerase chain reaction (PCR) reaction can be performed in one tube, and multiplex RT-PCR, which can simultaneously detect various viruses in one PCR reaction the introduction of the law allows for the rapid and accurate identification of many viruses. The product’s rapid and accurate detection of the Novel Coronavirus (2019-nCoV) gene allows not only to administer appropriate antiviral therapy to emergency patients, but also to reduce unnecessary antiviral use and hospitalization period, and to properly treat infected patients. It is expected to isolate the virus infected patients and block the spread of the virus in the hospital.

Principle

DiaPlexQ™ Novel Coronavirus (2019-nCoV) Detection Kit is in vitro diagnostic reagent to qualitative detection of ORF1a, N gene of Novel Coronavirus (2019-nCoV) by Multiplex OneStep qRT-PCR.

The kit includes 2X OneStep qRT-PCR Buffer, OneStep qRT-PCR Enzyme mix (including RTase, DNA polymerase and RNase inhibitor) and Primer & Probe Mixture. We also provide a Control Template (2019- nCoV), which allows us to compare the results with clinical sample results to ensure correct interpretation of the results.

Process

This test process is optimized for DiaPlexQ™ Novel Coronavirus (2019-nCoV) Detection Kit. This diagnostic kit is recommended to be carried out by professional personnel.

Sample collection

The kit can be used for nasopharyngeal swabs, oropharyngeal swabs, and sputum. Nasopharyngeal smear sample are collected from the posterior nasopharynx using a sterile swab and stored in a virus transport medium (VTM) and kept at 4 ° C ± 3 ° C for transport to the laboratory. VTM contains protein stabilizer, antibiotics for inhibiting bacterial/fungal growth, and buffer solution(Use 1~3 ㎖). Sputum samples should be rinsed with water in the morning and deeply coughed so that 1-3 ㎖ of sputum is collected in a sterile container and kept at 4 ℃ ± 3 ℃ for transport to the laboratory. Collected samples should be filled in with accurate information such as collection date, collection amount, type of sample, and patient information.

RNA Isolation

This DiaPlexQ™ Novel Coronavirus (2019-nCoV) Detection Kit is used commercial RNA prep kits, it is recommended that the RNA prep kit indicated in the table below. Prepare the total RNA from samples of patients by referring to the instruction manual of the kit that you want to use.

Multiplex OneStep qRT-PCR

1. Please thaw all reagents on the ice. After vortex, spin down.
2. Prepare PCR Master Mix by adding following reagent.
3. The amount of Master mix should be prepared by calculating additional amount corresponding to at least 1~2 reaction than the number including sample, control template (2019-nCoV), and NTC
   (Non-Template Control).
4. Mix master mix using vortex and spin down.
5. Dispense 15 ㎕ into a plate or strip tube suitable for the equipment using the manufactured master mix. (Be careful of cross contamination)
6. Add Template 5 ㎕. Labeling should be done to avoid confusion of template position.
7. After sealing with cap or film, spin down.
8. Install the prepared PCR mixture on the instrument and proceed with PCR under the following conditions.