Verification experiment of inactivation effect of Virus Transport Media

There are two types of Virus Transport Media added in the virus sampling tube, one is the inactivated preservation solution of the modified lytic virus protein extraction nucleic acid, and the other is the maintenance of the virus in vitro activity, its nucleic acid and antigen modified based on the transport medium Complete non-inactivated preservation solution. For inactivated preservation solutions, it is important to inactivate viruses efficiently and prevent secondary infection.

Different from the non-inactivated type, the inactivated Virus Transport Media is added with a high concentration of lysis salt, which can inactivate the virus efficiently and can effectively prevent the operator from secondary infection. But it also contains Rnase inhibitors, which can protect the viral nucleic acid from degradation, so that it can be subsequently detected by NT-PCR. The inactivation effect is experimentally verified below.

1. Inactivation verification materials

1 SPF chicken embryo (and hatched to 10 days old by itself)

2 Chicken infectious bronchitis virus IBV QXL87 strain

3 Normal saline (0.9% NaCl), autoclaved

4 Inactivated sample storage solution, 3 batches.

5 RNA extraction kit

Virus Transport Media inactivates viruses

2. Experimental methods

1. Add the prepared chicken infectious bronchitis virus QXL87 strain to the preservation solution, according to the ratio of 1 (viral solution): 10 (preservation solution), and set the room temperature at 18-26℃ for 45min. The virus fluid was inoculated into SPF chicken embryos, and allantoic fluid was harvested.

2. Inoculate the allantoic fluid harvested in 1 into 10 days old SPF chicken embryos according to the allantoic cavity inoculation method. Each sample is inoculated with 10 chicken embryos, 0.1 mL/piece, placed in a 37°C incubator for 144 h and discarded. After 24 h of dead chicken embryos, observe and record 24-44 h of chicken embryo deaths and the number of diseased live embryos after inoculation. Observation of chicken embryo lesions, and the detection of chicken infectious bronchitis virus nucleic acid on the harvested allantoic fluid, referring to GB/T 23197-2008 chicken infectious bronchitis diagnostic technology, using RNA extraction kit to extract RNA, and using one-step method Real -Time RT-qPCR for virus detection.

Group 1/2/3 added virus (100ul) + preservation solution (900ul); Group 4 added virus (100ul) + physiological saline (900ul); Group 5/6/7 added preservation solution (1000ul). Among them, the Virus Transport Media is in three batches.

3. Experimental results

According to the above experimental results, groups 1, 2 and 3 were inoculated with virus-containing preservatives, which showed that chicken embryos grew normally; group 4 was inoculated with virus-added physiological saline, and chicken embryo lesions; groups 5, 6, and 7 were inoculated with preservatives, showing No inhibition of chicken embryo growth. This indicates that the inactivated preservation solution can inactivate viruses.

Through this experiment, we can finally show that the three batches of random sampling of Desheng inactivated preservation solution can effectively inactivate the virus, and it has no inhibitory effect on the normal physiological function of the cells. Therefore, this inactivated Virus Transport Media It can efficiently inactivate various viruses and extract nucleic acids, which is suitable for RT-qPCR nucleic acid detection experiments. Of course, in view of the faster testing, which is directly used for the detection of patients diagnosed with New Crown, few companies in China will do so, because after all, New Crown virus is not so easy to obtain!