Virus sample preserving fluid for viral nucleic acid clinical detection and use method thereof

A virus nucleic acid and preservation solution technology, which is applied in the field of virus sample preservation solution and kits, can solve the problems of false negative detection and insufficient number of new coronavirus nucleic acids, so as to ensure the integrity and quantity, improve the detection accuracy, and ensure the free of charge. affected by degradation

Inactive Publication Date: 2020-06-19
NANJING YSY BIOTECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, the inventors of the present invention found that inactivating the virus by first incubating the sample at 56°C for 30 minutes will promote the degradation of the viral nucleic acid in the sample, which will lead to an insufficient amount of the new coronavirus nucleic acid in the sample, and eventually lead to detection false negative

Method used

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  • Virus sample preserving fluid for viral nucleic acid clinical detection and use method thereof
  • Virus sample preserving fluid for viral nucleic acid clinical detection and use method thereof
  • Virus sample preserving fluid for viral nucleic acid clinical detection and use method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Example 1: The effect of inactivation treatment at 56°C on the quality of intracellular RNA in samples preserved with Trizol

[0053] 72hpf juveniles were collected for extraction of total RNA, 15 fish in each group. After the water was blotted dry, total RNA was obtained by TriZol method.

[0054] 1) Divide into 3 experimental groups in total, and each group has 3 repetitions: in the first group, add 500 μl Trizol to each repetition group, put it in a 56-degree water bath, and after 30 minutes in the water bath, use a homogenizer to homogenize (with No agglomeration is the best); in group 2, add 500 μl Trizol to each repeated group, put it in a refrigerator at 4 degrees, and after standing for 30 minutes, use a homogenizer to homogenize (no agglomeration is the best); group 3 Add 500 μl 1xPBS to each repetition group, put it in a 56-degree water bath, and after 30 minutes of water bath, blot the PBS solution dry, add 500 μl Trizol, and use a homogenizer to homogenize (...

Embodiment 2

[0066] Example 2: The impact of samples stored in different preservation solutions on the quality of intracellular RNA after inactivation at 56°C

[0067] Collect 1800 μl of human cell solution per tube, centrifuge at 1500 g for 10 minutes, discard the supernatant, keep the precipitate, collect 6 tubes in total, and use the TriZol method to obtain total RNA.

[0068] 1) The 6 tubes of cells were equally divided into 3 experimental groups: the first group was added with 500ul RNA preservation solution 1 (100mM Tris (PH7.5), 12.5mM EDTA, 150mM Nacl, 250ug / ml proteinase K), and one tube was placed in 56 put the other tube in a 4-degree refrigerator and let it stand for 30 minutes; the second group was to add 500 μl RNA preservation solution 2 (0.5% SDS, 100mM Nacl, 10mM Tris-Hcl (PH8.0), 50mM EDTA, 100ug / ml proteinase K), put one tube in a 56°C water bath for 30 minutes, and put the other tube in a 4°C refrigerator for 30 minutes; the third group was to add 500μl RNA preservation...

Embodiment 3

[0080] Example 3: Effects of inactivation treatment at 56°C on the quality of intracellular RNA in samples stored in Hanks solution and Vazyme virus sample preservation solution

[0081]HEK-293 cells (self-cultivation), porcine epidemic diarrhea virus (Procine epidemic diarrhea virus, PEDV) (Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences) and lambda DNA (Takara #3010) were used as experimental samples.

[0082]

[0083] Fully dissolve PEDV with 10mL PBS, aliquot and store at -20°C until use. Configure the mixing tube according to the above table, and divide it into 1.5mL / tube, and place it in a refrigerator at 4°C, a water bath at 56°C, and a water bath at 92°C for different treatment times.

[0084] Novizyme Virus Sample Preservation Solution (Vazyme#R503) was pre-dispensed with 200 μL of absolute ethanol into a 1.5 mL RNase-free centrifuge tube, then added 500 μL of the sample preserved in the virus sample preservation solution, and vortexe...

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Abstract

The invention discloses a kit and virus sample preserving fluid capable of being used in a viral nucleic acid clinical detection procedure including a high-temperature inactivation step, and a viral nucleic acid clinical detection method using the sample preserving fluid or the kit. The sample preserving fluid, the kit and the clinical detection method have the advantages that the integrity and the quantity of virus nucleic acid can be ensured under the condition of utilizing high-temperature inactivated viruses, so that the virus nucleic acid can be effectively prevented from being degraded;and the detection accuracy of the virus nucleic acid in the clinical detection procedure is obviously improved.

Description

technical field [0001] The invention discloses a virus sample preservation solution and a kit that can be used in the clinical detection procedure of coronavirus nucleic acid containing a high-temperature inactivation step, and a coronavirus nucleic acid involving the use of the virus sample preservation solution or kit clinical testing program. Background technique [0002] The novel coronavirus (Severe Acute Respiratory Syndrome Coronavirus 2, or SARS-CoV-2) is believed to be the culprit behind the current epidemic of Corona Virus Disease 2019, or COVID-19. Whether carrying the new coronavirus has become one of the key clinical indicators for judging whether it is infected. Including two negative nucleic acid tests for the new coronavirus is also one of the clinical criteria for the cure of COVID-19 pneumonia. However, information from many sources shows that the false negative rate of nucleic acid detection of the new coronavirus in clinical testing is relatively high. ...

Claims

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Application Information

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IPC IPC(8): C12N7/00C12N7/04C12Q1/70C12Q1/6806C12R1/93
CPCC12N7/00C12N2770/20061C12Q1/6806C12Q1/701C12Q2521/537
Inventor 赵庆顺
Owner NANJING YSY BIOTECH CO LTD
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