Colloidal gold test strip for detecting nervous necrosis virus of grouper and preparation method thereof

A colloidal gold test strip, nerve necrosis technology, which is applied in biological testing, material inspection products, etc., can solve the problems of time-consuming and laborious detection methods for nerve necrosis virus, high detection conditions, and inability to detect in real time, and achieves high sensitivity, specificity, The detection method is simple and the effect of rapid detection

Inactive Publication Date: 2019-12-17
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] A colloidal gold test strip for detecting grouper nerve necrosis virus and its preparation and detection method provided by the present invention solve the problem that the existing detection method of nerve necrosis virus is time-consuming and laborious, requires high detection conditions and complicated detection procedures, and cannot The problem of real-time detection in the field

Method used

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  • Colloidal gold test strip for detecting nervous necrosis virus of grouper and preparation method thereof
  • Colloidal gold test strip for detecting nervous necrosis virus of grouper and preparation method thereof
  • Colloidal gold test strip for detecting nervous necrosis virus of grouper and preparation method thereof

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Experimental program
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Effect test

Embodiment 1

[0068] Such as figure 1 As shown, a colloidal gold test strip for detecting grouper neuronecrosis virus, the colloidal gold test strip includes a base plate 1 and a sample pad 2, a binding pad 3, and nitrocellulose that are sequentially lapped and fixed on the base plate 1 Membrane 4 and absorbent pad 5, the binding pad 3 is coated with a gold-labeled probe, and the gold-labeled probe is a thiol-modified nucleic acid aptamer at the 5' end, and its nucleotide sequence is: 5'-SH - AGCTACTGCTTTGGGGGT(sequence 1);

[0069] The nitrocellulose membrane 4 is provided with a detection line 41 and a quality control line 42, the detection line 41 is fixed with a capture probe, and the quality control line 42 is fixed with a quality control probe:

[0070] The nucleotide sequence of the capture probe is: TTCTTTTATTAGTTGATTT (sequence 2);

[0071] The nucleotide sequence of the quality control probe is: ACCCCCAAAGCAGTAGCT (sequence 3).

[0072] The preparation method of the colloidal g...

Embodiment 2

[0084] A method for detecting grouper neuronecrosis virus using colloidal gold test strip of the present invention, the detection method may further comprise the steps:

[0085] ① Virus incubation: Take 100 μl of phosphate buffered saline (PBS), add capture aptamer (Aptamer1) and amplification aptamer (Aptamer2), make the capture aptamer (Aptamer1) and amplification aptamer (Aptamer2) ) at a concentration of 200nM, and then add grouper neuronecrosis virus (RGNNV) infected cell lysates and incubate for 15min to obtain a mixture containing grouper neuronecrosis virus (RGNNV);

[0086] ②Strand displacement reaction: Add 2 μl of streptavidin (SA)-modified magnetic beads into the mixture, shake for 15 minutes to obtain Aptamer1-RGNNV-Aptamer2-magnetic bead complex, and separate Aptamer1-RGNNV- Aptamer2-magnetic bead complexes were collected, washed three times with phosphate buffered saline (PBST) containing Tween-20, and transferred to centrifuge tubes (EP tubes) for strand displa...

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Abstract

The invention, which relates to the technical field of virus detection, discloses a colloidal gold test strip for detecting nervous necrosis virus of a grouper and a preparation method thereof. The colloidal gold test strip comprises a bottom plate, a sample pad, a combination pad, a nitrocellulose membrane and an absorption pad, wherein the sample pad, the combination pad, the nitrocellulose membrane and the absorption pad are fixed on the bottom plate in a lap joint manner successively. The combination pad is coated with a gold-labeled probe. The nitrocellulose membrane is provided with a detection line and a quality control line; the detection line is fixedly provided with a capture probe; and the quality control line is fixedly provided with a quality control probe. The invention alsodiscloses a method for detecting the grouper nervous necrosis virus by adopting the colloidal gold test strip. According to the invention, the colloidal gold test strip is prepared based on the principle of combining the nucleic acid aptamer with the lateral flow biosensor and colloidal gold color development and has advantages of high sensitivity and specificity, simple assembling, and high portability; and the detection method operation is simple. The grouper nervous necrosis virus can be efficiently, accurately and quickly detected on site in real time.

Description

technical field [0001] The invention relates to the technical field of virus detection, in particular to a colloidal gold test strip for detecting grouper nerve necrosis virus and a preparation and detection method thereof. Background technique [0002] Neuronecrosis virus belongs to the beta-nodavirus genus of the Nodaviridae family and poses a huge threat to the aquaculture industry. It mainly targets the central nervous system of lower vertebrates and infects the retina and brain of fish infected by neuronecrosis virus. Obvious vacuole phenomenon can be observed. Grouper neuronecrosis virus (RGNNV), striped trevally neuronecrosis virus (SJNNV), striped star flounder neuronecrosis virus (BFNNV) and tabby puffer neuronecrosis virus (TPNNV) belong to the four genotypes of neuronecrosis virus, among which RGNNV is the worst. [0003] Grouper is a fish of great economic value. The mortality rate of larvae and juveniles can reach 100% after being infected by nerve necrosis vi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/52
CPCG01N33/52
Inventor 秦启伟刘嘉昕王劭雯曾令文俞也频李趁魏世娜
Owner SOUTH CHINA AGRI UNIV
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