Enzyme-linked immunoassay kit of structural protein antibody for seneca valley virus

An enzyme-linked immunoassay, structural protein technology, applied in immunoassays, viruses/phages, viruses, etc., can solve problems such as difficult clinical differential diagnosis, and achieve the effects of less antigen dosage, high sensitivity and broad market prospects.

Active Publication Date: 2017-10-17
CHINA ANIMAL HUSBANDRY IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After SVV infects pigs, although it will not cause large political and economic losses, the vesicular lesions caused are similar to those caused by foot-and-mouth disease virus, porcine vesicular disease, vesicular stomatitis, and porcine vesicular herpes. Diagnosis poses certain difficulties

Method used

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  • Enzyme-linked immunoassay kit of structural protein antibody for seneca valley virus
  • Enzyme-linked immunoassay kit of structural protein antibody for seneca valley virus
  • Enzyme-linked immunoassay kit of structural protein antibody for seneca valley virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043]Example 1, Preparation of Seneca Valley Virus Structural Protein Antibody ELISA Kit Coating Antigen

[0044] In this experiment, bioinformatics methods were used to accurately analyze the main epitopes of the Seneca Valley virus structural proteins VP1, VP2, and VP3, and suitable peptides were screened out. Four peptides were synthesized with an automatic peptide synthesizer, and the sequences were respectively As shown in Sequence 1, Sequence 2, Sequence 3 and Sequence 4 in the sequence listing, a newer and more complete coating antigen with a purity of about 80% is made, which can cover the main neutralizing antigenic epitope of Seneca Valley virus, and improve Antibody positive detection rate. The polypeptide synthesis method can be a conventional method. The following method is used in the present invention to synthesize the four polypeptides of the present invention as the coating agent of the kit of the present invention.

[0045] The coated antigen of the present...

Embodiment 2

[0074] Example 2, Preparation of Seneca Valley Virus Structural Protein Antibody ELISA Kit

[0075] Seneca Valley Virus Structural Protein Antibody ELISA Kit includes:

[0076] (1) 96-well detachable polystyrene enzyme-linked reaction plate coated with Seneca Valley virus antigen; 2×96 wells.

[0077] (2) Positive control serum: pig serum collected after artificial infection with Seneca Valley virus was used as the positive control serum of the kit (1 tube, 1.5ml / tube).

[0078] (3) Negative control serum: specific pathogen-free (SPF) pig serum, used as the negative control serum of the kit (1 tube, 1.5ml / tube).

[0079] (4) Enzyme-labeled secondary antibody: prepared by diluting 1:30000 with horseradish peroxidase-labeled rabbit anti-pig IgG (purchased from sigma company, product number A5670) as the stock solution, 2 bottles (12ml / bottle).

[0080] (5) Sample diluent: 0.01M phosphate buffer containing 0.5% (g / 100ml) casein, pH 7.4, 1 bottle (24ml / bottle).

[0081] (6) Sub...

Embodiment 3

[0087] Example 3, Sensitivity Test of Seneca Valley Virus Structural Protein Antibody ELISA Kit

[0088] 1. How to use the Seneca Valley Virus Structural Protein Antibody ELISA Kit

[0089] 1. Equilibration: Take the kit out of the refrigerated environment, and put it at room temperature for 30 minutes for later use; mix the liquid reagents before use.

[0090] 2. Dosing: dilute the 20-fold concentrated washing solution with distilled water or deionized water 20 times to obtain the washing buffer;

[0091] 3. Setting: 2 negative control wells and 2 positive control wells, and the rest are sample wells to be tested.

[0092] 4. Pre-dilution of the specimen to be tested: use the sample diluent to dilute the serum of the sample to be tested, the negative control serum, and the positive control serum at a ratio of 1:20.

[0093] 5. Adding samples: Add 100 μl of diluted samples to be tested in each well according to the preset setting. The time span of adding samples should be as ...

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Abstract

The invention discloses an enzyme-linked immunoassay kit of a structural protein antibody for a seneca valley virus. The kit comprises an elisa plate, positive control serum, negative control serum, an HRP-conjugated antibody, a sample diluent, a 20-fold concentrated detergent, a substrate solution A, a substrate solution B and a stop solution, wherein the elisa plate is coated with a structural protein epitope polypeptide composition for the seneca valley virus. The epitope polypeptide composition is one or any combination of more than two of a polypeptide as shown in a sequence 1, a polypeptide as shown in a sequence 2, a polypeptide as shown in a sequence 3 or a polypeptide as shown in a sequence 4 in the sequence table. The elisa plate is coated with a chemical synthetic antigen peptide, so that the kit is low in antigen dosage and high in sensitivity and specificity, and whether the structural protein antibody is infected by the seneca valley virus or not can be efficiently detected. The kit is high in sensitivity, good in specificity, convenient in operation, and has a good market prospect.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and more specifically, the invention relates to an enzyme-linked immunoassay kit for an antibody to a structural protein of Seneca Valley virus. Background technique [0002] Seneca Valley Virus (Seneca Valley Virus, SVV), also known as Senecavirus A (SVA), is the main pathogen of swine primary herpes disease (Swineidiopathicvesiculardisease, SIVD). After SVV infects pigs, although it will not cause large political and economic losses, the vesicular lesions caused by it are similar to those caused by foot-and-mouth disease virus, porcine vesicular disease, vesicular stomatitis, and porcine vesicular herpes. Diagnosis poses certain difficulties. Currently, SIVD has been reported in pig herds in North America, South America and Australia. In 2015, there were several SVV-infected pig herds in my country and Brazil with severe clinical and death symptoms, which caused serious economic ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/085G01N33/569
CPCC07K14/005C12N2770/32022G01N33/56983G01N33/6854G01N2333/085G01N2469/20
Inventor 董春娜张蕾张晓站肖进李静王飞巴利民张君齐鹏
Owner CHINA ANIMAL HUSBANDRY IND
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