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Foot and mouth disease virus non-structural protein antibody enzyme-linked immunodetection kit

A foot-and-mouth disease virus and non-structural protein technology, applied in the field of foot-and-mouth disease virus structural protein antibody ELISA kit, foot-and-mouth disease virus structural protein VP1 antibody ELISA kit field, can solve the problem of affecting the accuracy of the test results and cannot be used as a test Index, can not be used as a detection antibody and other issues, to achieve good market prospects, less antigen consumption, high sensitivity and specificity

Active Publication Date: 2014-06-18
CHINA ANIMAL HUSBANDRY IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Virus-infected animals can produce anti-2B antibodies, but this antibody has poor reproducibility in ELISA reactions and cannot be used as a detection antibody
2C antibody fades faster than 3ABC, and 2C antibody can be detected in immunized cattle, so it cannot be used as a detection indicator
Such prokaryotically expressed protein products contain a small amount of carrier proteins and host mycoproteins, and it is difficult to completely remove them. These miscellaneous proteins will produce non-specific binding with the test sample, thus affecting the accuracy of the test results.

Method used

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  • Foot and mouth disease virus non-structural protein antibody enzyme-linked immunodetection kit
  • Foot and mouth disease virus non-structural protein antibody enzyme-linked immunodetection kit
  • Foot and mouth disease virus non-structural protein antibody enzyme-linked immunodetection kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Embodiment 1, preparation of coated antigen by enzyme-linked immunosorbent assay kit for nonstructural protein antibody of foot-and-mouth disease virus

[0046] The present invention adopts the method of bioinformatics to accurately analyze the epitope of the nonstructural protein, and screens out the peptide segment suitable for ELISA detection from the main epitope on the 3B protein, that is, the nonstructural protein 3B epitope polypeptide of foot-and-mouth disease virus , and its sequence is shown in sequence 1 in the sequence listing. The polypeptide is used as the coating source of the kit of the present invention to prepare the non-structural protein ELISA kit for foot-and-mouth disease virus.

[0047] The coated antigen of the present invention can be prepared using an Applied Biosystem automatic polypeptide synthesizer (model 433A). Using the Merrifield solid-phase synthesis method, Fmoc (9-fluorenylmethyloxycarbonyl, 9-fluorenylmethyloxycarbonyl) modified ami...

Embodiment 2

[0078] Embodiment 2, the composition of foot-and-mouth disease virus nonstructural protein antibody ELISA kit

[0079] Foot-and-mouth disease virus non-structural protein ELISA kit includes:

[0080] (1) 96-well detachable polystyrene enzyme-linked reaction plate coated with foot-and-mouth disease virus antigen; 2×96 wells.

[0081] (2) Horseradish peroxidase-labeled rabbit anti-pig IgG polyclonal antibody (purchased from sigma, Cat. No. A5670), 2 vials (12ml each).

[0082] (3) Positive control serum: using the coated antigen artificially synthesized in Example 1 as the immunogen, the hyperimmune serum prepared by immunizing 25-35-day-old specific pathogen-free (SPF) pigs was added with 1000 U / ml streptavidin and 1000U / ml penicillin, sterilized by 0.2μm filter membrane, used as the positive control serum of the kit. 1 tube (1ml).

[0083] (4) Negative control serum: use normal pig serum without FMD antibody, add 1000U / ml streptomycin and 1000U / ml penicillin, pass through a...

Embodiment 3

[0090] Embodiment 3, the detection method of foot-and-mouth disease virus nonstructural protein antibody ELISA kit

[0091] 1. Equilibration: Take the kit out of the refrigerated environment, equilibrate at room temperature for 30 minutes before use, and mix the liquid reagents before use.

[0092] 2. Dosing: Dilute the concentrated washing liquid 20 times with distilled water or deionized water for later use.

[0093] 3. Setting: leave 1 blank well, 3 negative control wells, and 2 positive control wells, and the rest are sample wells to be tested.

[0094] 4. Pre-dilution of the sample to be tested: add 100 μl of sample diluent to each well of the dilution plate, and add 5 μl of the sample to be tested respectively.

[0095] 5. Adding samples: do not add samples to blank wells; add 100 μl negative control to each of the 3 negative control wells, add 100 μl positive control to each of the 2 positive control wells; add 100 μl diluted samples to be tested in the remaining wells...

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Abstract

The invention discloses a foot and mouth disease virus non-structural protein antibody enzyme-linked immunodetection kit. The kit comprises a foot and mouth disease virus non-structural protein 3B antigen epitope peptide-coated polymerase chain reaction plate and an enzyme-labeled antibody, wherein the foot and mouth disease virus non-structural protein 3B antigen epitope peptide is a polypeptide shown as a sequence 1 in a sequence table. The kit adopts a chemical synthesis non-structural protein 3B antigen peptide coated reaction plate and is small in antigen amount, high in sensitivity and high in specificity, and whether the foot and mouth disease virus infection exists can be efficiently detected. The kit is high in specificity, sensitive and high-efficiency and has good market prospects.

Description

technical field [0001] The invention belongs to the field of biotechnology. More specifically, the invention relates to an enzyme-linked immunosorbent detection kit for an antibody to the structural protein of foot-and-mouth disease virus, in particular to an enzyme-linked immunosorbent detection kit for an antibody to the structural protein VP1 of foot-and-mouth disease virus. Background technique [0002] Foot-and-Mouth Disease (FMD) is a global epidemic infectious disease that is widely distributed in the world and spread internationally. The International Bureau of Veterinary Medicine (OIE) listed foot-and-mouth disease as the first of Class A infectious diseases in the revised classification of animal infectious diseases in 1984. Foot-and-mouth disease is a severe infectious disease of artiodactyla animals caused by foot-and-mouth disease virus infection. It is often popular in cattle and pigs on a large scale. The fatality rate of foot and mouth disease is very low, ...

Claims

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Application Information

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IPC IPC(8): C07K14/09G01N33/68G01N33/569
CPCC07K14/005C12N2770/32122G01N33/56983G01N2333/09
Inventor 肖进齐鹏董春娜王楠赵洪涛张蕾张欣张艳宾张君李凤鸣郑应华
Owner CHINA ANIMAL HUSBANDRY IND
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