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Fowl leucovirus subgroup J specific antigenic polypeptide and application thereof

An avian leukemia virus and antigen polypeptide technology, which is applied in the biological field, can solve the problem of low specificity of the ALV-J antibody kit, and achieve the effects of being favorable for popularization and use, improving specificity and high specificity

Active Publication Date: 2017-02-22
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The object of the present invention is to provide a specific antigenic polypeptide of avian leukosis virus J subgroup (ALV-J) with strong specificity, better hydrophilicity, good reactivity and high purity, and the antigenic polypeptide has ALV-J specific antigen The epitope has a good biological reaction with the corresponding anti-ALV-J serum, and can be used as an antigen to stimulate the body to produce specific antibodies; and the use of this antigen polypeptide to construct has stronger specificity and stability, and is suitable for ALV-J serum Antibody detection ELISA detection technology to solve the technical problem of low specificity of the current ALV-J antibody kit

Method used

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  • Fowl leucovirus subgroup J specific antigenic polypeptide and application thereof
  • Fowl leucovirus subgroup J specific antigenic polypeptide and application thereof
  • Fowl leucovirus subgroup J specific antigenic polypeptide and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] The present embodiment provides a kind of indirect ELISA detection kit for specific detection ALV-J antibody, and its composition is as follows:

[0025] [1] A microtiter plate coated with an antigen polypeptide, the amino acid sequence of the antigen polypeptide is SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5 or Its derivative sequence is encoded by the gp85 partial gene of the envelope protein gene (env) of the virus, and has strong reactogenicity;

[0026] [2] enzyme-labeled antibody: horseradish peroxidase-labeled anti-chicken IgG;

[0027] [3] substrate solution preparation: 100mmol / L citric acid solution ((21g citric acid (C 6 h 8 O; H 2 0) Dissolve in deionized water, dilute to 1L) 24.3mL, 200mmol / LNa 2 HP0 4 .12H 2 0 (71.6g Na 2 HP0 4 .12H 2 0 dissolved in deionized water, dilute to 1L) 25.7mL and mix well, add 50mg of tetramethylbenzidine (TMB), add 50μL of 30% H 2 0 2 ;

[0028] [4] Stop solution (2mol / L) H 2 S0 4 : Get 177.8mL...

Embodiment 2

[0039] Use biological software to analyze the amino acid sequence of GP85 of ALV-J, the amino acid sequence SEQ ID NO.1 to SEQ ID NO.5 with better antigenicity, carry out conventional antigen polypeptide synthesis, choose one of the antigen polypeptides to coat the microtiter plate, An indirect ELISA detection kit (see Example 1 for composition and method) was prepared for specific detection of ALV-J antibody, and the serum of chickens of different ages artificially infected with ALV-J was detected. The detection results are shown in Table 1.

[0040] Table 1: ELISA detection effect based on the antigenic peptide sequence SEQ ID NO.1:

[0041]

[0042] It can be clearly seen from the results in the above table that the kit constructed by the polypeptide can detect specific antibodies in the sera of different day-old chickens infected with ALV-J.

Embodiment 3

[0044] In this example, the difference from Example 2 is that one of the five polypeptide sequences is optional, such as SEQ ID NO.3, only the first half of the sequence length is synthesized, and the derivation of the peptide is shortened by half. The synthetic shortened antigen polypeptide was used to coat the ELISA plate, and a kit was constructed to detect the serum of chickens artificially infected with ALV-J. The detection results are shown in Table 2. It can be seen that the ALV-J antibody-positive chickens can still be detected by using sequence-derived shortened antigen peptides to construct the kit, and the OD values ​​are all greater than 0.2.

[0045] Table 2: ELISA detection effect of the kit based on the sequence-derived shortened antigen peptide of SEQ ID NO.3:

[0046]

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Abstract

The invention provides a fowl leucovirus subgroup J specific antigenic polypeptide and an application thereof. The sequences of antigenic polypeptide are SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5 or deriving sequences of the sequences above. Besides, the antigenic polypeptide is coded by genes in gp 85 section of envelope protein gene (env) of a virus, and has strong specificity, good hydrophily, good reactivity and high purity. In addition, the antigenic polypeptide is used for preparing an indirect ELISA detection kit which can conduct specific detection on an ALV-J antibody, wherein the detection kit comprises one or more of enzyme labeled plates of ALV-J specific antigenic polypeptide above and related reagents, and the enzyme labeled plates and the reagents coat the detection kit. During the detection of ALV-J antibody, the kit has high specificity and low cost, is beneficial to popularization, and the effects are obviously superior to those of conventional kits and imported kits. The ALV-J specific antigenic polypeptide and the ELISA detection kit prepared by the antigenic polypeptide are applicable to evaluation of results of infection or purification of the ALV-J antibody; the ALV-J specific antigenic polypeptide is applicable to preparation of antibody of specific ALV-J.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an avian leukosis virus subgroup J (ALV-J) specific antigen polypeptide, an ELISA detection kit prepared by using it, and applications thereof. Background technique [0002] Avian leukosis subgroup J virus (ALV-J) is a C-type retrovirus, which is a recombinant virus integrated with endogenous virus and exogenous virus. It is mainly identified based on its unique envelope glycoprotein (gp85). classified as a separate subgroup. It not only causes myeloid cell tumors and other malignant tumors in broiler chickens, but also causes neoplastic diseases in laying hens, causing significant economic losses to the poultry industry. At present, there is no effective treatment and vaccine for subgroup J avian leukemia, and the disease can only be controlled by specific detection and elimination of positive chickens and population purification. The methods currently used to detect the avian leu...

Claims

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Application Information

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IPC IPC(8): G01N33/569
CPCG01N33/56983
Inventor 秦爱建田雪邵红霞钱琨叶建强
Owner YANGZHOU UNIV
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