ELISA (enzyme linked immunosorbent assay) kit for detecting PPRV (peste des petits ruminants virus) antibody

An enzyme-linked immunosorbent reagent, technology of Peste des petits ruminants, applied in the direction of virus/bacteriophage, antisense single-stranded RNA virus, virus, etc., can solve problems that have not been seen yet, and achieve good sensitivity, high specificity, and easy operation Effect

Active Publication Date: 2017-10-10
CHINA ANIMAL DISEASE CONTROL CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The most common reports at home and abroad use purified PPRV N and H proteins (full length) or partially recombinant N and H protein fragments containing the main antigenic epitopes of N protein as the coating antigen, and also use concentrated PPRV whole virus as the coating antigen. An indirect ELISA method was established to detect PPR serum antibodies by the antigen (Qiu Wenying, 2011; Balamurugan et al., 2007), but there is no report on the establishment of an indirect ELISA method using the H-F tandem fusion protein as the coated antigen

Method used

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  • ELISA (enzyme linked immunosorbent assay) kit for detecting PPRV (peste des petits ruminants virus) antibody
  • ELISA (enzyme linked immunosorbent assay) kit for detecting PPRV (peste des petits ruminants virus) antibody
  • ELISA (enzyme linked immunosorbent assay) kit for detecting PPRV (peste des petits ruminants virus) antibody

Examples

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Embodiment 1

[0033] Embodiment 1, soluble expression rmHF1

[0034] 1. Synthetic genes

[0035]This application has designed three kinds of PPRV H and F protein fusion genes, which are respectively the rmHF1-Y gene shown in the 4th-3558th position of SEQ ID No.1 and the rmHF1-Y gene shown in the 1st-3684th position of SEQ ID No.3 rmHF2-Y gene and rmHF1-W gene shown in positions 4-3558 of SEQ ID No.5. The difference in the nucleotide sequence between the rmHF1-Y gene and the rmHF2-Y gene is that the 5' end is different, and the 24th-3564th nucleotides of SEQ ID No.1 and the 150th-3690th nucleotides of SEQ ID No.3 are the same .

[0036] Both the rmHF1-Y gene and the rmHF1-W gene encode the protein rmHF1 shown in SEQ ID No.2, and the rmHF2-Y gene encodes the protein rmHF2 shown in SEQ ID No.3. The difference in amino acid sequence between rmHF1 and rmHF2 lies only in the amino terminal, and the amino acid residues 8-1184 of SEQ ID No. 2 and 51-1227 of SEQ ID No. 3 are the same.

[0037] ...

Embodiment 2

[0052] Example 2. Using rmHF1 protein as the coating antigen indirect ELISA method to detect antibodies against Peste des petits ruminants virus

[0053] The relevant solutions in this embodiment are as follows:

[0054] The preparation of PBS buffer solution with a concentration of 0.01M and a pH value of 7.4: 8.5g NaCl, 0.2g KCl, 2.9gNa 2 HPO 4 12H 2 O, 0.59g NaH 2 PO 4 2H 2 O, 1L deionized water.

[0055] Coating buffer: 0.05mol / L sodium carbonate-sodium bicarbonate buffer (pH9.6), solvent is water, solute and its concentration are as follows: Na 2 CO 3 1.59g / L and NaHCO 3 2.93g / L.

[0056] Washing solution: 0.5% Tween washing solution. The 0.5% Tween washing solution was prepared as follows: Tween 20 and sodium azide were added to the PBS buffer solution with a concentration of 0.01M and a pH value of 7.4 until the content of sodium azide was 5 g / L, Tween 20 The content was 5mL / L, and 0.5% Tween washing solution was obtained.

[0057] Blocking solution: 1% BS...

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Abstract

The invention discloses an ELISA (enzyme linked immunosorbent assay) kit for detecting a PPRV (peste des petits ruminants virus) antibody. The ELISA kit for detecting the PPRV antibody comprises an envelope antigen, wherein the envelope antigen is a recombinant PPRV H-F fusion protein which is a protein of a) or b) as follows: a) protein formed by an amino acid sequence shown in SEQ ID No.2; b) soluble protein obtained from the amino acid sequence shown in SEQ ID No.2 through substitution and/or deletion and/or addition of one or several amino acid residues. The ELISA kit for detecting the PPRV antibody not only can be used for diagnosing PPR (peste des petits ruminants), but also can be used for evaluating a vaccine immunity effect, further can detect PPR rapidly and accurately and is beneficial to clinical monitoring of PPR, thereby playing a positive role in better control of spread of PPR in China.

Description

technical field [0001] The invention relates to an enzyme-linked immunoassay kit for detecting antibodies to Peste des petits ruminants virus in the field of enzyme-linked immunoassays. Background technique [0002] Peste des petits ruminants (PPR) is an infectious disease notifiable by OIE, and it is also listed as a first-class animal infectious disease by the Chinese government. It has the characteristics of high morbidity and high mortality. Production brings serious economic losses, and also has a serious impact on international trade, so the disease has attracted great attention from many countries. Peste des Petits Ruminants (PPR) is an acute, febrile, contagious disease caused by Peste des Petits Ruminants Virus (PPRV), which is characterized by high morbidity and mortality. The control and elimination of epidemic diseases is inseparable from effective vaccines and fast and sensitive detection methods as technical support. Therefore, increasing the research and deve...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569C07K19/00C12N15/62C12N15/70
CPCC07K14/005C07K2319/00C12N15/70C12N2760/18422G01N33/56983G01N2333/12
Inventor 孙雨宋晓晖杨林王传彬董浩杨天意
Owner CHINA ANIMAL DISEASE CONTROL CENT
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