Polypeptide and ELISA detection reagent kit for detecting Newcastle disease virus antibody

A Newcastle disease virus and detection kit technology, applied in the field of agricultural biology, can solve the problems of poor specificity and sensitivity that cannot meet clinical requirements, and achieve the effects of easy standardization, good sensitivity and specificity

Active Publication Date: 2019-09-17
广纳达康(广州)生物科技有限公司
View PDF4 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, for a long time, the sensitivity of the ELISA detection kits for detecting Newcastle disease virus serum antibody levels developed in the early stage ca

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Polypeptide and ELISA detection reagent kit for detecting Newcastle disease virus antibody
  • Polypeptide and ELISA detection reagent kit for detecting Newcastle disease virus antibody
  • Polypeptide and ELISA detection reagent kit for detecting Newcastle disease virus antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Embodiment 1 detects the polypeptide analysis of chicken Newcastle disease virus antibody

[0050] According to the amino acid sequence of the chicken Newcastle disease virus HN protein, the relative conserved sites were analyzed by blast software, and the B of the protein was comprehensively analyzed in combination with the hydrophilicity analysis (hydrophilicity), flexibility (flexibility), and non-shielding (accessibility) of the protein sequence. For cell epitopes (http: / / crdd.osdd.net / raghava / bcepred / ), four sequences were selected as candidate polypeptide sequences, namely: NDV-H1, NDV-H2, NDV-H3 and NDV-H4.

[0051] According to the amino acid sequence of the F protein of chicken Newcastle disease virus, its relative conserved sites were analyzed by blast software, and the B of the protein was comprehensively analyzed in combination with the hydrophilicity analysis (hydrophilicity), flexibility (flexibility), and non-shielding (accessibility) of the protein sequen...

Embodiment 2

[0059] Embodiment 2 detects the screening of the polypeptide of chicken Newcastle disease virus antibody

[0060] The above-mentioned artificially synthesized polypeptide was coupled with BSA carrier protein by glutaraldehyde method to improve the coating efficiency of the polypeptide. The specific method is: 2 mg of carrier BSA is dissolved in 2 mL of PBS buffer (0.01 mol / L, pH 7.4), dialyzed in PBS buffer at room temperature overnight; 1 mg / mL of polypeptide is added to 2 mL of carrier BSA solution, and the After stirring, 1 mL of 0.2% glutaraldehyde solution was added dropwise to the above mixture, and stirred while adding. Stir at room temperature for 4 hours, add 400 μL of 1 mol / L glycerol to the above mixture, and continue stirring for 1 hour to terminate the reaction; put the above mixture into a dialysis bag, place it in PBS buffer and dialyze overnight at 4°C; collect the peptide carrier pair Combined, store at -80°C.

[0061] The coupled polypeptides obtained above...

Embodiment 3

[0065] Example 3 Determination of the Optimum Coating Concentration of Antigen and Optimum Serum Dilution of Newcastle Disease Virus ELISA Antibody Detection Kit

[0066] Determine the optimal coating concentration of the antigen and the optimal dilution of the serum according to the matrix titration method, and use the coating solution to mix the NDV-H4 polypeptide and the BSA conjugate at 1:100, 1:200, 1:400, 1: After dilution of 800 and 1:1600, add them to the microtiter plate respectively, add 1 column for each dilution, 100 μL per well, and place it at 2-8°C for 15 hours. Discard the coating solution, add 200 μL of blocking solution (5% skim milk) to each well, incubate at 37° C. for 1 hour, discard the blocking solution in the well. Use the sample diluent to dilute the positive serum of Newcastle disease virus 1:320, 1:640, 1:1280, 1:2560, 1:5120, dilute the negative serum 1:40, 1:80, and dilute the positive serum and negative serum Each dilution of each dilution was ad...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses polypeptide and ELISA detection reagent kit for detecting a Newcastle disease virus antibody. The reagent kit is used for coating an elisa plate through screened Newcastle disease HN protein polypeptide. Through detection of combining capacity of a serum antibody and polypeptide in samples, and through assistance with the antibody of goat anti-chicken Ig gamma labelled with horseradish peroxidase, the level of Newcastle disease virus antibody is detected. The reagent kit is used for detecting avian influenza virus H5, H7, H9 subtype antibodies, chicken infectious Fabricius' bursal disease virus antibodies, and chicken infectious bronchitis virus antibodies to be negative. The reagent kit can detect that the hemagglutination inhibition (HI) titer is greater than or equal to 4log<2> serum, and can meet clinical serum sensitivity detection requirements. When the reagent kit and a hemagglutination inhibition method are both used for detecting clinical serum, the coincidence rate is as high as 98.7%. The reagent kit is simple to operate, free from special material requirements, good in result judgment objectivity, free from influence by subjective judgment by operators, and suitable for being used as a tool for detecting and monitoring the level of the antibodies after immunization with a Newcastle disease vaccine.

Description

technical field [0001] The invention relates to the field of agricultural biotechnology, in particular to a polypeptide for detecting Newcastle disease virus antibody and an ELISA detection kit thereof. Background technique [0002] Newcastle disease (ND) is an acute and highly contagious infectious disease of chickens and various poultry caused by Newcastle disease virus. The disease spreads rapidly and has a high fatality rate. The main features of the disease are dyspnea, diarrhea, nervous disturbance, mucosal and serosal hemorrhage. The disease can cause significant economic losses, and is stipulated by OIE as a disease that must be reported. Therefore, countries all over the world attach great importance to the occurrence and prevalence of this disease. [0003] Newcastle disease virus (NDV) belongs to the Paramyxoviridae family and the genus Avian Mumps virus. The complete virus particles are nearly round, with a diameter of 100-500 nm. The virus has an envelope stru...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C07K14/125G01N33/569G01N33/543
CPCC07K14/005G01N33/56983G01N33/54306C12N2760/18122
Inventor 张安定成泽任海洋
Owner 广纳达康(广州)生物科技有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap