Eureka AIR delivers breakthrough ideas for toughest innovation challenges, trusted by R&D personnel around the world.

H3 type flu virus hemagglutinin space conformation simulation antigen epitope and application thereof

An antigenic epitope, influenza virus technology, applied in the field of biomedicine, can solve the problems of time-consuming and laborious, unable to obtain spatial conformation epitopes, etc.

Inactive Publication Date: 2010-06-02
CHINA PHARM UNIV
View PDF1 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Conventional epitope mapping methods are time-consuming and laborious and cannot obtain spatial conformational epitopes

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
  • H3 type flu virus hemagglutinin space conformation simulation antigen epitope and application thereof
  • H3 type flu virus hemagglutinin space conformation simulation antigen epitope and application thereof
  • H3 type flu virus hemagglutinin space conformation simulation antigen epitope and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1, Prokaryotic expression of influenza virus hemagglutinin fragment HA1

[0045] 1) Cloning of influenza virus hemagglutinin fragment HA1 gene ha1: On MDCK cells growing into a single layer, inoculate H3 influenza virus A / Swine / Guangdong / 6 / 2004 (H3N2) strain, 37 ° C 5% CO 2 Culture in an incubator for 48 hours. When obvious cell lesions appear, the cells are collected, the cells are lysed by repeated freezing and thawing, centrifuged to remove the precipitate, and the supernatant is taken. Total RNA was extracted with Trizol reagent (product of Invitrogen Company), and a pair of primers were designed according to the hemagglutinin gene sequence of influenza virus A / Swine / Guangdong / 6 / 2004 (H3N2) strain (Gene ID: AY852277), followed by reverse transcription RT-PCR was carried out under the action of enzyme (product of Invitrogen Company) and Taq enzyme (product of Takara Company) to obtain the ha1 gene, which was identified by DNA sequencing. The two primer seque...

Embodiment 2

[0050] Example 2, Purification and Identification of Anti-recombinant HA1 Rabbit Serum IgG

[0051] 1) Purified recombinant HA1 was used to immunize New Zealand rabbits: the purified recombinant HA1 was adjusted to 200 μg / mL with 0.01 M, pH 7.4 PBS, mixed with an equal volume of Freund's adjuvant, emulsified, and subcutaneously multi-pointed on the back to immunize New Zealand rabbits For rabbits, the second immunization is performed two weeks after the first immunization, and the third immunization is performed two weeks after the second immunization. The second and third doses were emulsified with an equal volume of Freund's incomplete adjuvant, and were injected subcutaneously at multiple points on the back. After three immunizations, the titer of the antibody was tested. When the titer reached 1:32, blood was collected from the heart, and the serum was separated.

[0052] 2) Purification of anti-recombinant HA1 rabbit serum IgG: use saturated ammonium sulfate fractional p...

Embodiment 3

[0053] Embodiment 3, phage random peptide library screening

[0054] 1) The phage random peptide library is the M13 phage twelve peptide library (Ph.D.-12TMPhage Display Peptide Library Kit) commercialized by BioLabs

[0055] 2) Screening method: Coat the ELISA plate with anti-recombinant HA1 rabbit serum IgG (100 μg / mL) diluted in the coating solution (carbonate buffer solution with pH 9.6), overnight at 4°C, add blocking solution (3% BSA) at 4°C for 2-3 hours, wash with TBST, and spin dry the washing solution. Add diluted phage random peptide library to each well, and the number of phage added in each round should be 2×10 10-11 CFU, incubate at room temperature for 1 hour, shake off the liquid, rinse with TBST buffer containing 0.1% tween-20 (v / v), and finally use 100 μl pH 2.2 glycine buffer to elute the IgG that specifically binds to the anti-recombinant HA1 rabbit serum phage, and immediately neutralized with 10 μl pH 9.8 Tris buffer, 10 μl was taken for phage titer det...

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 relates to polypeptide biological technical field, which discloses a spatial conformation mimic antigen epitope of H3-type influenza virus hemagglutinin and a relative application. The sequence of the spatial conformation mimic antigen epitope is SEQ ID No. 1. The invention utilizes anti-recombination HA1 rabbit serum IgG selection phage 12 peptide library, uses phage ELISA, DNA sequencing, western bolt, and software analysis to analyze selected clone, and selects the small peptide from random phage 12 peptide library, which can be specially combined with anti-recombination HA1 rabbit serum IgG. The inventive spatial conformation mimic antigen epitope can prepare diagnostic reagent box of H3-type influenza virus.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and relates to an H3 type influenza virus hemagglutinin space conformation mimic antigen epitope and application thereof. Background technique [0002] Influenza virus belongs to the Orthomyxoviridae family, which is a single-strand negative-sense, segmented RNA virus. The genome consists of 8 segments and encodes 10 viral proteins. According to the different antigenic types of the internal proteins of influenza virus, it is divided into A (A), There are three types of B (B) and C (C), and the type A influenza virus is divided into different serotypes according to the antigenicity of the influenza virus surface protein hemagglutinin (HA) and neuraminidase. Now 15 different HA subtypes (H1-H15). H3 influenza virus can infect humans, poultry, pigs, horses and other mammals. In 1968, H3N2 influenza virus caused a sudden influenza pandemic and seriously endangered public safety. [0003] Hemagg...

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
Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/08C07K14/705G01N33/569
Inventor 钟辉邓小昭姚文娟董莉莉于娟储春丽刁振宇张云
Owner CHINA PHARM UNIV
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
Eureka Blog
Learn More
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com