Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

B cell epitope polypeptide of cysteine protease inhibitor of trichina at intestinal tract period, hybridoma cell strain, monoclonal antibody and application

A hybridoma cell line and monoclonal antibody technology, applied in the field of immunology, can solve the problems of complex ES antigen components, hindering practical application, and long production cycle

Active Publication Date: 2020-06-19
JILIN UNIV
View PDF5 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the ES antigen has complex components, cumbersome preparation, long production cycle, uneven batch quality, and serious diagnostic blind spots (cannot be detected before 19 days after infection) and cross-reactivity, which hinder its practical application.

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
  • B cell epitope polypeptide of cysteine protease inhibitor of trichina at intestinal tract period, hybridoma cell strain, monoclonal antibody and application
  • B cell epitope polypeptide of cysteine protease inhibitor of trichina at intestinal tract period, hybridoma cell strain, monoclonal antibody and application
  • B cell epitope polypeptide of cysteine protease inhibitor of trichina at intestinal tract period, hybridoma cell strain, monoclonal antibody and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] Prokaryotic expression and purification of embodiment 1Ts-WN10 protein

[0064] 1. Primer Design

[0065] According to the registered TsWN10 gene sequence in Genbank (accession number: EU263325), design PCR amplification primers, the sequence is as follows:

[0066] TsWN10-EcoRI-atg:5'-TAAC GAATTC ATGCAGATACTTGGTGA-3' (as shown in SEQ ID No.3)

[0067] TsWN10-XhoI-tta:5'-GACG CTCGAG TTAACATTCAACA-3' (as shown in SEQ ID No.4)

[0068] The underlined part is the introduced EcoRI and XhoI restriction sites, and the expected length of the amplified product is 1187bp.

[0069] 2. Extraction and reverse transcription of Trichinella spiralis T1 (T.spiralis) RNA

[0070] Take one Trichinella spiralis T1 (T.spiralis) conservation mouse in this experiment, kill it by neck dislocation, peel off the skin, tail, viscera and claws, wash the carcass and mince it, put it in 300ml containing 1% HCl and 1% stomach The protease digestion solution was stirred and digested in a 37°C...

Embodiment 2

[0091] Example 2 Preparation of anti-Ts-WN10 protein monoclonal antibody

[0092] 1. Mice Immunization

[0093] Five 6-week-old female BALB / c mice were immunized with the purified recombinant Ts-WN10 protein for 5 times with a two-week interval between each immunization.

[0094] One week after the 4th immunization and 5th immunization, blood was collected from the tail of the mice, and the serum was separated (centrifuged at 3000rpm at 4°C for 30min), and the antibody level was detected by Ts-WN10 indirect ELISA method. The Ts-WN10 indirect ELISA method is operated as follows: Ts-WN10 protein refolded and purified on the column is diluted with coating solution (0.01M NaOH PH12), the coating amount is 0.125ug / well, 100ul per well. Coat at 37°C for 2h or overnight at 4°C. After that, the plate was washed 3 times with PBST (containing 0.05% Tween 20). Block with blocking solution (5% skim milk) at 37°C for 2h. After that, the plate was washed 3 times with PBST. Serum to be ...

Embodiment 3

[0102] Identification of embodiment 3 monoclonal antibody

[0103] 1. Subclass identification of monoclonal antibodies

[0104] Subclass identification of the monoclonal antibody obtained in Example 2 was carried out according to the operating instructions of the antibody subclass identification kit. The results show that the heavy chain of the monoclonal antibody of the present invention is IgG1 and the light chain is κ chain, see Table 1 for details.

[0105] Table 1 Identification of monoclonal antibody subclasses

[0106] Monoclonal antibody name lgG1 lgG2a wxya lgG3 lm w wxya kappa chain lambda chain WN10-1H9 0.837 0.062 0.045 0.048 0.031 0.046 0.7 5 7 0.039

[0107] 2. Ts-WN10 competition ELISA test

[0108] Coating and sealing are the same as Ts-WN10 indirect ELISA, starting from 1:1, diluting the positive and negative sera of Trichinella spiralis infection by 2 times from 1:1, taking 50ul of the diluted serum to be tested a...

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

PropertyMeasurementUnit
Concentrationaaaaaaaaaa
Login to View More

Abstract

The invention relates to the field of immunology, in particular to B cell epitope polypeptide of a cysteine protease inhibitor (Ts-WN10) of trichina at an intestinal tract period, a hybridoma cell strain, a monoclonal antibody and an application of the B cell epitope polypeptide, the hybridoma cell strain and the monoclonal antibody. The hybridoma cell strain which can secrete Ts-WN10 protein resistant specific antibodies can be obtained, Ts-WN10 protein specific B cell epitope polypeptide recognized by the monoclonal antibody secreted by the hybridoma cell strain is identified, and the B cellepitope polypeptide, the hybridoma cell strain and the monoclonal antibody have important significance in diagnosis of trichina diseases, and have important significance in detection of trichina multiple hosts, by sandwich ELSIA which is used for establishing competitive ELISA and detecting a circulating antigen and development of a subunit vaccine.

Description

technical field [0001] The invention relates to the field of immunology, in particular to a B cell epitope polypeptide, a hybridoma cell line, a monoclonal antibody and an application thereof of a trichinella spiralis intestinal stage cysteine ​​protease inhibitor. Background technique [0002] Trichinellosis is mainly caused by the host eating raw or semi-raw meat containing trichinella, and pork is the main source of human infection. The average incubation period of human trichinellosis is 2 weeks. The main symptoms of patients in the acute stage are fever, severe muscle pain, severe diarrhea, facial edema, and eosinophilia, etc. The symptoms can last for several weeks and lead to body failure, especially severe infection Severe damage to the myocardium and brain may occur, and even death may occur. [0003] In view of the great threat and harm caused by trichinella spiralis to public health and safety, the World Organization for Animal Health (OIE) listed trichinellosis ...

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): C07K14/81C07K16/38C12N15/15C12N5/20G01N33/68G01N33/577G01N33/53A61K39/395A61P33/10C12R1/91
CPCC07K14/8139C07K16/38G01N33/6893G01N33/577G01N33/5308A61P33/10C07K2317/76C07K2317/33C07K2317/34G01N2333/8139A61P33/00C07K16/18C07K2317/92G01N2333/4353C07K16/20C07K2317/56C07K2317/565G01N33/56905
Inventor 刘明远刘晓雷刘琰杨勇唐斌
Owner JILIN UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap 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