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Monoclonal antibody for resisting porcine epidemic diarrhea viruses and application thereof

A porcine epidemic diarrhea, monoclonal antibody technology, applied in the direction of antiviral agents, antiviral immunoglobulins, antibodies, etc., can solve the problems of time-consuming and expensive

Active Publication Date: 2016-04-06
LUOYANG PULIKE WANTAI BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the methods for detecting porcine epidemic diarrhea virus include virus isolation and identification and serum examination, but it takes too long; although the PCR method is sensitive, it requires expensive instruments and professional operators

Method used

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  • Monoclonal antibody for resisting porcine epidemic diarrhea viruses and application thereof
  • Monoclonal antibody for resisting porcine epidemic diarrhea viruses and application thereof
  • Monoclonal antibody for resisting porcine epidemic diarrhea viruses and application thereof

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Experimental program
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Embodiment approach

[0028] As an embodiment of the present invention, the antibody is monoclonal antibody PEDV-McAB1, the amino acid sequence of the heavy chain variable region of the monoclonal antibody PEDV-McAB1 is SEQ ID No. 2, and the amino acid sequence of the light chain variable region is SEQ ID No. 4.

[0029] The relative affinity constant of the monoclonal antibody PEDV-McAB1 is 0.78 μg / ml, that is to say, the binding strength of the monoclonal antibody PEDV-McAB1 to the antigenic determinant of the antigen is very high, which is better than that of the anti-antibody in the prior art. The binding strength of porcine epidemic diarrhea virus monoclonal antibody; The neutralizing antibody titer of described monoclonal antibody is greater than 1:512, that is to say that described monoclonal antibody PEDV-McAB1 has good neutralizing activity, can suppress porcine epidemic CDV superinfects target cells.

[0030] The term "genetically engineered antibody" can be expressed by appropriate host...

Embodiment 1

[0084] Embodiment 1 Preparation, purification, identification and inspection of anti-porcine epidemic diarrhea virus monoclonal antibody

[0085] 1.1 Preparation and purification of monoclonal antibody against porcine epidemic diarrhea virus

[0086] The porcine influenza virus HN1301 strain (see Chinese patent CN104513827A) was purified according to the method in Zhang Liyan et al. Hybridoma cells, two monoclonal antibodies were produced in mice.

[0087] The two monoclonal antibodies were prepared according to Chen Dan et al. ) operating method for purifying monoclonal antibodies by octanoic acid-ammonium sulfate combined precipitation method to purify anti-porcine epidemic diarrhea virus monoclonal antibody PEDV-McAB1 and anti-porcine epidemic diarrhea virus monoclonal antibody PEDV-McAB2 respectively.

[0088] 1.2 Identification of monoclonal antibodies against porcine epidemic diarrhea virus

[0089] 1.2.1 Identification of monoclonal antibody types and subclasses

[...

Embodiment 2

[0120] The determination of embodiment 2 monoclonal antibody variable region sequence

[0121] 2.1 Design of primers for heavy chain and light chain variable region of monoclonal antibody

[0122] According to the sequence characteristics of the mouse monoclonal antibody, the heavy chain variable region primer sequence was designed:

[0123] P1: 5'-ACTAGTTGACGTGGTCTCTAGGGTCACTTTAGTTTTTCCT-3'

[0124] P2: 5'-CGGAAGCTTCCAGCGRCCARKCCATATACIGRTGG-3'

[0125] Design light chain variable region primer sequences:

[0126] P3: 5'-GCCATCTCATGRAGWCATTKWCYCAAGTCTTT-3'

[0127] P4: 5'-CGGACGCTTACTGCCTGGTAAGAAGATGGA-3'

[0128] 2.2 Determination of the sequence of the variable region of the monoclonal antibody

[0129] According to Zhang Aihua et al. (Zhang Aihua, Guan Lan, Wang Zhiyou et al.. Cloning and sequence analysis of light and heavy chain variable region genes of mouse anti-CD molecular monoclonal antibodies. Chinese Journal of Biological Products, 2001, 15(2): 65-68 ) estab...

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Abstract

The invention provides a variable region sequence for a rat monoclonal antibody specially combined with porcine epidemic diarrhea viruses, a prepared antibody or an antibody fragment thereof, and vaccine and a kit prepared from the antibody. The medicine composition can effectively prevent and treat porcine epidemic diarrhea virus infection. The kit can effectively detect the porcine epidemic diarrhea viruses, is high in sensitivity, and carry out detecting fast.

Description

technical field [0001] The invention relates to an anti-porcine epidemic diarrhea virus monoclonal antibody, and applications such as a pharmaceutical composition and a kit prepared by using the monoclonal antibody, and belongs to the field of biomedicine. Background technique [0002] Porcine epidemic diarrhea (porcineepidemicdiarrhea, PED) is an intestinal infectious disease caused by porcine epidemic diarrhea virus (porcineepidemicdiarrheavirus, PEDV), which mainly causes vomiting, watery diarrhea, dehydration and other symptoms in suckling piglets. Death, the mortality rate can reach 50%-100%. In recent years, since the winter of 2010, the epidemic has broken out in most parts of my country, and then it has also broken out on a large scale in South Korea, Japan, Taiwan and other regions. In the first half of 2013, the disease also broke out in North America and spread rapidly, causing serious losses to the global breeding industry. [0003] At present, the methods for ...

Claims

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Application Information

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IPC IPC(8): C07K16/10A61K39/395A61P31/14G01N33/577G01N33/569
CPCA61K2039/505C07K16/10
Inventor 田克恭张云静
Owner LUOYANG PULIKE WANTAI BIOTECH
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