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Canine parvovirus hybridoma cells, monoclonal antibodies and applications

A hybridoma, monoclonal antibody technology, applied in the field of monoclonal antibodies, can solve problems such as low sensitivity and achieve good neutralizing activity

Active Publication Date: 2019-04-12
LUOYANG PULIKE WANTAI BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The invention also provides a canine parvovirus ELISA detection kit, which can be used for fast, convenient and accurate detection of canine parvovirus, and overcomes the problem of low sensitivity in the detection of the prior art. Gold rapid test strips have higher sensitivity

Method used

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  • Canine parvovirus hybridoma cells, monoclonal antibodies and applications
  • Canine parvovirus hybridoma cells, monoclonal antibodies and applications
  • Canine parvovirus hybridoma cells, monoclonal antibodies and applications

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] Example 1 Preparation, purification, identification and testing of anti-canine parvovirus monoclonal antibody

[0073] 1.1 Preparation and purification of monoclonal antibody against canine parvovirus

[0074] The canine parvovirus CVCC AV298 strain was prepared according to the operation method of Harlow E et al. (Harlow E, Lane D. Antibodies: laboratory manual [M]. New York: Cold Spring Harbor Laboratory Press, 1998, 139-312). strain, hybridoma 10H4 strain, further prepared and purified anti-canine parvovirus monoclonal antibody 10B11 and anti-canine parvovirus monoclonal antibody 10H4 respectively.

[0075] Among them, the preservation number of the hybridoma cell 10B11 strain is CCTCC No: C201578, which was preserved in the China Center for Type Culture Collection, and the preservation address was Wuhan, Wuhan University, China, and the preservation time was May 20, 2015. It secretes anti-canine parvovirus Monoclonal antibody 10B11; hybridoma cell 10H4 strain prese...

Embodiment 2

[0109] Example 2 Pairing of Monoclonal Antibodies

[0110] 2.1 Selection of monoclonal antibody pairing mode

[0111] When screening monoclonal antibody pairing combinations, the following factors are mainly considered: first, the activity of the monoclonal antibody; second, whether there is a non-specific reaction between the monoclonal antibody and other viruses except canine parvovirus; third, the background color development.

[0112] 2.2 Detection of Monoclonal Antibody Activity

[0113] The canine parvovirus CVCC AV298 strain was selected, diluted to 0.01HA, and tested for different matching modes. The results are shown in Table 4:

[0114] Table 4 Monoclonal Antibody Matching Detection

[0115]

[0116] Note: + means positive reaction, - means negative reaction.

[0117] The results showed that except for the solid-phase monoclonal antibody 10B11 and the enzyme-labeled monoclonal antibody 10H4, the detection of the 0.01HA canine parvovirus CVCC AV298 strain virus ...

Embodiment 3

[0119] Example 3 Preparation of Canine Parvovirus ELISA Kit

[0120] 3.1 Horseradish peroxidase (HRP) labeling of monoclonal antibodies

[0121] According to the monoclonal antibody 10H4 prepared in Example 1, according to Tijssen P et al. (Tijssen P, Kurstak E. Highly efficient and simple methods for the preparation of peroxidase and active peroxidase antibody conjugates for enzyme immunoassays [J]. 457) for horseradish peroxidase (HRP) labeling of anti-canine parvovirus monoclonal antibodies.

[0122] 3.2 Immobilization of monoclonal antibodies

[0123] The monoclonal antibody 2 prepared in Example 1 was immobilized on the nitrocellulose membrane with a BioDot XYZ3050 three-dimensional spraying platform.

[0124] 3.3 Composition of the detection kit

[0125] The detection kit includes detection test strips, sample processing solution, sample processing tube, and sample preservation solution.

[0126] (1) Detection test strips (the side schematic diagram is shown in figu...

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Abstract

The invention provides a hybridoma prepared from canine parvovirus antigens. An anti-canine-parvovirus monoclonal antibody secreted from the hybridoma has quite high relative affinity constant and good neutralizing activity, and a vaccine composition prepared from the monoclonal antibody can effectively prevent and treat canine parvovirus infections. A canine parvovirus detection kit prepared with two kinds of monoclonal antibodies has the advantages of quickness, convenience, accuracy, good sensitivity, specificity and repeatability and has higher sensitivity than colloidal gold-labeled commercial test strips.

Description

technical field [0001] The invention relates to a canine parvovirus hybridoma cell, a monoclonal antibody produced by it, a vaccine composition prepared by it, an immune detection system and its application, and belongs to the field of biotechnology. Background technique [0002] Canine parvovirus (Canine parvovirus, CPV) exists widely in nature, causing animal infection of canine parvovirus disease, which can occur all year round. The main clinical symptoms are vomiting, diarrhea and leukopenia, and the morbidity and mortality are relatively high. Dogs are the main natural host of CPV. Dogs of all ages and sexes are susceptible. CPV can also infect other canids such as minks, foxes, wolves, coyotes, crab-eating foxes, South American dogs, and Asian foxes and weasels. Animals, puppies are the most susceptible, with antigen drift, can also infect cats. CPV can be excreted with the feces and urine of sick animals, polluting the surrounding environment, and the feces of rehabi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/20C07K16/08G01N33/577G01N33/569C12R1/91
Inventor 田克恭郝丽影
Owner LUOYANG PULIKE WANTAI BIOTECH
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