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A kind of anti-channel catfish virus nucleocapsid protein monoclonal antibody and its application

A channel catfish and monoclonal antibody technology, applied in the field of immunology, can solve the problems of cumbersome operation, inability to meet the work requirements of rapid diagnosis of pathogens, long detection cycle, etc., and achieve simple and convenient preparation, strong immunogenicity and strong specificity Effect

Active Publication Date: 2020-03-10
CHINESE ACAD OF INSPECTION & QUARANTINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these methods are accurate and reliable, they cannot meet the actual work needs of rapid diagnosis of pathogens and timely control of the epidemic due to the disadvantages of cumbersome operation and long detection cycle. Therefore, it is important to establish more sensitive and rapid detection technology for the prevention and treatment of this disease significance

Method used

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  • A kind of anti-channel catfish virus nucleocapsid protein monoclonal antibody and its application
  • A kind of anti-channel catfish virus nucleocapsid protein monoclonal antibody and its application
  • A kind of anti-channel catfish virus nucleocapsid protein monoclonal antibody and its application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1 Preparation of anti-channel catfish virus monoclonal antibody

[0030] 1. Antigen preparation and purification

[0031] The channel catfish virus suspension was purified by differential centrifugation, that is, the channel catfish virus suspension was first passed through 8000r / min, and centrifuged for 30min; mol / LPBS resuspended. After the concentration was measured by a spectrophotometer, it was used to immunize mice.

[0032] 2. Immunization of Mice

[0033]The mice used for immunization were 5-week-old female BALB / c mice of SPF grade. Each mouse was given the above-mentioned purified channel catfish virus suspension as an antigen for basic immunization by intraperitoneal injection, and the virus suspension was mixed with Freund's complete adjuvant 1:1, and injected intraperitoneally; 2 weeks later, booster immunization, channel catfish virus (30 micrograms) mixed with Freund's incomplete adjuvant 1:1, injected intraperitoneally; after that, boosted ...

Embodiment 2

[0045] Example 2 Characterization of anti-channel catfish virus nucleocapsid protein monoclonal antibody identification

[0046] 1. Isotype identification of monoclonal antibodies

[0047] Method: The typing kit (SBA Clonotyping TM System / HRP), according to its instructions, the Ig subtype of the monoclonal antibody secreted by the mouse hybridoma cell line CCV-8B6 of the present invention is identified.

[0048] Results: The subtype of the monoclonal antibody secreted by the mouse hybridoma cell line CCV-8B6 of the present invention is IgG3 k chain.

[0049] 2. Antigenic site analysis of monoclonal antibodies

[0050] Western blotting:

[0051] Purified channel catfish virus monoclonal antibody was used as the detection group, normal channel catfish ovary cell lysate was used as negative control, serum of immune mice was used as positive control, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis ( SDS-PAGE) to separate proteins, transfer the proteins on the ge...

Embodiment 3

[0060] Embodiment 3 detects the composition of the colloidal gold test strip of channel catfish virus

[0061] Detect the colloidal gold test strip of channel catfish virus, this colloidal gold test paper comprises the water-absorbing pad that connects successively, basement membrane (nitrocellulose membrane), gold mark pad and sample pad; Described basement membrane is provided with and detects T line and Quality control C line, described quality control C line is coated with goat anti-mouse IgG antibody (commercially purchased gain), and described detection T line is coated with primary antibody (i.e. rabbit anti-channel catfish virus polyclonal antibody); the gold label pad contains a colloidal gold-labeled secondary antibody (the monoclonal antibody against channel catfish virus nucleocapsid protein secreted by the hybridoma cell line CCV-8B6 with the preservation number CGMCC No.13843).

[0062] Among them, preparation of rabbit polyclonal antibody against channel catfish...

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Abstract

The invention discloses a monoclonal antibody for resisting channel catfish virus nucleocapsid protein. The monoclonal antibody is secreted by a mouse hybridoma cell strain of which the preservation number is CGMCC No.13843. The invention also discloses a colloidal gold test strip for detecting the channel catfish virus. The invention further discloses application of the monoclonal antibody, the hybridoma cell strain and the colloidal gold test strip. The monoclonal antibody for resisting the channel catfish virus nucleocapsid protein is high in specificity, the channel catfish virus can be accurately detected, and favorable conditions for preventing and the diagnosing channel catfish virus are provided.

Description

technical field [0001] The invention relates to the field of immunology, and more specifically relates to a monoclonal antibody against nucleoprotein of channel catfish virus, a hybridoma cell line and a colloidal gold test strip for detecting channel catfish virus. Background technique [0002] Channel catfish (Ictalurus punctatus), also known as channel catfish, belongs to the Siluriformes family. It has the characteristics of relatively fast growth, tender meat and high sales price, and is welcomed by consumers and aquaculture industry. With the gradual expansion of its breeding scale, the increasing prevalence of channel catfish virus disease (CCVD) caused by virus infection has caused huge economic losses to its farming industry. [0003] Channel catfish virus disease (CCVD) was first prevalent in North America. The optimum water temperature for an outbreak is 25-30°C. The disease is a disease caused by a herpes virus (Channel catfish virus (CCV) for short), which ca...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/08C12N5/20G01N33/532G01N33/543G01N33/569G01N33/577
CPCC07K16/085G01N33/532G01N33/543G01N33/56994G01N33/577C07K2317/33
Inventor 景宏丽吴绍强林祥梅高隆英张旻王娜
Owner CHINESE ACAD OF INSPECTION & QUARANTINE