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Method for preparing chicken pox virus antibody

A fowl pox virus and antibody technology, applied in the direction of antiviral immunoglobulin, serum immunoglobulin, instruments, etc., can solve the problems of low sensitivity, poor detection effect, inability to meet fowl pox diagnosis, vaccine testing and other problems, and achieve antibody High purity and specificity

Inactive Publication Date: 2012-10-31
北京健翔和牧生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although preventive vaccines have been on the market, effectively reducing the incidence and prevalence of the disease, the control and detection of the virus still require specific, high-sensitivity, and rapid detection methods because of the diversity and complexity of fowlpox virus antigens , the detection and diagnosis techniques based on antiserum cannot meet the needs of fowl pox diagnosis and vaccine inspection
[0003] There are many methods for chickenpox antigen detection and disease diagnosis. The traditional serological method has been used until now, that is, the use of immune serum for immunological tests, but this method has disadvantages such as poor detection effect and low sensitivity.

Method used

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  • Method for preparing chicken pox virus antibody

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Effect test

Embodiment 1

[0035] The preparation of embodiment 1 fowlpox virus antibody

[0036] 1. Virus culture and crude antigen preparation

[0037] Chicken embryonic fibroblasts grow into a monolayer and infect and inoculate 102 strains of fowlpox virus. After 4 to 5 days, digest the cells, centrifuge at 1500g for 10 minutes at 4°C, collect the precipitate, and wash with isotonic buffer (10mM Tris, pH8.0, 10mM NaCl, 5mM EDTA) after washing, dissolve the diluted virus with hypotonic buffer (10mM Tris, pH8.0, 10mM KCl, 5mM EDTA) containing 0.025% mercaptoethanol, 0.1% Triton-X100, centrifuge at 200g for 5min at 4°C, Remove cell debris and DNA.

[0038] 2. Concentration of virus

[0039] Use a 0.5-0.75 μm membrane cassette filter or a 0.5-0.7 μm hollow fiber column to remove cell debris. Rinse the filter with 2LPBS buffer (300kD cutoff), sterilize with 2L of 0.1M NaOH solution, and empty with normal saline. Prepare 5 L of permeation buffer (20 mM Tris, pH 7.8, 0.2 M NaCl, 1 mM MgCl 2 ) to concen...

Embodiment 2

[0061] Embodiment 2 neutralization test, fluorescent antibody staining

[0062] The purified antibody prepared in Example 1 was diluted in multiples and mixed with the same volume of antigen diluent (200TCID 50 / 0.1ml), overnight at room temperature, 0.1ml was added to the grown chicken embryo fibroblasts and cultured for 14 days to observe the results. Results found 101-10 4 Antibodies at double dilution neutralized the virus without any pathological changes in the cells, 10 5 The cell group of the diluted antibody has obvious cytopathic changes, so the neutralizing titer of the purified antibody is 10 4 .

[0063] After the virus strain infected QT-353~4 days, the cells were digested and resuspended in the culture medium to make the cell density reach 10 5 cells / ml, centrifuge at low speed, add 100 μl of cell solution on the slide, fix with 3% formalin (made in PBS) for 30 minutes, wash with PBS containing 0.1M glycine, add PBS containing 0.1% Trito-X100 and place for 15...

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Abstract

The invention discloses a preparation method of a chicken pox virus antibody, belonging to the technical field of biology. The method comprises the following steps: (1) culturing the chicken pox virus, and preparing a coarse antigen; (2) preparing the concentrated and purified antigen; (3) preparing the animal immune and antiserum; and (4) purifying the antibody. The antibody prepared by the method disclosed by the invention has the advantages of high purity and favorable specificity, and can be widely used for preparing reagents for detecting chicken pox virus infection.

Description

technical field [0001] The invention relates to a preparation method of virus antibody, in particular to a preparation method of fowlpox virus antibody, and belongs to the field of biotechnology. Background technique [0002] Fowlpox is an important disease in the poultry industry, which seriously endangers the development of the poultry industry, causing hypertrophic skin lesions (larynx, trachea), coughing and death in chickens. Although preventive vaccines have been on the market, effectively reducing the incidence and prevalence of the disease, the control and detection of the virus still require specific, high-sensitivity, and rapid detection methods because of the diversity and complexity of fowlpox virus antigens However, antiserum-based detection and diagnostic techniques cannot meet the needs of fowl pox diagnosis and vaccine testing. [0003] There are many methods for fowlpox antigen detection and disease diagnosis. The traditional serological method has been use...

Claims

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Application Information

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IPC IPC(8): C07K16/08C07K16/06G01N33/569
Inventor 张澍吕宏亮徐明伟高斌战
Owner 北京健翔和牧生物科技有限公司
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