Anti hepatitis B virus chicken yolk immune globulin IgY and its separating method

An immunoglobulin and hepatitis B virus technology, applied in the field of specific antibodies and their separation, can solve problems such as complex methods, and achieve the effects of simple process, high purity and high activity

Inactive Publication Date: 2007-03-28
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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  • Anti hepatitis B virus chicken yolk immune globulin IgY and its separating method
  • Anti hepatitis B virus chicken yolk immune globulin IgY and its separating method

Examples

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Embodiment 1

[0020] (1) Animal immunization Six-month-old laying hens were kept in isolation during immunization and laying, and 60 μg / ml hepatitis B virus surface antigen HbsAg was mixed with an equal volume of adjuvant as the immunogen. Take 1ml of this immunogen solution, and inject each chicken intramuscularly at four points. For the first immunization, the antigen is mixed with complete adjuvant, and a booster injection is given two weeks later, and after that, the booster injection is given every other month. During the booster immunization, the antigen is mixed with incomplete adjuvant. During the immunization period, eggs are collected every day and stored at 4°C to be separated;

[0021] (2) Separation and purification Separate chicken yolk, dilute with 8 times the volume of deionized water, adjust the pH to 5 with 0.1M HCl, add sodium chloride to 0.15M, place at 4°C for 5.5 hours, and centrifuge at 3500 rpm Separation for 23 minutes, discard the precipitate, add sodium sulfate t...

Embodiment 2

[0025] (1) Animal immunization Six-month-old laying hens were kept in isolation during immunization and laying, and 60 μg / ml hepatitis B virus surface antigen HbsAg was mixed with an equal volume of adjuvant as the immunogen. Take 1ml of this immunogen solution, and inject each chicken intramuscularly at four points. For the first immunization, the antigen is mixed with complete adjuvant, and a booster injection is given two weeks later, and after that, the booster injection is given every other month. During the booster immunization, the antigen is mixed with incomplete adjuvant. During the immunization period, eggs are collected every day and stored at 4°C to be separated;

[0026] (2) Separation and Purification Separate chicken egg yolk, dilute it with 6 times the volume of deionized water, adjust the pH to 5.5 with 0.1M HCl, add sodium chloride to 0.2M, place at 4°C for 5 hours, and centrifuge at 3500 rpm Separation for 20 minutes, discard the precipitate, add sodium sul...

Embodiment 3

[0028](1) Animal immunization Six-month-old laying hens were kept in isolation during immunization and laying, and 60 μg / ml hepatitis B virus surface antigen HbsAg was mixed with an equal volume of adjuvant as the immunogen. Take 1ml of this immunogen solution, and inject each chicken intramuscularly at four points. For the first immunization, the antigen is mixed with complete adjuvant, and a booster injection is given two weeks later, and after that, the booster injection is given every other month. During the booster immunization, the antigen is mixed with incomplete adjuvant. During the immunization period, eggs are collected every day and stored at 4°C to be separated;

[0029] (2) Separation and Purification Separate chicken egg yolk, dilute it with 9 times the volume of deionized water, adjust the pH to 4.5 with 0.1M HCl, add sodium chloride to 0.1M, place at 4°C for 6 hours, and centrifuge at 3500 rpm Separation for 25 minutes, discard the precipitate, add sodium sulf...

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Abstract

The invention discloses anti-hepatitis B virus immunoglobulin of yolk and separation method. The separation method includes the following steps: animal immune; separating and purifying yolk; diluting by de-ionized water; adjusting pH; adding sodium chloride; setting for 5-6h; centrifugal separation; removing precipitation; adding sodium sulfate into supernatant to process the first salting out; centrifugalizing out protein precipitation; adding de-ionized water until the precipitation be fully dissolved; adding sodium sulfate to process the second salting out; centrifugal separation; the gained protein precipitation is added into de-ionized water until it is fully dissolved; filtering by gel-column; the eluted protein flow is monitored by ultraviolet photometer to absorb the 280 nm light wave and collect the first spectral band, concentrated by reversal dialysis, dialyzed to remove salt to gain the anti-hepatitis B virus IgY. The invention has the advantages of simple technology, low cost, and better sample quality.

Description

technical field [0001] The invention relates to a specific antibody and its separation method, in particular to an anti-hepatitis B virus immunoglobulin and its separation method. Background technique [0002] Hepatitis B is a serious infectious disease, and there are about 200 million hepatitis B virus carriers in the world. Hepatitis B virus surface antigen HbsAg is a group of proteins that constitute the outer membrane of the virus. One of its important functions is to recognize the receptors on the liver cell membrane, make the virus contact with the cell, and then invade the cell for replication. When HbsAg is positive in the blood, it is a sign of HBV infection. HbsAg appears in acute or chronic hepatitis and "healthy" virus carriers. Therefore, detection of hepatitis B virus surface antigen HbsAg is of great significance to the diagnosis and treatment of the disease. In terms of immune detection, specific antibody IgG obtained from the blood of rabbits, sheep and ot...

Claims

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Application Information

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IPC IPC(8): C07K16/02
Inventor 孙淑清马向辉
Owner TIANJIN UNIV
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