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Process for selectively separating IgY antibody from anseriforme birds eggs and IgY antibody produced therefrom

An avian egg, selective technology, applied in the direction of antibodies, egg-derived immunoglobulins, analytical materials, etc., can solve the problem of successful purification that does not provide any hint, complex and expensive

Inactive Publication Date: 2003-12-31
GOOD BIOTECH CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Furthermore, because the IgY (ΔFc) antibody exists only in birds of the order Anseriformes (Order Anseriformes) including ducks and geese, and because the lipid content in the yolk of birds of the order Anseriformes is reported to be higher than that of chickens and turkeys, etc. Galliform birds, so the above-mentioned traditional method does not provide any hint for the successful purification of IgY(ΔFc) antibody
IgY(ΔFc) antibodies have only been co-precipitated with IgY from duck serum, and this method is complicated and expensive, but still no separate IgY(ΔFc) antibodies have been selectively isolated from egg yolk

Method used

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  • Process for selectively separating IgY antibody from anseriforme birds eggs and IgY antibody produced therefrom
  • Process for selectively separating IgY antibody from anseriforme birds eggs and IgY antibody produced therefrom
  • Process for selectively separating IgY antibody from anseriforme birds eggs and IgY antibody produced therefrom

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] Example 1: Immunization program for stimulating specific antibody production

[0056] Twelve artificially bred ducks (Anasplatyrhynchos var. domestica) aged 16 weeks for antibody and egg production were housed separately. The ducks received an initial subcutaneous injection of 1-5 mg / ml of C-reactive protein (CRP; purified from human ascites), formulated in phosphate buffer at pH 7.5 with The volume is completely emulsified in Freund's adjuvant. The concentration of antigen used is generally in the range of 1 to 5 mg / ml. Following this initial injection, young hens received three additional injections of 1-5 mg antigen every two weeks. One week later, eggs were collected, labeled and stored at 4°C until antibody extraction and purification. During the experiment, the booster procedure was repeated every four weeks. Blood sampling was performed on day 7 after each booster injection. Each blood sample was centrifuged and the resulting serum collected.

Embodiment 2

[0057] Example 2: Antibody Extraction by Duck Egg Yolk

[0058] The egg yolk collected from the eggs laid by the highly immune ducks in Example 1 was washed thoroughly with a weak column of distilled water to thereby remove the egg white. Measure the volume of egg yolk, and then fully mix it with distilled water in an amount 10 times the measured volume of egg yolk. Subsequently, the mixture was kept at 4°C for at least two hours, and then centrifuged for one hour at 10,000 rpm in a Hitachi CR22F centrifuge. A light supernatant layer and a semi-solid easily disturbed layer form in the centrifuge tube.

Embodiment 3

[0059] Embodiment 3: Treat with adsorbent

[0060]In the crude extract prepared in Example 2, one of the following adsorbents was added: 2% (w / v) fumed silica (purchased from Sigma), 3% (w / v) di Silica (Sigma), 3% (w / v) Celite diatomaceous earth (available from Celite Corporation) and 3% or 5% (w / v) Celite diatomaceous earth hyflo-Cel (Celite Corporation). The resulting suspension was incubated at 4°C for 60 minutes with gentle agitation. After the incubation was completed, the adsorbent was precipitated with a Hitachi CR-22F centrifuge at 20,000 rpm at 4°C, and the supernatant and precipitate were collected respectively. A 10 μl sample was taken from each supernatant for non-reducing SDS-polyacrylamide gel electrophoresis analysis (SDS-PAGE).

[0061] like figure 1 As shown, fumed silica had the best adsorption activity in terms of the amount of antibody adsorbed by the adsorbent, and almost no antibody was left in the flow-through solution. Silica exhibits a somewhat wea...

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PUM

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Abstract

The present invention is the method of separating and purifying yolk antibody from egg of anseriformes bird. The required yolk antibody is separated through adsorption and chromatographic step with one kind of water unsoluble and un-charged adsorbent, and the IgY antibody is deposited via salting-out step. The present invention also relates to the yolk antibody and its various application.

Description

technical field [0001] The present invention mainly relates to a method for rapidly separating and purifying yolk antibodies (especially IgY(ΔFc) antibodies) from yolk of Anseriformes birds, and the yolk antibodies obtained by the method. Specifically, the present invention relates to a method for isolating and purifying yolk antibodies from yolks of birds of the order Anseriformes, which uses a water-insoluble and non-charged adsorbent to perform adsorption chromatography to achieve the desired results. Isolation of desired egg yolk antibodies and differential precipitation of the IgY by a salting-out process. The present invention also relates to the use of these IgY antibodies in quantitative or qualitative immunoassays, or for the preparation of pharmaceutical compositions targeting an interesting etiological antigen. Background technique [0002] Antibodies have been widely used in various biological research and clinical applications. Sera obtained from highly immune...

Claims

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Application Information

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IPC IPC(8): A61K39/395C07K16/02G01N33/563G01N33/96
Inventor 邱义能
Owner GOOD BIOTECH CORP
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