Preparation method of multivalence anti-venin yolk antibody

A technology of egg yolk antibody and snake venomous chicken, applied in the direction of immunoglobulin from eggs, etc., can solve the problems of difficult storage and transportation, backward purification process, high allergic reaction rate, etc., achieve considerable social and economic benefits, broad application prospects, preparation low cost effect

Inactive Publication Date: 2007-10-03
广州医学院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In summary, the antivenoms produced in the prior art have the following deficiencies: one is that the products are mostly liquid preparations, which are not easy to store and transport, and have a short validity period; Re-inject the corresponding antivenom serum, but the condition of the snakebite is acute, progresses rapidly, and the condition is dangerous and the mortality rate is high, and the diagnosis before the medication will inevitably cause delay in the treatment time; the third is that the refining and purification process is backward, so that the product is less than Less active and higher rate of allergic reactions
Moreover, the low yield and high cost lead to high prices, which are greatly limited in clinical application.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1: Preparation of trivalent (cobra + viper + Agkistrodon) antivenom IgY

[0024] (1) Preparation of 3 kinds of snake venom antigens:

[0025] Preparation of 3 kinds of single venom antigens: Extract the venoms of adult Chinese cobra, domestic viper, and Agkistrodon with a self-made snake venom extractor (patent number: ZL200420046279.9), and add the venom at a ratio of 1:4 (V / V) Diluted with distilled water, centrifuged, and vacuum-dried at low temperature to obtain a freeze-dried powder. Dissolve 50mg of snake venom freeze-dried powder in 5ml of 0.1M phosphate buffered saline (PBS) solution, add 0.5ml of 0.25% glutaraldehyde and place it at 30°C for 2 hours, then use SephadexG-25 chromatography to desalt and determine snake venom LD 50 , and fully emulsified with Freund's adjuvant at 1:1 (V / V) to make antigen.

[0026] (2) Immunize hens with 3 kinds of snake venom attenuated antigens:

[0027] Take 22-week-old Laihang hens (body weight: 1.0±0.1kg), and inje...

Embodiment 2

[0033] The difference from Example 1 is that in the step (2), the attenuated single snake venom antigens of 3 kinds (Chinese cobra, domestic viper, Agkistrodon acutus) are mixed in a ratio of 1:1:1 . A 22-week-old Laihang hen (body weight: 1.0±0.1kg) was taken, and the mixed snake venom antigen was injected into the body of the chicken through subcutaneous and muscular sites of wings, chest, abdomen and back of the chicken. The interval between the first immunization and the second immunization was 2 weeks, and the interval between subsequent immunizations was 4 weeks. The eggs were collected 2 weeks after the initial immunization, and the eggs collected after 2 weeks were trivalent antivenom IgY chicken eggs.

[0034] In the step (3), the ratio of deionized water to egg yolk liquid is 5:1, adjust the pH to 4, place at 4°C for 6 hours, centrifuge at 4°C for 15min / 19000g, and concentrate the supernatant by ultrafiltration to a solution concentration of 15mg / ml; Then use ammon...

Embodiment 3

[0036]The difference from Examples 1 and 2 is that in the step (2), the attenuated single venom antigens of Chinese cobra, domestic viper, and Agkistrodon acutus are passed through chicken wings, chest, abdomen and back respectively. Three 22-week-old Laihang hens (body weight: 1.0±0.1kg) were injected subcutaneously and intramuscularly. The interval between the first immunization and the second immunization was 2 weeks, and the interval between subsequent immunizations was 4 weeks. The eggs were collected at the second week after the initial immunization, and the eggs collected after 2 weeks were eggs of three different monovalent anti-venom IgY eggs.

[0037] In the step (3), three kinds of monovalent antivenom IgY are directly extracted by water dilution method, and then these three kinds of monovalent antivenom IgY are mixed uniformly in a ratio of 1:1:1.

[0038] In the step (3), the ratio of deionized water to egg yolk liquid is 12:1, adjust the pH to 8, place at 4°C fo...

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Abstract

This invention relates to a preparation method of multivalent anti-snake venom chicken yolk antibody. It includes following steps:(1) collect different kinds of snake venom antigen; (2) preparation of chicken ovum of anti-snake venom chicken yolk antibody, and infuse snake venom to chicken for immunization, after 20 to 4 weeks continuously collect egg for 6 to 10 mouths.(3) extract multivalent anti-snake venom chicken yolk antibody. This product can replace horse serum for injection, and be able to developed as oral using drug which has extensive application prospects in prevention of snake bite.

Description

technical field [0001] The invention relates to a preparation method of snake venom antibody, in particular to a preparation method of anti-snake venom chicken egg yolk antibody (Immunoglobulin-yolk, IgY). More specifically, it is a preparation method of multivalent anti-snake venom chicken egg yolk antibody. Background technique [0002] At present, the existing routine production of antivenom serum is prepared by immunizing horses. Usually, horses are first immunized with detoxified snake venom. After the plasma is digested with gastric enzymes and salted out with ammonium sulfate, it is further refined and purified to remove A polyclonal antiserum preparation with Fc fragment IgG as the main component. [0003] In summary, the antivenoms produced in the prior art have the following deficiencies: one is that the products are mostly liquid preparations, which are not easy to store and transport, and have a short validity period; Re-inject the corresponding antivenom serum...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/02
Inventor 孔天翰董伟华刘四红祁俊华陈熹陈学文
Owner 广州医学院
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