CN1072507C

CN1072507C - Method of extracting yolk antibody

Info

Publication number: CN1072507C
Application number: CN 97108886
Authority: CN
Inventors: 凌育燊; 郭予强; 杨连楷; 李健东; 罗映霞; 詹乐群; 余双祥; 邱深本; 朱国锋
Original assignee: INST OF FOWL SCIENCES GUANGDONG PROV
Current assignee: INST OF FOWL SCIENCES GUANGDONG PROV
Priority date: 1997-04-18
Filing date: 1997-04-18
Publication date: 2001-10-10
Anticipated expiration: 2017-04-18
Also published as: CN1196957A

Abstract

The present invention belongs to a method for manufacturing an animal immune antibody. Most of yolk antibody extracting methods at home and abroad have the purpose of immunology experiments in an experimental laboratory to extract IgY in yolk antibodies; therefore, most of the yolk antibody extracting methods have the defects of high purity requirements, complex operation, low output, low temperature operation requirements and difficult production and popularization. The yolk antibody extracting method provided by the present invention can use a conventional dilute liquid and a displacement liquid at normal temperature, and the operation is easy; therefore, the present invention is easy to organize scaled production.

Description

A kind of method of extracting yolk antibody

The invention belongs to fowl, poultry immune antibody and method for making.

Utilize immune antibody control people and animals, or some infectious disease of birds, existing quite long history, traditional antiserum production be utilize white mice, exempt from, animals such as horse, cattle, sheep and pig by certain immunity and booster immunization after, collect its serum and make.Because among the fowl group behind vaccine virus immunization, corresponding disease still happens occasionally, as replenishing of vaccine, domestic in the later stage eighties, begin common use high-immunity egg yellow liquor, but this class high-immunity egg yellow liquor prepares by non-SPF hen egg, thereby its widely used result, inevitably bring the people for sending out source of disease, through the former propagation of egg vertical infection sexually transmitted disease (STD), as the Sha Shi bacillus, escherichia coli, leukemia virus etc., the significant quantities of fat that contains with egg yolk itself, compositions such as protein, also may cause similar anaphylactoid " egg yolk disease " problem after injecting the fowl body, so the research of extracting yolk antibody liquid just having put on the agenda, is to want to seek the IgG succedaneum of serum to the most of purpose of yolk antibody method for extracting both at home and abroad, requirement purity height, thereby generally yield poorly, the method for extracting that the sub-fraction yolk antibody is arranged also is to carry out trivial operations at low temperatures, equipment requirements is higher, cost is higher, though some use at the scene, because of equipment, operation and cost problem and can not promoting.

The objective of the invention is characteristics, propose a kind of each method of system that is suitable for mass production, easy and simple to handle, egg yolk extracting antibody that cost is low, and obtain satisfactory result according to yolk antibody.

The implementation process of method for extracting provided by the present invention is such: behind the conventional cleaning-sterilizing high-immunity egg shell surface, shell and get egg yolk, remove the albumen that adheres on the membrane of yolk as far as possible, use aquesterilisa rinsing egg yolk three times repeatedly, puncture membrane of yolk after draining, collect egg yolk, in egg yolk: the ratio of diluent=1: 5 (volume ratio) adds 0.1M, PH is 5.5 NaCl solution, after stirring, stir on the limit, the limit is the stock solution of the Polyethylene Glycol of adding 60% slowly, making final concentration is 3.5%, fully behind the mixing, in room temperature 18-20 ℃ of following standing over night, with Cotton Gossypii and filtered through gauze, remove free-fat and sediment, collect filtrate and filter above-mentioned filtrate to remove fine impurities and antibacterial with the malleation duplex strainer that the filter membrane aperture is respectively 0.8 μ m and 0.45 μ m composition, using molecular cut off at last is that 10000 ultrafilter is removed residual Polyethylene Glycol and other salt, and suitably concentrate, do the active inspection of conventional antibody then, microbial check and safety testing obtain qualified finished product after the product packing.The method for extracting of continuing to use in antibody method for extracting provided by the invention and the prior art has following advantage and characteristics:

1, to be about the total protein concentration of 83.5%～95% extract be 3.70%-4.23 gram % to effective extraction ratio of yolk antibody, and wherein globulin accounts for the 43.9%-52.9% (adopting biuret method to measure) of total protein, and antibody activity meets the requirements.

2, product does not contain pathogenic microorganism alive, and is safe, do not have any toxicity, side effect.

3, this method overall process all can at room temperature be operated, and equipment requirements is not high, and is easy and simple to handle, and cost is lower, is easy to tissue and produces in enormous quantities.

Embodiment:

Getting egg shells and gets Huang, remove the attached albumen that sticks on the membrane of yolk as far as possible, then egg yolk is placed sterile purified water, cyclic washing three times, to remove the albumen on the membrane of yolk, puncture membrane of yolk after draining, collect egg yolk, after egg yolk stirred, the 0.1M that adds 5 times of volumes of egg yolk homogenate, PH is 5.5 NaCl diluent, make the dilution egg yolk liquid, slowly add 60% Polyethylene Glycol stock solution then, making final concentration is 3.5%, fully behind the mixing in room temperature 18-20 ℃ of following standing over night, use Cotton Gossypii and filtered through gauze then, remove sediment, collect filtrate, reuse filter membrane hole is toward the malleation duplex strainer of difference 0.8 μ m and 0.45 μ m, filter above-mentioned filtrate, remove fine impurities and antibacterial, the reuse molecular cut off is that 10000 ultrafilter is removed residual Polyethylene Glycol and other salt, and suitably concentrates, carry out antibody activity inspection and packing and become product, through measuring with biuret method, the effective extraction ratio average out to of antibody more than 90% in the product, extract total protein concentration average out to 3.97%, it is average 48.46% that globulin accounts for total protein, and antibody activity meets the requirements.

Claims

1, stud bird poultry is used the method for extracting of Yolk immune antibody, the feature of this method is: at first the egg yolk of high-immunity egg is made egg yolk homogenate, the 0.1M, the pH that add 5 times of homogenate volumes in egg yolk homogenate are 5.5 sodium chloride diluent, it is 3.5% that the Polyethylene Glycol that adds 60% concentration then makes it final concentration, under room temperature, left standstill 24 hours, remove by filter fat, sediment, fine impurities and antibacterial by three times then, just obtain the antibody finished product.

2, according to the antibody method for extracting described in the claim 1, it is characterized in that filtering step is: earlier with Cotton Gossypii or filtered through gauze sediment, reuse filter membrane aperture is the malleation duplex strainer that 0.8 μ m and 0.45 μ m form, remove fine impurities and antibacterial, at last, using molecular cut off is that 10 000 ultrafilter is removed residual Polyethylene Glycol and other salt, filters overall process and all carries out under room temperature.