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Method for rapidly preparing large number of chick embryo fibroblasts

A fibroblast and chicken embryo technology, applied in the field of cell culture, can solve the problems of long operation time, easy pollution, unsuitable for mass production, etc., and achieve the effect of increased harvest and short operation time

Inactive Publication Date: 2017-01-11
ZHAOQING DAHUANONG BIOLOGIC PHARMA +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The operation steps are numerous and easy to pollute, and the operation time is usually about 3 hours, and the harvested chicken embryo fibroblasts are very few, which is not suitable for mass production at all.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] A method for rapidly preparing a large amount of chicken embryo fibroblasts, comprising the following steps:

[0035] 1) Take chicken embryos: take 11-day-old SPF chicken embryos, put them into the ultra-clean table, wipe and disinfect them with 75% alcohol cotton, sterilize the embryo eggs with flames, and use sterile tweezers to break the egg shells , use new sterile tweezers to take out the chicken embryo;

[0036] 2) trypsin washing: cut the chicken embryo taken out in step 1) into 0.5-1cm block tissue in diameter, then add 0.25wt% trypsin solution to wash twice, the volume of trypsin solution is 5 times the volume of the tissue block ;

[0037] 3) Homogenization: use a homogenizer under aseptic conditions to pulverize and homogenize the chicken embryo treated in step 2) until the diameter of the chicken embryo tissue block is as small as 1 mm;

[0038] 4) Digestion: add 0.25wt% trypsin solution to the homogenized tissue block in step 3), the volume ratio of tryps...

Embodiment 2

[0044] A method for rapidly preparing a large amount of chicken embryo fibroblasts, comprising the following steps:

[0045] 1) Take chicken embryos: Take 9-day-old SPF chicken embryos, put them into the ultra-clean table, wipe and disinfect them with 75% alcohol cotton, sterilize the embryo eggs with flames, and use sterile tweezers to break the eggshells , use new sterile tweezers to take out the chicken embryo;

[0046] 2) trypsin washing: cut the chicken embryo taken out in step 1) into a 0.5-1cm block tissue in diameter, then add 0.25wt% trypsin solution to rinse for 3 times, the volume of the trypsin solution is 3 times the volume of the tissue block ;

[0047] 3) Homogenization: use a homogenizer under aseptic conditions to pulverize and homogenize the chicken embryo treated in step 2) until the diameter of the chicken embryo tissue block is as small as 1mm;

[0048] 4) Digestion: add 0.25wt% trypsin solution to the homogenized tissue block in step 3), the volume rati...

Embodiment 3

[0053] A method for rapidly preparing a large amount of chicken embryo fibroblasts, comprising the following steps:

[0054] 1) Take chicken embryos: take 10-day-old SPF chicken embryos, put them into the ultra-clean table, wipe and disinfect them with 75% alcohol cotton, sterilize the embryo eggs with flames, and use sterile tweezers to break the egg shells , use new sterile tweezers to take out the chicken embryo;

[0055] 2) trypsin washing: cut the chicken embryo taken out in step 1) into a block tissue with a diameter of 0.5-1cm, then add 0.25wt% trypsin solution to wash 3 times, the volume of the trypsin solution is equal to the volume of the tissue block;

[0056] 3) Homogenization: use a homogenizer under aseptic conditions to pulverize and homogenize the chicken embryo treated in step 2) until the diameter of the chicken embryo tissue block is as small as 1 mm;

[0057]4) Digestion: add 0.25wt% trypsin solution to the homogenized tissue block in step 3), the volume r...

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Abstract

The invention discloses a method for rapidly preparing a large number of chick embryo fibroblasts. The method comprises steps as follows: 1) taking of chick embryos; 2) pancreatin washing; 3) homogenization; 4) digestion; 5) dispersion; 6) subculture, and obtaining of the chick embryo fibroblasts. The yield of the chick embryo fibroblasts obtained with the method is more than 20 times that of chick embryo fibroblasts obtained with traditional methods, and the operation time is short. Practices prove that each chick embryo can be transferred in 10-20 T225 cell bottles.

Description

technical field [0001] The invention relates to a method for rapidly preparing a large amount of chicken embryo fibroblasts, which belongs to the technical field of cell culture. Background technique [0002] In addition to being used in related research in the field of virology, chicken embryo fibroblasts (CEF) are also widely used in the production of veterinary vaccines, such as infectious bursal disease, Marek's disease, Newcastle disease, etc., and can also be used to express some genetic engineering products Wait. [0003] At present, there are various methods for isolating chicken embryo fibroblasts, which are mainly suitable for the preparation of a small amount of chicken embryo fibroblasts in the laboratory, which usually includes aseptically taking chicken embryos, removing the head, limbs, internal organs, etc., chopping them up, and using PBS or Hank's solution washing, trypsinization, blowing, gauze filtration, centrifugation to recover cells, cell counting, c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/073
CPCC12N5/0603C12N2509/00
Inventor 陈瑞爱徐家华王新秋孙贝贝任常宝谢小雨
Owner ZHAOQING DAHUANONG BIOLOGIC PHARMA
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