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Isolated culture method for chicken muscle satellite cells

A technique for separating and culturing satellite cells, which is applied in the field of separation and culturing of chicken muscle satellite cells, can solve the problems of poor preservation, low cell viability, and small number of separations, and achieve the effects of short time consumption, good effect, and reduced pollution

Inactive Publication Date: 2020-02-07
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This method combines the principles of loose muscle fibers and free fibrous stromal cells, and optimizes the enzymes used for extracting cells, the attachment medium, the method of purifying cells, and the subsequent cell differentiation method, and solves the problem of cell proliferation in the prior art to the greatest extent. Defects and deficiencies of low activity rate, small number of separations, and poor preservation

Method used

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  • Isolated culture method for chicken muscle satellite cells
  • Isolated culture method for chicken muscle satellite cells
  • Isolated culture method for chicken muscle satellite cells

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Experimental program
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Effect test

Embodiment 1

[0051] (1) Preliminary preparation

[0052] 12 embryonic age SPF eggs (Jinan Xinshengda Biological Engineering Co., Ltd.), 100-mesh cell filter (biosharp), 400-mesh cell filter (biosharp), complete medium, self-made mixed enzyme solution, differentiation medium

[0053]

[0054] ② Complete medium formula: DMEM-F12 medium (Gibco) 180ml

[0055] Fetal bovine serum (Gibco) 30ml (ie 15%)

[0056] Double antibody 4ml (ie 2%)

[0057] ③ Formula for inducing differentiation medium: DMEM-F12 medium (Gibco) 48ml

[0058] Fetal bovine serum (Gibco) 2ml (ie 4%)

[0059] ④ Double Antibody Formula: Penicillin (100kU / L), Streptomycin (100mg / L) mixed solution

[0060]

[0061] Ultrapure water (DDW) 1000ml

[0062] (2) Operation process

[0063] 1. Isolate cells

[0064] ① Take 12-embryo-age SPF eggs, disinfect the bulk with alcohol cotton, knock open with sterile tweezers, take out the chicken embryos and place them on a sterile operating table;

[0065] ②Wipe the legs with alcoh...

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Abstract

The invention belongs to the biotechnology field, and particularly relates to an isolated culture method for chicken muscle satellite cells. The isolated culture method for the chicken muscle satellite cells comprises the following steps of: adopting an SPF (Specific Pathogen Free) chicken, and taking the muscle tissues of the SPF chicken; adding a mixed enzyme solution containing collagenase, type I and pancreatin, wherein a ratio of the collagenase, type I to the pancreatin is 1-2:1; digesting at a temperature of 37 DEG C; and through two-time differential attachment purification for 30 minutes, obtaining a target cell. By use of the method, the defects and the deficiencies in the prior art that a cell survival rate is low, a separation amount is small and storage properties are poor canbe effectively eliminated.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for separating and culturing chicken muscle satellite cells. Background technique [0002] The isolation and purification of static muscle satellite cells commonly used at home and abroad often requires the use of expensive fluorescence-activated cell sorter (FACS) or magnetic-activated cell sorter (MACS), which is complicated and expensive [1] . The simple laboratory separation methods currently used are generally tissue attachment method, two-step digestion method, and single enzymatic hydrolysis method: 1. The tissue attachment method takes a long time, the number of cells obtained is small, and the activity rate is low, which has been gradually eliminated. [2] ; 2. The two-step digestion method usually uses two enzymes to digest in two steps, which takes a long time, is easy to pollute, has low cell viability, and poor adherence (it needs to be coated with g...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077
CPCC12N5/0659C12N2500/84C12N2509/00C12N2509/10
Inventor 戴巍刘学忠顾建红
Owner YANGZHOU UNIV
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