Blood serum substituting agent and blood serum free rainbow trout embryo cell line culture medium

A serum replacement and embryonic cell technology, applied in embryonic cells, tissue culture, animal cells, etc., can solve the problems of complex purification and identification process of cell products, difficulty in separation and purification of cell products, allergic reactions of heterogeneous serum, etc., to avoid cell Allergic reaction, low price, fast growth effect

Inactive Publication Date: 2008-02-20
HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0004] The purpose of the present invention is to solve the problem of high cost, inhumane, unstable quality of serum used in culture medium, certain substances affecting cell growth and the expression of some cell functions, difficult separation and purification of cell products, purification and identification of various cell products in the later stage The process is complicated, it may cause allergic reactions to heterogeneous serum and there is no serum-free embryonic cell line medium suitable for rainbow trout at present, and a serum replacement agent and serum-free rainbow trout embryonic cell line medium are provided

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  • Blood serum substituting agent and blood serum free rainbow trout embryo cell line culture medium

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specific Embodiment approach 1

[0010] Embodiment 1: The serum replacement agent in this embodiment consists of 0.3g recombinant human insulin-like growth factor-1 (rhIGF-1), 0.5g recombinant human basic fibroblast growth factor (rh-bFGF), 0.65g platelet-derived Growth factor B (PDGF-B), 1.5g bovine serum albumin (BSA), 5g bovine transferrin, 34g β-mercaptoethanol, 0.294g glutamic acid, 0.110g sodium pyruvate and 2-3g non-essential amino acids.

[0011] The serum replacement agent of this embodiment has a wide range of applications and can be used for serum-free culture of fish and animal cells. The non-essential amino acids in this embodiment were purchased from Shanghai Weike Biochemical Reagent Co., Ltd., which did not contain glutamic acid. The serum replacement agent of this embodiment does not contain cholesterol.

specific Embodiment approach 2

[0012] Specific embodiment two: In this embodiment, the serum-free rainbow trout embryo cell line culture medium is composed of RPMI-1640 medium and the serum replacement agent described in specific embodiment one; every 1000mL serum-free rainbow trout embryo cell line medium contains the specific embodiment. 44.354-45.354 g of the above-mentioned serum replacement agent, and the balance is RPMI-1640 medium.

specific Embodiment approach 3

[0013] Specific embodiment three: present embodiment rainbow trout embryonic cell line culture method:

[0014] 1. Collection of rainbow trout embryos

[0015] In this embodiment, the rainbow trout embryos are obtained from high-quality rainbow trout fertilized eggs of the Bohai Cold Water Fish Experimental Station of the Heilongjiang Fisheries Research Institute of the Chinese Academy of Fishery Sciences.

[0016] 2. Embryo (blastocyst-late gastrulation) cell isolation

[0017] After the collected rainbow trout embryos passed the microscopic examination, they were cultured at 18°C ​​until the late stage of blastula or late gastrula, and then the rainbow trout embryos were repeatedly dipped in 1×PBS solution for 3 to 4 times, and then washed with 70% alcohol Soak for disinfection and sterilization, and then wash three times with 1×PBS solution with a pH value of 7.4, then put it in RPMI-1640 complete medium, use pointed tweezers to peel off the egg membrane to separate the ce...

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Abstract

A serum alternative and serum-free rainbow trout embryo cell line medium relates to a serum alternative and a serum-free embryo cell line medium. The invention solves the problems of too high cost and unstable quality of the serum used in the medium, and the present lack of suitable rainbow trout serum-free embryo cell line medium. The serum alternative consists of 0.3g rhIGF-1, 0.5g rh-bFGF, 0.65g PDGF-B, 1.5g bovine serum albumin, 5g bovine transferring, 34g Beta- mercaptoethanol, 0.294g glutamatic acid, 0.110g sodium pyruvate, and 2 to 3g nonessential amino acid. The medium consists of RPMI-1640 medium and the serum alternate. The serum alternate of the invention has price as low as 60 percent of that of bovine serum and stable quality. The rainbow trout embryo cell line medium cultured by the serum-free rainbow trout embryo cell line medium of the invention has the advantages of rapid breeding speed and long regenerating cycle.

Description

technical field [0001] The invention relates to a serum replacement agent and a serum-free culture medium for embryonic cell lines. Background technique [0002] Synthetic media used for culturing cells in vitro require the addition of some natural components, the main natural component being serum, especially bovine serum. However, the use of serum in the culture medium has the following disadvantages: 1. High cost of serum: 500mL bovine serum price is 3,000-4,000 yuan; 2. Inhumane collection of living organisms; 3. The quality of serum is unstable, and there are differences between batches, which affects the experimental data 4. Serum contains complex components, some of which can inhibit the growth of certain cells, affect the expression of cell growth and certain functions of cells, and have a dedifferentiation effect on the in vitro culture of certain cells; 5 .A large number of complex proteins in serum bring great difficulties to the separation and purification of va...

Claims

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Application Information

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IPC IPC(8): C12N5/06C12N5/073
Inventor 薛淑群尹洪滨
Owner HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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