Human cell culture medium and preparation method thereof

A medium and cell technology, applied in the field of human cell culture medium and its preparation, can solve the problems of high cost, convenience, poor medium preservation and application, etc., and achieve the goal of promoting synthesis, maintaining biological characteristics, and reducing the use of serum. Effect

Inactive Publication Date: 2018-08-21
EASTERN UNION STEM CELL & GENE ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0013] (1) Cells are susceptible to certain mechanical and chemical factors in serum-free medium, and the preservation and application of the medium are not as convenient as traditional synthetic medium
[0014] (2) Higher cost
[0015] (3) Highly targeted, a serum-free medium may only be suitable for the cultivation of a certain type of cells

Method used

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  • Human cell culture medium and preparation method thereof
  • Human cell culture medium and preparation method thereof
  • Human cell culture medium and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] A medium for cultivating human cells, consisting of 20 kinds of amino acids, 11 kinds of vitamins, 13 kinds of inorganic salts, 2 kinds of bases, 2 kinds of energy metabolism substances such as nucleosides and glucose, and 2 kinds of lipids such as linoleic acid And precursor substances and putrescine and other three components.

[0060] Among them, the formula of amino acid components is: glycine 0.27~0.41mmol / L; L-valine 0.53~0.80mmol / L; L-isoleucine 0.52~0.78mmol / L; L-leucine 0.53~0.79mmol / L; L-phenylalanine 0.26~0.39mmol / L; L-methionine 0.13~0.20mmol / L; L-tryptophan 0.05~0.08mmol / L; L-threonine 0.53~0.79mmol / L; L-serine 0.27~0.41mmol / L; L-alanine 0.20~0.29mmol / L; L-proline 0.27~0.41mmol / L; L-cystine 0.22~0.33mmol / L; L -Tyrosine disodium salt 0.30~0.45mmol / L; L-lysine hydrochloride 0.54~0.81mmol / L; L-arginine hydrochloride 0.41~0.62mmol / L; L-histidine salt Salt 0.14~0.22mmol / L; L-glutamine 2.72~4.08mmol / L; L-glutamic acid 0.34~0.51mmol / L; L-asparagine 0.14~0.21mm...

Embodiment 2

[0072] Prepare 1000mL human cell culture medium:

[0073] (1) Weigh 0.34mmol of glycine, 0.66mmol of L-valine, 0.65mmol of L-isoleucine, 0.66mmol of L-leucine, 0.32mmol of L-phenylalanine, and 0.16mmol of L-methionine. mmol, L-tryptophan 0.06mmol, L-threonine 0.66mmol, L-serine 0.34mmol, L-alanine 0.25mmol, L-proline 0.34mmol, L-cystine 0.28mmol, L- Tyrosine disodium salt 0.38mmol, L-lysine hydrochloride 0.68mmol, L-arginine hydrochloride 0.52mmol, L-histidine hydrochloride 0.18mmol, L-glutamine 3.4mmol, L-glutamic acid 0.43mmol, L-asparagine 0.17mmol, L-aspartic acid 0.2mmol, folic acid 7.8E-03mmol, thiamine 9.7E-03mmol, riboflavin 8.7E-04mmol, vitamin B122. 1E-04mmol, biotin 4.8E-05mmol, calcium pantothenate 6.9E-03mmol, pyridoxal hydrochloride 1.6E-02mmol, pyridoxine hydrochloride 5.8E-05mmol, inositol 5.2E-02mmol, choline chloride 4.3E- 02mmol, Nicotinamide 2.6E-02mmol, Sodium Chloride (NaCl) 88mmol, Sodium Bicarbonate (NaHCO 3)32mmol, sodium dihydrogen phosphate dihydr...

Embodiment 3

[0078] The method for cultivating umbilical cord mesenchymal stem cells using the human-derived cell culture medium prepared in Example 2:

[0079] (1) adding fetal bovine serum (FBS) with a volume fraction of 7% to the human-derived cell culture medium prepared in Example 2;

[0080] In order to compare the test effect, the culture medium of the control group was used: using Iskoff's modified medium (IMDM) as the base medium, adding 7% fetal bovine serum (FBS) by volume fraction;

[0081] (2) Take the third-generation umbilical cord mesenchymal stem cells, add physiological saline and centrifuge to wash them, then inoculate them into two culture flasks respectively, and add the culture medium of the experimental group and the control group respectively;

[0082] (3) At 37°C, 5% carbon dioxide (CO 2 ) conditions and cultured to 90% confluency when subcultured.

[0083] Such as figure 1 Shown is the cell morphology at passage 6. Wherein A adopts the human cell culture mediu...

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Abstract

The invention discloses a human cell culture medium which comprises amino acids, vitamins, inorganic salts, hypoxanthine, thymidine, glucose, sodium pyruvate, linoleic acid, lipoic acid, putrescine and 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid. Due to improvement on components of the culture medium, according to the growth and metabolism characteristics of a human cell line, components such as pyridoxine hydrochloride, copper sulfate, ferrous sulfate, zinc sulfate, magnesium chloride, disodium hydrogen phosphate, sodium hypoxanthine, linoleic acid, lipoic acid, putrescine, thymidine and the like are increased, a microenvironment for growth of cells in the human body is met, and synthesis of human cell bio-enzymes can be promoted. The concentration of each component in the culture medium is optimized, the instability brought by a novel component is reduced, biological characteristics of the cultured cells are well maintained, and usage of serum during cell culture can be reduced or avoided. According to the test, the culture medium can be suitable for adherent culture and suspension culture and is applicable to the culturing of most of the human cells.

Description

technical field [0001] The invention relates to a composition for cell culture, in particular to a human cell culture medium and a preparation method thereof. Background technique [0002] Since the establishment of tissue and cell culture technology, the use of human cells to carry out modern biopharmaceuticals, especially the use of human cells for tissue engineering research or biotherapy, has continued to flourish. Among these technologies, the development of suitable medium is the most core and basic key technology. [0003] By far, in vitro expansion of human cells is most commonly performed using basal media supplemented with fetal bovine serum. However, using fetal bovine serum to cultivate cells also brings many problems. First of all, fetal bovine serum albumin is a heterologous protein. After the cells cultured with fetal bovine serum are used for treatment, it may cause infection in the treated person, which is potentially dangerous. Second, fetal bovine serum...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/00
CPCC12N5/0018C12N2500/12C12N2500/14C12N2500/16C12N2500/20C12N2500/22C12N2500/24C12N2500/32C12N2500/34C12N2500/36C12N2500/38C12N2500/40C12N2500/46C12N2501/999
Inventor 陆梦晓刘洋郑媛媛刘红琼范星洲盛小琦贡波吴明远
Owner EASTERN UNION STEM CELL & GENE ENG
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