Serum-free culture medium for in-vitro culture and proliferation of mesenchymal stem cell

A serum-free medium and stem cell technology, applied in the field of serum-free medium, can solve the problems of threats, scientific research and treatment interference, and large differences in components, so as to avoid potential dangers and promote the normal growth and proliferation of cells.

Pending Publication Date: 2017-07-07
JIANGSU YINUOWAN CELL CLINIC CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the use of fetal bovine serum can satisfy the normal growth and proliferation of mesenchymal stem cells, it can also provide anchorage factors to meet the anchorage growth of anchorage-dependent cells; however, the disadvantages of using animal serum are becoming more and more obvious with the development of treatment research; animals The presence of unknown proteins with unclear components in serum has brought great interference to scientific research and treatment; and often the batches are different, and the composition of animal serum is also very different, and the stability is poor, which has a great impact on the stability of research. Serum may also carry unknown infectious substances and immunogenic heterologous proteins, which will pose a great threat to treatment; in summary, in order to better cultivate and expand mesenchymal stem cells in vitro, to overcome animal serum carrying In the process of cell culture, it is necessary to look for substitutes such as FBS or adopt serum-free culture conditions, and cancel the use of non-human products to reduce potential risks; Serum-based Mesenchymal Stem Cell Culture Medium

Method used

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  • Serum-free culture medium for in-vitro culture and proliferation of mesenchymal stem cell
  • Serum-free culture medium for in-vitro culture and proliferation of mesenchymal stem cell
  • Serum-free culture medium for in-vitro culture and proliferation of mesenchymal stem cell

Examples

Experimental program
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Effect test

Embodiment 1

[0031] DMEM is used as the base medium, and its components are as follows,

[0032]

[0033] Dissolve the above components in ultrapure water or triple distilled water, stir and dissolve at room temperature (18-25°C), adjust the pH to 7.1±0.1 with 0.5mol / L hydrochloric acid, and set the volume to 1L, 0.22um filter membrane After filtration and sterilization, it can be used for in vitro culture and expansion of mesenchymal stem cells.

[0034] Take adipose-derived mesenchymal stem cells as an example:

[0035] In vitro culture of adipose-derived mesenchymal stem cells (ADMSCs): place adipose tissue in a 75cm 2In the cell culture flask, add an equal volume of PBS (phosphate buffer saline) containing 1% double-antibody (penicillin and streptomycin) and 5ug / ml amphotericin B, screw the cap tightly, shake vigorously for 5-10s Afterwards, it was allowed to stand still, and the adipose tissue was layered with PBS, and the adipose tissue was located in the upper layer, and then t...

Embodiment 2

[0039] Based on DMEM / F12, its components are as follows:

[0040]

[0041] Dissolve the above components in ultrapure water or triple distilled water, stir and dissolve at room temperature (18-25°C), and adjust the pH to 7.1±0.1 with 0.5mol / L hydrochloric acid, dilute to 1L, filter through 0.22um After the membrane is filtered and sterilized, it can be used for in vitro culture and expansion of mesenchymal stem cells.

[0042] Take umbilical cord mesenchymal stem cells as an example:

[0043] In vitro culture of umbilical cord mesenchymal stem cells (UCMSCs): Take out the umbilical cord from the collection bottle, place the umbilical cord in a culture dish pre-added with sterile saline, wash away the residual blood of the umbilical cord; sterilize the washed umbilical cord with high pressure Cut the umbilical cord into several small pieces of 2-3cm with bacteria scissors, rinse again, cut the umbilical cord longitudinally, remove one umbilical vein and two umbilical arteri...

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Abstract

The invention discloses a serum-free culture medium for in-vitro culture and extension of a mesenchymal stem cell, and belongs to the technical field of biology. The serum-free culture medium for the in-vitro culture and the extension of the mesenchymal stem cell is definite in chemical composition. According to the serum-free culture medium, a growth factor, insulin, oleic acid, transferin (iron-free), sodium selenite, bovine serum albumin, an antioxidant (beta-mercaptoethanol) dexamethasone and gentamicin are added to a basic culture medium, so that the mesenchymal stem cell can be attached to a culture plate matrix; the in-vitro culture and the extension can be realized under a serum-free condition; a multidirectional differentiation potency characteristic of the mesenchymal stem cell can be maintained; and an adipocyte, an osteoblast and a chondrocyte can be induced under treatment of an in-vitro inducing culture medium. The serum-free culture medium can effectively avoid a potential risk caused by use of a cell product cultured by a serum culture medium.

Description

technical field [0001] The invention relates to a cell culture medium in the field of biotechnology, which is a serum-free culture medium for culturing and expanding mesenchymal stem cells in vitro. Background technique [0002] Mesenchymal stem cells (mesenchymal stem / stromal cells, MSCs) are a type of adult stem cells derived from the mesoderm; they also have multi-directional differentiation potential. Under suitable conditions in vivo and in vitro, MSCs can differentiate into osteoblasts. Chondrocytes, adipocytes, fibroblasts, muscle cells, etc.; due to their high-efficiency differentiation potential, their clinical application has attracted much attention, and they have been used in the treatment of various diseases. MSCs can secrete a large number of biologically active molecules including a variety of growth factors, cytokines and chemokines, etc., and regulate the expression of various genes through the influence of the microenvironment in which they are located, and...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
CPCC12N5/0662C12N5/0667C12N5/0668C12N2500/05C12N2500/24C12N2500/30C12N2500/44C12N2500/90C12N2501/11C12N2501/115C12N2501/135C12N2501/15C12N2501/30C12N2501/33
Inventor 胡鹏曾骥孟庄莉莉
Owner JIANGSU YINUOWAN CELL CLINIC CO LTD
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