A method for selectively purifying and expanding stem cells in large quantities

Abstract

The invention discloses a method for selectively purifying and expanding stem cells in large quantities. The method includes selectively expanding stem cells using absorbent cotton fibers or modified absorbent cotton fibers. The use of absorbent cotton fiber-based polymer materials can effectively increase the value of purified stem cells and maintain the same characteristics of stem cells, which is conducive to the realization of large-scale expansion of stem cells.

Description

technical field

[0001] The invention relates to the field of biomedicine, in particular to a method for selectively purifying and expanding stem cells in large quantities based on absorbent cotton fibers. The use of absorbent cotton fibers can effectively increase the value of stem cells and maintain the characteristics of stem cells, which is conducive to the realization of a large number of stem cells. Background technique

[0002] Stem cells play an important role in life science research, medical treatment and drug screening. However, the growth conditions of stem cells are harsh, the number in the organism is rare, and they are prone to aging and differentiation under general culture conditions. Therefore, how to obtain a large number of stem cells is one of the difficult problems in the field of life sciences.

[0003] In addition to hematopoietic stem cells, other types of stem cells generally require adherent growth or aggregate growth (neural stem cells) to achiev...

Examples

Embodiment 1

[0021] 1. Extract single-strand fibers from the medical degreasing surface, and the fiber diameter is 5-10 μm.

[0022] 2. Plasma surface treatment of absorbent cotton fibers (PDC-M plasma cleaning machine). The conditions of ammonia plasma treatment were: input power 20W; treatment time 60s; activation frequency 13.65MHz; pressure 15 Pa; ammonia gas flow rate 150 mL / min.

[0023] 3. After the surface treatment of the defatted fiber is completed, use ultraviolet light to irradiate for 10 hours to sterilize for later use.

[0024] 4. Use 1% agarose (Ultrapure agarose, Invitrogen, Carlsbad, CA) to coat the bottom of the culture dish, and then inoculate PC12 cell suspension, for example, inoculate 2×10 cells per well in a six-well plate. 4 cells / mL.

[0025] 5. Put the sterilized degreased fibers in a petri dish and culture them for 5 days. During the culture period, replace the culture medium every 2-3 days. Finally, collect the cell balls on the fibers. The diameter of the ce...

Embodiment 2

[0028] Basically the same as in Example 1, the difference is that the anti-adsorption cell material used is gelatin, and the treatment conditions of ammonia plasma are: treatment time is 10s; frequency is 10kHz; pressure is 10 Pa; ammonia gas flow rate is 50 mL / min, high temperature and high pressure sterilization is used after the surface treatment of defatted fiber is completed; the present invention cultures adipose-derived mesenchymal stem cells for 9 days, and the number ranges from 2×10 4 Cells / mL expanded to 7×10 5 cells / mL, the expression levels of genes Oct 4, Sox 2 and Nanog, which represent the characteristics of stem cells, also increased significantly.

Embodiment 3

[0030] Basically the same as Example 1, the difference is that the treatment conditions of ammonia plasma are: treatment time is 35s; frequency is 40kHz; pressure is 5 pa; ammonia gas flow rate is 100 mL / min, after the surface treatment of degreased fiber is completed, high temperature Autoclave sterilization; after the surface treatment of defatted fibers is completed, soak and sterilize with iodine solution, and the present invention cultivates induced pluripotent stem cells for 15 days, and the number ranges from 2×10 4 Cells / mL expanded to 3×10 6 cells / mL, the expression levels of genes Oct 4, Sox 2 and Nanog, which represent the characteristics of stem cells, also increased significantly.

technical field

[0001] The invention relates to the field of biomedicine, in particular to a method for selectively purifying and expanding stem cells in large quantities based on absorbent cotton fibers. The use of absorbent cotton fibers can effectively increase the value of stem cells and maintain the characteristics of stem cells, which is conducive to the realization of a large number of stem cells. Background technique

[0002] Stem cells play an important role in life science research, medical treatment and drug screening. However, the growth conditions of stem cells are harsh, the number in the organism is rare, and they are prone to aging and differentiation under general culture conditions. Therefore, how to obtain a large number of stem cells is one of the difficult problems in the field of life sciences.

[0003] In addition to hematopoietic stem cells, other types of stem cells generally require adherent growth or aggregate growth (neural stem cells) to achiev...

Examples

Embodiment 1

[0021] 1. Extract single-strand fibers from the medical degreasing surface, and the fiber diameter is 5-10 μm.

[0022] 2. Plasma surface treatment of absorbent cotton fibers (PDC-M plasma cleaning machine). The conditions of ammonia plasma treatment were: input power 20W; treatment time 60s; activation frequency 13.65MHz; pressure 15 Pa; ammonia gas flow rate 150 mL / min.

[0023] 3. After the surface treatment of the defatted fiber is completed, use ultraviolet light to irradiate for 10 hours to sterilize for later use.

[0024] 4. Use 1% agarose (Ultrapure agarose, Invitrogen, Carlsbad, CA) to coat the bottom of the culture dish, and then inoculate PC12 cell suspension, for example, inoculate 2×10 cells per well in a six-well plate. 4 cells / mL.

[0025] 5. Put the sterilized degreased fibers in a petri dish and culture them for 5 days. During the culture period, replace the culture medium every 2-3 days. Finally, collect the cell balls on the fibers. The diameter of the ce...

Embodiment 2

[0028] Basically the same as in Example 1, the difference is that the anti-adsorption cell material used is gelatin, and the treatment conditions of ammonia plasma are: treatment time is 10s; frequency is 10kHz; pressure is 10 Pa; ammonia gas flow rate is 50 mL / min, high temperature and high pressure sterilization is used after the surface treatment of defatted fiber is completed; the present invention cultures adipose-derived mesenchymal stem cells for 9 days, and the number ranges from 2×10 4 Cells / mL expanded to 7×10 5 cells / mL, the expression levels of genes Oct 4, Sox 2 and Nanog, which represent the characteristics of stem cells, also increased significantly.

Embodiment 3

[0030] Basically the same as Example 1, the difference is that the treatment conditions of ammonia plasma are: treatment time is 35s; frequency is 40kHz; pressure is 5 pa; ammonia gas flow rate is 100 mL / min, after the surface treatment of degreased fiber is completed, high temperature Autoclave sterilization; after the surface treatment of defatted fibers is completed, soak and sterilize with iodine solution, and the present invention cultivates induced pluripotent stem cells for 15 days, and the number ranges from 2×10 4 Cells / mL expanded to 3×10 6 cells / mL, the expression levels of genes Oct 4, Sox 2 and Nanog, which represent the characteristics of stem cells, also increased significantly.

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