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Culture apparatus used for temperature sensitive cells, preparation method thereof and cell culturing method

A temperature-sensitive, cell culture technology, applied in tissue cell/virus culture devices, biochemical equipment and methods, animal cells, etc., can solve the problems of cell surface proteins, ion channels, cell signal damage, etc., to reduce death , the effect of reducing damage, improving quality and activity

Inactive Publication Date: 2013-12-11
上海瀚正生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to provide a temperature-sensitive cell culture device to solve the problem that the methods for removing cells in tissues in tissue engineering in the prior art are basically the application of biological enzyme digestion or direct cell scraping. Methods Technical issues causing damage to cell surface proteins, ion channels, cell signaling, etc.
[0007] Another object of the present invention is to provide the above-mentioned temperature-sensitive cell culture device preparation method to solve the problem that in the prior art, the methods for removing cells in tissues in tissue engineering are basically using biological enzyme digestion or direct cell scraping. Falling method, the technical problem that these methods cause damage to cell surface proteins, ion channels, cell signals, etc.

Method used

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  • Culture apparatus used for temperature sensitive cells, preparation method thereof and cell culturing method
  • Culture apparatus used for temperature sensitive cells, preparation method thereof and cell culturing method
  • Culture apparatus used for temperature sensitive cells, preparation method thereof and cell culturing method

Examples

Experimental program
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Effect test

Embodiment 1

[0032] Example 1 Preparation of temperature-sensitive polystyrene culture sheet by spin coating method and cultivation of human umbilical cord mesenchymal stem cells

[0033] (1) Synthesis of polystyrene (PS): In a 25ml round-bottom single-necked flask, add 10ml of dry toluene solvent, add 7.5g of styrene and 1.5g of diisopropyl xanthate (DIP) into the flask Stir evenly, add 0.5g AIBN at room temperature, stir until the AIBN is completely dissolved, seal with a rubber stopper, and cycle three times with nitrogen gas-freezing-pumping. React at 75°C for 6 hours, remove toluene by rotary evaporation, add THF to dissolve, and precipitate in petroleum ether to obtain 7.5 g of polystyrene (PS).

[0034] (2) Synthesis of PS-PNIPAAm: Add 1.0 g PS, 1.2 g isopropylacrylamide monomer, and 10 mg AIBN to 6 ml of dry 1.4 dioxane, stir and dissolve at room temperature. Nitrogen-freezing-pumping cycles three times, react at 80°C for 10 h, remove 1.4 dioxane by rotary evaporation, add THF to ...

Embodiment 2

[0039] Example two dental pulp mesenchymal stem cells were cultured in the graft

[0040] (1) In a 10ml round-bottom single-necked flask, add 5ml of dry toluene solvent, add 12g of styrene and 2.4g of diisopropyl xanthate disulfide (DIP) into the flask and stir evenly, add 0.8g of AIBN at room temperature, and stir Until the AIBN is completely dissolved, seal it with a rubber stopper, and cycle three times with nitrogen gas-freezing-pumping. React at 75°C for 5 hours, remove toluene by rotary evaporation, add THF to dissolve, and precipitate in petroleum ether to obtain polystyrene (PS).

[0041] (2) Synthesis of PS-PNIPAAm: Add 3.0 g PS, 6.0 g isopropylacrylamide monomer, and 150 mg AIBN to 10 ml of dry 1.4 dioxane, stir and dissolve at room temperature. Nitrogen-freezing-pumping cycle five times, react at 90°C for 15 hours, remove 1.4 dioxane by rotary evaporation, add THF to dissolve, and precipitate in ether to obtain PS-PNIPAAm block polymer.

[0042] (3) Dissolving the...

Embodiment 3

[0045] Example 3 Growth and detachment of human amniotic mesenchymal stem cells on the surface of the graft

[0046] (1) Synthesis of PS-PNIPAAm block copolymer: Add 5.0g PS, 10g isopropylacrylamide monomer, and 80mg AIBN to 50ml dry 1.4 dioxane, stir and dissolve at room temperature. Nitrogen-freezing-pumping cycles three times, react at 100°C for 12 hours, remove 1.4 dioxane by rotary evaporation, add THF to dissolve, and precipitate in ether to obtain PS-PNIPAAm block polymer.

[0047] (2) Dissolving the prepared PS-PNIPAAm block polymer in tetrahydrofuran. Polystyrene material cut into 5cm 2 Place the polystyrene sheet on the vacuum suction cup of the coating machine, drop the block polymer solution onto the polystyrene sheet with a syringe, and rotate at 8000rpm for 40s to ensure uniform suspension coating. After obtaining the grafted pieces, put them into an oven at 100°C and dry them for 3 hours for later use.

[0048] (3) Sterilization treatment of temperature-sensi...

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Abstract

The invention provides a culture apparatus used for temperature sensitive cells, a preparation method thereof and a cell cultureing method. The culture apparatus used for temperature sensitive cells comprises a polystyrene body; and the polystyrene body is grafted with a polystyrene-poly N-isopropylacrylamide block polymer. Compared with existing technologies, when the culture apparatus used for temperature sensitive cells is used for cell culturing, it is possible to obtain complete cultured cell layers and extracellular matrixes from deposition of the culture cell layers without damage by simple temperature variation, digestion by pancreatic enzyme or dissociation by chemical reagents is avoided, the death of cells is reduced, damages to the extracellular matrixes are lessened, and quality and activity of the cultured cells are improved.

Description

technical field [0001] The invention relates to a cell culture device, a preparation method, and a cell culture method, in particular to a temperature-sensitive cell culture device, a preparation method, and a cell culture method. Background technique [0002] Cell therapy is a treatment method based on functional cells, which can be used as an independent treatment method or combined with conventional surgery, chemical drugs and other treatment methods. Its mechanism of action is: 1) direct action of cells to repair damaged tissues and organs or kill tumor cells; 2) indirect action of cells to secrete related cytokines or bioactive molecules to regulate the proliferation and function of the patient's own cells. Therapeutic cells may be of the patient's own or allogeneic origin; the treatment may be general infusion or transplantation. Cell therapy is widely used in bone marrow transplantation, advanced liver cirrhosis, femoral head necrosis, malignant tumors, myocardial in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12M3/04C12N5/0775C08J7/12C08L25/06C08F112/08C08F293/00
Inventor 齐念民杜明明冯见陈彦田
Owner 上海瀚正生物技术有限公司
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