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Method for preparing human adipose-derived stem cell membrane based on warm dish

An adipose stem cell and cell membrane technology, which is applied to the field of preparing human adipose stem cell membranes in a temperature-sensitive culture dish, can solve the problems of reducing preparation efficiency, increasing pollution risks, and taking a long time, so as to improve preparation efficiency and avoid cell adhesion failure. Uniform, short processing time results

Pending Publication Date: 2020-10-30
SHANGHAI ICELL BIOTECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The disadvantages of the current method for preparing stem cell membranes using a petri dish are as follows: First, the petri dish needs to be coated, and the specific method is to use reagents with strong adhesion functions, such as serum, gelatin, cohesin, etc. For coating, the coating time is long, which reduces the preparation efficiency and increases the risk of contamination; second, the cells need to be resuscitated and re-cultivated, which takes a long time; Separation speed and integrity need further improvement

Method used

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  • Method for preparing human adipose-derived stem cell membrane based on warm dish
  • Method for preparing human adipose-derived stem cell membrane based on warm dish
  • Method for preparing human adipose-derived stem cell membrane based on warm dish

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] In this example, the complete medium consisted of Gibco DMEM / F-12 basal medium, 8% AventaCELL-Helios serum replacement, and 8 ng / ml EGF.

[0084] The method for preparing human adipose stem cell sheet based on temperature-sensitive culture dish includes the following steps:

[0085] Step a. Isolation of adipose stem cells:

[0086] 1) Remove the upper layer of oil and the lower bloody swelling fluid of the extracted human adipose tissue, and mix the adipose tissue with normal saline (such as figure 1 shown), the volume ratio of fat to normal saline is 1:1, centrifuged (the adipose tissue after centrifugation is as figure 2 shown), remove the upper layer of fat and the bloody liquid at the bottom of the fat after centrifugation;

[0087] 2) Repeat the above steps 3 times to obtain middle layer pure adipose tissue;

[0088] 3) Add collagenase solution for injection prepared with physiological saline in proportion by volume, and after mixing, put it into a 37°C shaking...

Embodiment 2

[0115] In this example, the complete medium consisted of low glucose DMEM basal medium, 10% AventaCELL-Helios serum replacement, and 10 ng / ml EGF.

[0116] The method for preparing human adipose stem cell sheet based on temperature-sensitive culture plate includes the following steps:

[0117] Step a. Isolation of adipose stem cells:

[0118] 1) Remove the upper layer of oil and the bloody swelling liquid in the lower layer of the extracted human adipose tissue, mix the fat with physiological saline, and the volume ratio of the fat to the physiological saline is 1:1, centrifuge, and remove the upper layer of fat and the bloody liquid at the bottom of the centrifuged fat;

[0119] 2) Repeat the above steps 3 times to obtain middle layer pure adipose tissue;

[0120] 3) Add collagenase solution for injection prepared with physiological saline in proportion by volume, and after mixing, put it into a 37°C shaking water bath to digest and decompose;

[0121] 4) Add physiological ...

Embodiment 3

[0148] In this example, the complete medium consisted of α-MEM basal medium, 5% AventaCELL-Helios serum replacement, and 5 ng / ml EGF.

[0149] The method for preparing human adipose stem cell sheet based on temperature-sensitive culture dish includes the following steps:

[0150] Step a. Preparation of adipose stem cell sheet

[0151] 1) Take a temperature-sensitive petri dish and add a complete medium to make the medium completely cover the temperature-sensitive petri dish, which can be placed in an incubator or an ultra-clean workbench at room temperature;

[0152] 2) Take out the cryopreserved adipose stem cells from liquid nitrogen for rapid recovery. The cryopreservation solution is a mixture of DMSO and human serum albumin injection. The volume ratio of DMSO and human serum albumin injection is 5:95; according to 1×107 Cells / ml cells were directly inoculated into temperature-sensitive culture dishes or temperature-sensitive culture plates; cultured in a 37°C, 5% CO2 inc...

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Abstract

The invention discloses a method for preparing a human adipose-derived stem cell membrane based on a warm dish, which comprises the following steps: (1) taking a temperature-sensitive culture dish ora temperature-sensitive culture plate, adding an infiltration liquid, soaking, and sucking out the infiltration liquid; (2) inoculating cryopreserved adipose-derived stem cells or cultured adipose-derived stem cells into a temperature-sensitive culture dish or a temperature-sensitive culture plate, and culturing to prepare an adipose-derived stem cell membrane; (3) obtaining the adipose-derived stem cell membrane; and (4) re-culturing the adipose-derived stem cell membrane. According to the method, a warm dish or a warm dish plate does not need to be coated, cryopreserved cells can be adoptedfor direct inoculation, the separation speed of a cell membrane sheet and the warm dish is high, the integrity is high, and therefore steps and time can be saved, and the preparation efficiency can beimproved.

Description

technical field [0001] The invention belongs to the technical field of stem cell regeneration medicine, and relates to a method for preparing human-derived adipose stem cell membranes based on a warm dish, in particular to a method for preparing human adipose stem cell membranes by directly attaching to a temperature-sensitive culture dish that is damaged and affected. Background technique [0002] Stem cells are a kind of primitive undifferentiated cells with multi-directional differentiation potential and self-replication ability, and are the primitive cells that form various tissues and organs of mammalian tissues. It has extremely high application value in many fields such as repairing the body, organ transplantation, medical beauty and so on. Stem cell research has become a major hotspot in the field of life sciences. [0003] Human adipose stem cells are mostly derived from adipose tissue obtained through cosmetic liposuction. After chemical enzymatic or mechanical se...

Claims

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Application Information

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IPC IPC(8): C12N5/0775A61L27/38A61L27/50
CPCC12N5/0667A61L27/3834A61L27/3895A61L27/50A61L2300/412
Inventor 王淑玲王莉李宁袁惟芯邵小燕
Owner SHANGHAI ICELL BIOTECH
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