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Clinical application-grade adipose-derived stem cells and preparation method thereof

An adipose stem cell, clinical application technology, applied in cell dissociation methods, biochemical equipment and methods, applications, etc., can solve the problems of low cell survival rate, low yield of nucleated cells, long separation and preparation time, etc. short time effect

Active Publication Date: 2020-03-17
SHANGHAI ICELL BIOTECH +4
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, the currently known separation and preparation methods for adipose stem cells have the following disadvantages: 1. The yield of nucleated cells is low; 2. The time for separation and preparation is long; 3. The cell survival rate is low
In addition, the current traditional passage technology limits the application of adipose-derived stem cells to less than 6 generations, which greatly limits the application of adipose-derived stem cells.

Method used

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  • Clinical application-grade adipose-derived stem cells and preparation method thereof
  • Clinical application-grade adipose-derived stem cells and preparation method thereof
  • Clinical application-grade adipose-derived stem cells and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1 Preparation of Adipose Stem Cells

[0071] 1. Preparation of collection bottles

[0072] A fat sample preservation solution was prepared, and the components of the preservation solution included: 40 ml of DMEM, 0.8 ml of penicillin and streptomycin (containing 10,000 IU / ml of penicillin and 10,000 μg / ml of streptomycin; Gibco). Add the prepared preservation solution into the collection bottle, marked as A. Add the same 40ml volume of normal saline to the collection bottle, labeled B.

[0073] 2. Screening of fat samples

[0074] The requirements for fat samples are as follows:

[0075] 1. Hepatitis B, hepatitis C, syphilis, AIDS, CMV, EBV, HTLV virus tests are negative,

[0076] Its detection method is a routine detection method in the art.

[0077] 2. The donor is in good physical condition, without genetic family history, malignant tumor, autoimmune disease, acute or chronic infectious disease, congenital disease, blood system disease or other diseases ...

Embodiment 2

[0103] Embodiment 2 Digestive fluid of the present invention different component distribution ratio and culture effect (weight percentage)

[0104]

[0105] Table 1: The proportion and effect data of the components used in the digestive juice

Embodiment 3

[0106] Note: The unit of cell number in Table 2 is ×10 5 NC / mL Fat Example 3, Detection of Adipose Stem Cells

[0107] 1. Observation of cell morphology

[0108] Adherent cells can usually be seen separating out from the adipose stem cells on the third day, and the cells can reach 80% confluence on the 7th to 10th day. After subculture, the cells were evenly distributed and grew into a spindle. Such as figure 1 shown.

[0109] 2. Detection of nucleated cells per milliliter of adipose tissue

[0110] The number of nucleated cells obtained from the adipose tissue preserved in the preservation solution was: 5.12±0.34×10 5 NC;

[0111] The number of nucleated cells obtained from adipose tissue preserved in normal saline was: 3.24±0.58×10 5 N.C.

[0112] 3. Amplification times of adipose-derived stem cells

[0113] The cells after each digestion were counted, and the amplification factor of each generation was calculated. Such as figure 2 . The amplification factor of ...

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Abstract

The invention discloses a clinical application-grade adipose-derived stem cells and a preparation method thereof. The preparation method comprises the following steps: carrying out primary culture onseparated adipose-derived stem cells, digesting, subculturing and screening out cells with cell clone groups, and carrying out subculturing inoculation. The adipose-derived stem cells prepared by themethod disclosed by the invention are high in nucleated cell yield, short in separation time and high in cell survival rate, and the passage can reach more than 10 generations.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a method for separating and preparing stem cells. Background technique [0002] Adipose tissue is abundant in the human body and easy to obtain. A large number of adipose stem cells obtained through liposuction not only have multi-directional differentiation potential in vivo and in vitro, but also can act like adipocytes, chondrocytes, muscle cells, and osteoblasts under the action of different inducing factors. Cells, nerve cells, glial cells and islet cells differentiate, and can secrete a variety of pro-angiogenic factors and anti-apoptotic factors. Human adipose stem cells come from a wide range of sources, and can be obtained from any human body through liposuction or fat resection. It is safe and painless, and it is stable in vitro culture, fast in expansion, and not easy to age. [0003] However, the currently known method for separating and preparing adi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775A01N1/02
CPCC12N5/0667A01N1/0221C12N2501/91C12N2509/00
Inventor 袁惟芯张钰邵小燕王春慧赵雅宁戴果鲜张光辉严小敏
Owner SHANGHAI ICELL BIOTECH
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