Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Method for preparing clinical-level adipose-derived stem cells

An adipose stem cell and adipose tissue technology, applied in the field of cell culture technology and biological therapy, can solve the problems of increased risk, low vitality, and non-compliance with clinical application requirements.

Active Publication Date: 2019-04-19
JILIN TUO HUA BIOTECH
View PDF5 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, most of the isolation methods of adipose stem cells are separated by enzymatic digestion, but the number of obtained cells is insufficient, and their viability is low, aging often occurs during long-term passage, and the addition of exogenous serum and factors in the medium does not meet the clinical application requirements. These factors have greatly increased the risk of clinical use

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for preparing clinical-level adipose-derived stem cells
  • Method for preparing clinical-level adipose-derived stem cells
  • Method for preparing clinical-level adipose-derived stem cells

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0024] The invention provides a method for preparing clinical-grade adipose stem cells, the steps are as follows:

[0025] (1) The adipose tissue is washed with double anti-salt water, shredded, and impurities, grease and other substances are removed;

[0026] (2) Repeated injection of chylodious adipose tissue through a syringe to shorten the digestion time and increase the number and vitality of stem cells;

[0027] (3) Digest the adipose tissue with a mixed enzyme of type I collagenase, hyaluronidase, deoxyribonuclease I, and trypsin, filter, wash, and discard the supernatant to obtain adipose stem cells.

[0028] Further, the method of the present invention also includes the following steps:

[0029] (4) Inoculate the adipose-derived stem cells in a-MEM medium for culture, which contains cell nutrient supplements, EGF, bFGF, TGF, HGF, PDGF, and does not contain exogenous serum and other non-conforming clinical-grade culture reagents.

[0030] As a preferred solution, the...

Embodiment 1

[0036] Example 1: Isolation and Preparation of Adipose Stem Cells

[0037] Adipose stem cells were isolated and prepared by the following steps:

[0038] 1) Obtain adipose tissue from cosmetic liposuction fluid or surgical fat mass, and wash the adipose tissue with double-resistant saline solution containing penicillin and streptomycin mixture, wherein the working concentration of penicillin is 100U / ml, and the working concentration of streptomycin is 0.1mg / ml, shredded to remove impurities such as grease;

[0039] 2) Take out 30ml from the adipose tissue obtained in step 1) and add it to a 50ml syringe, and repeatedly inject the syringe for 3 minutes, 30 times per minute, to make the adipose tissue chylolysis; wherein, the duration of the repeated injection of the syringe is 1 minute to 5 minutes, 10-30 times per minute; if the time is too short, the adipose tissue will not be fully emulsified; if the time is too long, the stem cells will be damaged and the recovery effect w...

Embodiment 2

[0043] Embodiment 2: Adipose stem cell bacteria, fungus, endotoxin, mycoplasma detection prepared by the present invention

[0044] The cell growth state of the adipose stem cells prepared by the method of Example 1 was observed under a microscope, subcultured when the degree of confluence was 80%-90%, and samples were taken after subculture to detect bacteria, fungi, endotoxin and mycoplasma. The test results showed that there was no colony growth after cultured on Columbia blood agar and Sabauro agar plate culture, showing that bacteria and fungi were negative; DNA fluorescent staining method showed no particulate mycoplasma after Hoechst3325 staining, showing that mycoplasma was negative; The test for endotoxin was negative; the above tests were all derived from the pharmacopoeia test methods, and the results showed that they were all negative and met the conditions for reinfusion of biological therapy.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
The average diameteraaaaaaaaaa
Login to View More

Abstract

The invention relates to a method for preparing clinical-level adipose-derived stem cells. Specifically, the method includes the steps of firstly, washing adipose tissue with double-resistance salinecontaining mycillin mixed liquid, and cutting off and removing adipose; secondly, conducting chylosis on the adipose tissue obtained in the first step; thirdly, adding the adipose tissue subjected tochylosis in the second step to an a-MEM containing an enzyme mixture to be digested, wherein the enzyme mixture contains I-type collagenase, hyaluronidase, deoxyribonuclease I and trypsin; fourthly, centrifuging digested liquid in the third step, and removing supernate to obtain the adipose-derived stem cells.

Description

technical field [0001] The invention relates to the fields of cell culture technology and biotherapy, in particular to a method for preparing clinical-grade adipose stem cells. Background technique [0002] Adipose derived stem cells (ADSC) are adult mesenchymal stem cells isolated from adipose tissue with self-renewal and multidirectional differentiation potential. It has some common characteristics of stem cells, and has self-renewal, high proliferation ability and differentiation potential to adipocytes, osteoblasts, chondrocytes, and nerve cells under suitable induction conditions in vitro, and has attracted widespread attention. ADSC cells can proliferate stably in vitro and have a low death rate. At the same time, it has the advantages of easy material collection, a large number of stem cells can be obtained from a small amount of tissue, suitable for large-scale culture, and little damage to the body. Moreover, it has a wide range of sources and a large reserve in the...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N5/0775
CPCC12N5/0667C12N2509/10
Inventor 姜丽君李超
Owner JILIN TUO HUA BIOTECH
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com