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

Method for separating adipose-derived stem cells from fat and preparation in use

A technology of mesenchymal stem cells and fat, applied in the field of isolation and culture of autologous adipose-derived mesenchymal stem cells, can solve the problems of inapplicable acquisition of a large number of adipose-derived mesenchymal stem cells, low cell yield, and low cell viability

Active Publication Date: 2018-03-30
FIVE DIMENSION BY INCOSC HEALTH MANAGEMENT JIANGSU
View PDF9 Cites 9 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0021] Although the prior art discloses some methods for culturing adipose-derived mesenchymal stem cells such as the above-mentioned ones, the inventors have found that these methods do not seem to be suitable for obtaining a large amount of adipose-derived mesenchymal stem cells from fat. For example, these methods are used in Low cell yield and / or low cell viability when harvesting adipose-derived mesenchymal stem cells from fat

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 separating adipose-derived stem cells from fat and preparation in use
  • Method for separating adipose-derived stem cells from fat and preparation in use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0111] Example 1: Isolation and culture of adipose-derived mesenchymal stem cells

[0112] (1) Put the fat sample mixed with swelling fluid obtained by liposuction into a centrifuge tube, and centrifuge at 800rpm for 3min;

[0113](2) Remove the top layer of fat, and pipette the suspended fat particles and the bottom cell sediment respectively, and set aside;

[0114] (31) Add mixed enzyme (0.05% collagenase type I+0.05% collagenase type II+0.03% neutral protease+0.03% DNase) with volume ratio 1: 1 to the fat granule gained in step (2), at 37 Centrifuge at 100rpm for 30min; after digestion, add an equal volume of complete medium to stop digestion, centrifuge at 1500rpm for 5min, discard the supernatant, and resuspend the cell pellet in PBS;

[0115] (32) Resuspend the bottom cell pellet obtained in step (2) with PBS, add an equal amount of erythrocyte lysate and let it stand for 10 minutes to lyse; after the erythrocytes are fully lysed, centrifuge at 1500 rpm for 5 minutes...

Embodiment 2

[0123] Example 2: Isolation and culture of adipose-derived mesenchymal stem cells

[0124] (1) Put the fat sample mixed with swelling fluid obtained by liposuction into a centrifuge tube, and centrifuge at 500rpm for 5min;

[0125] (2) Remove the top layer of fat, and pipette the suspended fat particles and the bottom cell sediment respectively, and set aside;

[0126] (31) Add mixed enzyme (0.05% collagenase type I+0.05% collagenase type II+0.03% neutral protease+0.03% DNase) with volume ratio 1:1.2 in the fat granule gained in step (2), at 37 Centrifuge at 100rpm for 20min; after digestion, add an equal volume of complete medium to stop digestion, centrifuge at 2000rpm for 3min, discard the supernatant, and resuspend the cell pellet in PBS;

[0127] (32) Resuspend the bottom cell pellet obtained in step (2) with PBS, add an equal amount of erythrocyte lysate and let it stand for 5 minutes to lyse; after the erythrocytes are fully lysed, centrifuge at 2000 rpm for 3 minute...

Embodiment 3

[0134] Example 3: Isolation and culture of adipose-derived mesenchymal stem cells

[0135] (1) Put the fat sample mixed with swelling fluid obtained by liposuction into a centrifuge tube, and centrifuge at 1000rpm for 2min;

[0136] (2) Remove the top layer of fat, and pipette the suspended fat particles and the bottom cell sediment respectively, and set aside;

[0137] (31) Add mixed enzymes (0.05% collagenase type I+0.05% collagenase type II+0.03% neutral protease+0.03% DNase) with a volume ratio of 1:0.8 to the fat granules obtained in step (2), at 37 Centrifuge at 100rpm for 40 minutes; after digestion, add an equal volume of complete medium to stop digestion, centrifuge at 1000 for 8 minutes, discard the supernatant, and resuspend the cell pellet in PBS;

[0138] (32) Resuspend the bottom cell pellet obtained in step (2) with PBS, add an equal amount of erythrocyte lysate and let it stand for 15 minutes to lyse; after the erythrocytes are fully lysed, centrifuge at 100...

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

No PUM Login to View More

Abstract

The invention discloses a method for separating adipose-derived stem cells from fat and a preparation in use. On one hand, the method for separating the adipose-derived stem cells from fat comprises the steps of conducting liposuction to obtain a fat sample doped with inflation liquid to be subjected to centrifugation; removing the uppermost-layer fat, and removing suspension fat particles and bottom-layer cell sediment respectively; adding the fat particles into digestive enzyme mixed liquid, conducting vibration digestion, stopping the digestion, conducting centrifugation, and re-suspendingcell sediment; re-suspending the cell sediment at the bottom layer, adding erythrocyte pyrolysis liquid to conduct pyrolysis, conducting centrifugal, and re-suspending the cell sediment; merging to obtain cell suspension, filtering, and separating out the adipose-derived stem cells serving as primary passage. The invention further relates to a method for separating and cultivating the adipose-derived stem cells. The prepared adipose-derived stem cell preparations are used for separating the digestive enzyme mixed liquid of the adipose-derived stem cells, and are used for cultivating a culturemedium of the adipose-derived stem cells. The method for separating the adipose-derived stem cells from fat has various advantages as shown in the description.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a method for isolating adipose mesenchymal stem cells from fat, in particular to a method for isolating and culturing autologous adipose mesenchymal stem cells. The present invention also relates to the culture medium and related test solution used in the above-mentioned culture of autologous adipose-derived mesenchymal stem cells. When the method of the present invention is used to isolate and culture the adipose-derived mesenchymal stem cells, the excellent technical effect of the present invention can be presented. The obtained autologous adipose-derived mesenchymal stem cells can be used for autologous application. Background technique [0002] Mesenchymal stem cells (MCS) are hot spots of adult stem cells with multi-directional differentiation potential. It is a cell population with bone formation ability and hematopoietic function found in most adult organs. It can be extracted ...

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
Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0775
CPCC12N5/0667C12N2501/105C12N2501/11C12N2501/115C12N2509/00C12N2501/15
Inventor 王芳梅俊
Owner FIVE DIMENSION BY INCOSC HEALTH MANAGEMENT JIANGSU
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