Culture medium and method for architectonic stem cell

An adult stem cell and culture method technology, applied in the field of culture medium of adult stem cells, can solve the problems affecting self-renewal ability and differentiation potential, etc.

Inactive Publication Date: 2011-07-20
SHANGHAI JIAOTONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, whether BIO has an effect on the Wnt signaling pathway of human HS / PCs, and further affects its self-renewal ability and differentiation potential, has not yet been reported.

Method used

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  • Culture medium and method for architectonic stem cell
  • Culture medium and method for architectonic stem cell
  • Culture medium and method for architectonic stem cell

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0124] The synthesis of embodiment 1 BIO, MeBIO

[0125] Reagents and experimental conditions: (a) chloral hydrate, Na2SO4, NH2OH·HCl, H2O, HCl; (b) H2SO4, 60-80°C; (c) 3-indole acetate, p-toluenesulfonic acid, EtOH, 70°C (d) NH2OH·HCl, KOH, 30% EtOH, reflux; (e) CH3I, NaH, DMF; (f) 3-indole acetate, Na2CO3, MeOH, 25°C; (g) NH2OH·HCl , Py, 120°C

[0126] Synthesis of N-(oximinoacetyl)m-bromoaniline (2)

[0127] Chloral hydrate (3.73g, 22.55mmol) and anhydrous sodium sulfate (25.52g, 179.72mmol) were added into water (56mL), stirred at 35°C until dissolved. Separately take m-bromoaniline (1, 3.50g, 20.35mmol), add water (14.5mL), concentrated hydrochloric acid (2.2mL) and hydroxylamine hydrochloride (4.8g, 20mL) aqueous solution respectively, shake this mixture up to hydrate as above In chloral, stir well, react at 80-90°C for 2h, cool to 50°C, filter with suction, wash the filter cake with water to obtain white solid 2 (4.566g, yield 92.3%), mp151-153°C, 1HNMR (300Hz, DMSO...

Embodiment 2BI

[0141] The preparation of embodiment 2BIO, MeBIO solution

[0142] BIO and MeBIO powders were dissolved in DMSO to prepare a 10 mM stock solution and stored at -20°C. Freeze and thaw before use, then dilute with medium (DMEM / F-12) 100 times (100 μM), 1000 times (10 μM), 10000 times (1 μM), and finally add 20 μL to the cultured cells (final volume 1 mL) , for final drug concentrations were 2 μM, 0.2 μM and 0.02 μM, respectively.

Embodiment 3

[0143] Example 3 The acquisition of umbilical cord blood mononuclear cells (mono-nuclear cells, MNC) and CD34 + Cell separation and purification

[0144] Umbilical cord blood samples were anticoagulated with sodium citrate, and Ficoll separation solution was used to separate cord blood to obtain cord blood mononuclear cells (Xu Wenrong, Hu Jiabo, Gu Keliang, et al. Experimental study on the density separation of cord blood hematopoietic stem / progenitor cells[J] . Journal of Clinical Laboratory. 2001, 19(2): 73-76). Press 300μL / 10 8 Add the amount of cells to Buffer 3, and resuspend the cells before use. Buffer3: PBS / 0.5%BSA / 2mM EDTA (magnetic sorting buffer)

[0145] CD34 + Cell sorting steps:

[0146] (1) Immunomagnetic bead labeling

[0147] Add FcR-blocking Reagent (100μL / 108 cells), mix gently, then add anti-CD34 + Magnetic beads (100μL / 10 8 cells), mix gently and thoroughly, incubate at 4°C for 30 minutes, take out every 10 minutes during this period, and shake ge...

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Abstract

The invention discloses a medium of adult stem cell, which is characterized by the following: comprising GSK-3 depressant and early stage hematopoiesis factor; improving external augment ability of adult stem cell; maintaining self regenerate ability and differentiation potency. This invention also relates to appliance of the medium in culturing adult stem cell of animal and culturing method of adult stem cell. This invention possesses the advantages of simple operation, lower cost and easy to control.

Description

technical field [0001] The invention belongs to the field of bioengineering, and in particular relates to a culture medium and a culture method of adult stem cells. Background technique [0002] In recent years, hematopoietic stem cell transplantation (Haematopoietic stem cell transplantation, HSCT) has become an important means of treatment of hematological malignancies, solid tumors, autoimmune diseases and certain gene defect diseases, such as leukemia, systemic lupus erythematosus, aplastic Anemia, AIDS, etc. Although allogeneic transplantation has relatively good curative effect and low recurrence rate, there are certain restrictions on the source of donors, and it is prone to severe graft-versus-host disease and high mortality. Although umbilical cord blood stem cell transplantation has the advantages of abundant sources, weak antigenicity, and few complications after transplantation, it also has the disadvantage of insufficient quantity, which limits adult transplant...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/071C12N5/074
Inventor 韩伟姜俊芬毛振民张爱英
Owner SHANGHAI JIAOTONG UNIV
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