A culture system and application thereof for expansion of umbilical cord blood hematopoietic stem cells

A technology of hematopoietic stem cells and culture system, which is applied to blood/immune system cells, animal cells, vertebrate cells, etc., can solve the problems of limited number of cells and limited wide application, and achieves reduced cell structure, high separation efficiency, and operation saving. effect of steps

Active Publication Date: 2020-05-08
SHUNHAO CELL BIOTECHNOLOGY (TIANJIN) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the limited number of cells contained in an umbilical cord blood, this greatly limits its wide application in clinical practice.

Method used

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  • A culture system and application thereof for expansion of umbilical cord blood hematopoietic stem cells
  • A culture system and application thereof for expansion of umbilical cord blood hematopoietic stem cells
  • A culture system and application thereof for expansion of umbilical cord blood hematopoietic stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] (1) 50-80ml of collected umbilical cord blood is slowly added to a 50ml centrifuge tube containing lymphatic separation solution at a ratio of 2:1, centrifuged at 22°C, 1500-3000rpm, centrifuged for 10-20min, the upper layer is platelet-rich plasma , Heparin is used as anticoagulant in the blood collection bag, and the upper serum accounts for 45-50% of the total volume. Slowly suck out the cells of the third layer of albuginea with a pipette and set aside.

[0038] (2) The platelet-rich plasma is stored in an ultra-low temperature refrigerator at -80°C, and after 12 hours, it is quickly taken out and placed in a 37°C water bath for resuscitation. Repeat this 5 times. Centrifuge at 25°C, 3500rpm, 20min, take the supernatant, and filter with 0.22μm. Store in a refrigerator at 4°C for later use.

[0039] (3) Use the Peprotech ELISA kit to detect the platelet lysate prepared in step (2), which is rich in cytokines TGF-β, IGF, PDGF, VEGF, FGF, and analyze the cytokine content ...

example 2

[0054] (1) 50-80ml of collected umbilical cord blood is slowly added to a 50ml centrifuge tube containing lymphatic separation solution at a ratio of 2:1, centrifuged at 22°C, 1500-3000rpm, centrifuged for 10-20min, the upper layer is platelet-rich plasma , Heparin is used as anticoagulant in the blood collection bag, and the upper serum accounts for 45-50% of the total volume. Slowly suck out the cells of the third layer of albuginea with a pipette and set aside.

[0055] (2) The platelet-rich plasma is stored in an ultra-low temperature refrigerator at -80°C, and after 12 hours, it is quickly taken out and placed in a 37°C water bath for resuscitation, and this is repeated 5 times. Centrifuge at 25°C, 3500rpm, 20min, take the supernatant, and filter with 0.22μm. Keep in refrigerator at 4°C for later use.

[0056] (3) After resuspending the albuginea cells prepared in step (1) with physiological saline, centrifuge, 1200rpm, 10min, use MACS-CD34isolation kit for the cell pellet a...

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Abstract

The present invention discloses a culture system used for umbilical cord blood hematopoiesis stem cell amplification and application thereof, the culture system includes an AIM-V culture medium, platelet lysate PL, stem cell factor SCF, human FMS-like tyrosine kinase 3 ligand Flt-3L , G-CSF and IL-3, and angiopoietin-likeprotein 3 (ANGPTL3), small molecule compound StemRegenin (SR1) and pyrimidine-indole molecule UM171 are also added. By adding of the appropriate concentration of ANGPTL3, SR1 and UM171 for combination, the number of hematopoiesis stem cell population is greatly increased, the culture system is used for umbilical cord blood hematopoiesis stem cell growth and amplification, to be more important, implantation capacity and hematopoietic system reconstruction capacity after hematopoietic stem cells are transplanted into a receptor are significantly improved, and the culture system opens up new use prospects in clinical amplification of umbilical cord blood stem cells.

Description

Technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a culture system for umbilical cord blood hematopoietic stem cell expansion and its application. Background technique [0002] Hematopoietic stem cell transplantation is one of the most effective treatment methods for the treatment of leukemia and lymphoma and other malignant tumors, metabolic diseases, autoimmune diseases, and innate immune deficiencies and other diseases that seriously endanger human health. The number of hematopoietic stem cells (HSC) in the bone marrow is low, and it is difficult to effectively reconstruct hematopoietic reconstitution in patients after transplantation. This has also become a bottleneck in the clinical application of HSC, and about 30-40% of patients cannot find the same HLA type The donor has lost the opportunity for treatment. Umbilical cord blood (UCB) is another important source of hematopoietic stem cells in addition to bone marrow and...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0789
Inventor 张云李相国高畅孙丽娟王冠华
Owner SHUNHAO CELL BIOTECHNOLOGY (TIANJIN) CO LTD
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