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Method of amplifying hematopoietic stem cells of umbilical cord blood in vitro

A technology for in vitro expansion of hematopoietic stem cells, applied in the field of in vitro expansion of umbilical cord blood hematopoietic stem cells, can solve problems such as low expansion efficiency that cannot meet clinical needs, insufficient number of umbilical cord blood hematopoietic stem cell transplants, and inability to expand umbilical cord blood hematopoietic stem cells , to achieve the effect of good application prospects

Inactive Publication Date: 2017-12-08
YUNNAN SHUNXI REGENERATION MEDICAL ENG CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problem of insufficient number of umbilical cord blood hematopoietic stem cell transplantation, and the disadvantages that the umbilical cord blood hematopoietic stem cells cannot be expanded at the current stage, the amplification efficiency is low and the number of cells cannot meet the clinical demand, the present invention provides a method for expanding umbilical cord blood hematopoietic stem cells in vitro Method to obtain a sufficient number of umbilical cord blood hematopoietic stem cells for clinical use

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  • Method of amplifying hematopoietic stem cells of umbilical cord blood in vitro
  • Method of amplifying hematopoietic stem cells of umbilical cord blood in vitro
  • Method of amplifying hematopoietic stem cells of umbilical cord blood in vitro

Examples

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Effect test

Embodiment 1

[0043] A method for expanding umbilical cord blood hematopoietic stem cells in vitro, comprising the steps of:

[0044] Step (1), centrifuge cord blood at 650g for 15 minutes to remove plasma, then add an equal volume of normal saline to the remaining blood cells to dilute the blood cells;

[0045] Step (2), take the centrifuge tube added with Ficoll-diatrizoate solution, and then add the blood cells diluted in step (1) along the wall of the centrifuge tube to make the boundary clear. The volume ratio of sucrose-diatrizoate meglumine solution is 4:3. Centrifuge at 1800 rpm for 30 minutes at 18°C, then take the buffy coat layer and wash it twice with normal saline. The volume of normal saline used each time is white Three times the volume of the membrane layer, cleaned by centrifugation, the centrifugation speed is 1200 rpm, and the centrifugation time is 5 minutes each time to obtain CBMC (cord blood mononuclear cells);

[0046] Wherein, every 100mL polysucrose-diatrizoate so...

Embodiment 2

[0055] A method for expanding umbilical cord blood hematopoietic stem cells in vitro, comprising the steps of:

[0056] Step (1), centrifuge cord blood at 700g for 20 minutes to remove plasma, then add an equal volume of normal saline to the remaining blood cells to dilute the blood cells;

[0057] Step (2), take the centrifuge tube added with Ficoll-diatrizoate solution, and then add the blood cells diluted in step (1) along the wall of the centrifuge tube to make the boundary clear. The volume ratio of sucrose-diatrizoate meglumine solution is 4:3, centrifuge at 2000 rpm for 45 minutes at 20°C, then take the buffy coat layer and wash it with normal saline 2~3 times, the volume of normal saline used each time 4 times the volume of the buffy coat, cleaned by centrifugation, the centrifugation speed is 1500 rpm, each centrifugation time is 8 minutes, and CBMC (cord blood mononuclear cells) are obtained;

[0058] Wherein, every 100mL polysucrose-diatrizoate solution contains po...

Embodiment 3

[0067] A method for expanding umbilical cord blood hematopoietic stem cells in vitro, comprising the steps of:

[0068] Step (1), centrifuge cord blood at 660g for 18 minutes to remove plasma, then add an equal volume of normal saline to the remaining blood cells to dilute the blood cells;

[0069] Step (2), take the centrifuge tube added with Ficoll-diatrizoate solution, and then add the blood cells diluted in step (1) along the wall of the centrifuge tube to make the boundary clear. The volume ratio of sucrose-diatrizoate meglumine solution is 4:3, centrifuge at 1900 rpm for 39 minutes at 19°C, then take the buffy coat layer and wash it twice with normal saline, the volume of normal saline used each time is white 3.5 times the volume of the film layer, cleaned by centrifugation, the centrifugation speed is 1300 rpm, and the centrifugation time is 7 minutes each time to obtain CBMC (cord blood mononuclear cells);

[0070] Wherein, every 100mL polysucrose-diatrizoate solution...

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Abstract

The invention relates to a method of amplifying hematopoietic stem cells of umbilical cord blood in vitro and belongs to the technical field of biomedicines. The method amplifies the hematopoietic stem cells of umbilical cord blood in vitro by means of CBMC separation of a mononuclear cell of umbilical cord blood, CD34 positive cell enrichment and CD34 positive cell expansion, and the amplified hematopoietic stem cells of umbilical cord blood are detected relatively. According to the method provided by the invention, the hematopoietic stem cells of umbilical cord blood can be amplified in a multiple fold manner, and the cells meet the clinical application standard. By expansion in vitro, the quality of implanting the hematopoietic stem cells of umbilical cord blood for clinical use of an adult is met. No treatment methods affecting the cell state are employed, and a condition culture medium of mesenchymal stem cells is used for amplifying the hematopoietic stem cells. Compared with a control group, the method can raise the proportion of the D34 positive hematopoietic stem cells from 0.28% to 13.95%, and has a good application prospect.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a method material for expanding umbilical cord blood hematopoietic stem cells in vitro. Background technique [0002] Bone marrow hematopoietic stem cell transplantation has been used to treat malignant hematopoietic diseases. However, transplantation of hematopoietic stem cells from the bone marrow is painful and carries risks of infection and complications. Moreover, the number of bone marrow hematopoietic stem cells cannot always satisfy all transplant requirements. Compared with bone marrow hematopoietic stem cells, umbilical cord blood hematopoietic stem cells have better cell proliferation and immune tolerance. However, unfortunately, cord blood hematopoietic stem cells also have limitations, that is, the number of cells is low, which can only meet the needs of children for transplantation. According to literature reports, the successful transplantation o...

Claims

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Application Information

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IPC IPC(8): C12N5/0789
CPCC12N5/0647C12N2502/1388
Inventor 王亚君李一佳陈曦闫姗姗
Owner YUNNAN SHUNXI REGENERATION MEDICAL ENG CO LTD
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