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Method for preparing autologous hematopoietic stem cells, kit, the stem cells and application

A technology of hematopoietic stem cells and somatic cells, applied in the field of biomedicine, can solve the problems of low safety, less available quantity, insufficient sources of hematopoietic stem cells, etc., and achieve the effect of production speed and safety advantages.

Active Publication Date: 2015-03-18
深圳百年干细胞技术研究院有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

This patented technology allows individuals with different types of blood clotting disorders (hemophilia) from their own bone marrow to produce specific type of pluripocytes called platelets that help heal damaged tissues quickly without causing rejection symptoms after transplant surgery. These new plasma units are then given back into patients' bodies where they stimulate natural growth processes within them faster than traditional methods like chemotherapy.

Problems solved by technology

This patents discusses methods for producing large amounts of useful quantities of both natural ones like those found within each individual' s own body but there may still remain some small numbers of these populations due to factors including ethnicity issues associated with living bodies. Additionally, traditional techniques involving collecting them often result in low yields of desired products because many individuals cannot access all parts needed during this process.

Method used

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  • Method for preparing autologous hematopoietic stem cells, kit, the stem cells and application
  • Method for preparing autologous hematopoietic stem cells, kit, the stem cells and application
  • Method for preparing autologous hematopoietic stem cells, kit, the stem cells and application

Examples

Experimental program
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Effect test

Embodiment 1

[0059] The preparation of embodiment 1 kit

[0060] It is operated in a safe operation bench with a cleanliness level of 10-100, and is prepared under a low temperature of 4-10 degrees. In 500ml of RPMI640 culture medium, add: 10μM Y-27632 (C14H21N3O 2HCl); 10ng / mL stem cell factor; 10ng / mL interleukin 3; 10ng / mL interleukin 6; 10ng / mL Interleukin 11; 10 ng / mL macrophage colony-stimulating factor (M-CSF); 10 ng / mL granulocyte colony-stimulating factor (G-CSF); 10 μg / mL fucoidan; 10 μg / mL sulfuric acid Dextran; 10 nM AMD3100 (1,1′-[1,4-phenylenebis(methylene)]-bis-1,4,8,11-tetraazacyclotetradecane); 10 μM M-Malendronate sodium trihydrate; Pamidronate disodium at 10 μM.

[0061] Thoroughly mix and dissolve, then filter and sterilize through a filter with a pore size of 0.22 microns, and prepare cell culture fluid.

[0062] The cell culture solution is placed in the kit, so as to prepare the kit for producing protein-induced autologous hematopoietic stem cells.

Embodiment 2

[0063] Example 2 Preparation of protein-induced autologous hematopoietic stem cells (M-PiHPS) in mice

[0064] Select 3-5 month-old Balb / C male mice (purchased from the Chinese Academy of Medical Sciences School of Radiology (Tianjin), animal qualification certificate number: SCXK (Jin) 2010-0002), collect all their blood, and use conventional Ficoll centrifugation The technology centrifuges and separates blood to obtain blood mononuclear cells. The obtained mononuclear cells were divided into 5×10 6 Individual / ml density, inoculate in the cell culture dish, and add the cell culture solution prepared in embodiment 1 wherein, at 37 ℃ and 5% CO 2 Protein-induced autologous hematopoietic stem cells were obtained by culturing them in an incubator for 5 days. Gently pipette the cultured cells to suspend the cells completely, then collect the cells in a centrifuge tube, perform the centrifugation-saline suspension cell washing operation, repeat 3 times, suspend protein-induced aut...

Embodiment 3

[0065] Example 3 Preparation of human protein-induced autologous hematopoietic stem cells (M-PiHPS)

[0066] Collect 50ml of peripheral blood from the human body, and use conventional Ficoll centrifugal separation technology to centrifuge and separate the blood to obtain blood mononuclear cells. The obtained mononuclear cells were divided into 5×10 6 Individual / ml density, inoculate in the cell culture dish, and add the cell culture solution prepared in embodiment 1 wherein, at 37 ℃ and 5% CO 2 Protein-induced autologous hematopoietic stem cells were obtained by culturing them in an incubator for 5 days. Gently pipette the cultured cells to suspend the cells completely, then collect the cells in a centrifuge tube, perform the centrifugation-saline suspension cell washing operation, repeat 3 times, suspend protein-induced autologous hematopoietic stem cells with normal saline, and obtain protein-induced autologous hematopoietic stem cells Autologous hematopoietic stem cells. ...

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Abstract

The invention provides a method for preparing autologous hematopoietic stem cells. The method includes following steps of cultivating somatic cells in a cell culture fluid for 3-6 days and collecting the generated autologous hematopoietic stem cells. The cell culture fluid is an RPMI1640 culture fluid containing: 1-100 [mu]M of Y-27632 (C14H21N3O.2HCl), 1-100 ng/ml of stem cell factors, 1-100 ng/ml of interleukin 3, 1-100 ng/ml of interleukin 6, 1-100 ng/ml of interleukin 11, 1-100 ng/ml of a macrophage colony stimulating factor (M-CSF), 1-100 ng/ml of a granulocyte colony-stimulating factor (G-CSF), 1-100 [mu]g/ml of fucoidin, 1-100 [mu]g/ml of dextran sulfate, 1-100 nM of AMD3100 (1,1'-[1,4-phenylenebis(methylene)]-bis-1,4,8,11-tetrazacyclotetradecane), 1-100 [mu]M of M-alendronate sodium (Malendronate sodium trihydrate), and 1-100 [mu]M of disodium pamidronate. The invention provides a method for preparing the autologous hematopoietic stem cells, a kit, the stem cells and an application. In addition, the method is advantaged in yield, purity, and producing speed of the production of the protein-induced autologous hematopoietic stem cells.

Description

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Claims

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Application Information

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Owner 深圳百年干细胞技术研究院有限公司
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