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Kit for processing human marrow, cord blood and peripheral blood cells and cell processing method

A technology of peripheral blood stem cells and umbilical cord blood, which is applied in the field of umbilical cord blood, peripheral blood stem cell isolation kits and stem cell isolation, medium stem cells, and human bone marrow, and can solve the problems of unclear marker reaction, low cell activity, and long acquisition time, etc. Achieve the effects of complete stem cell types, small cell fluid volume, and short separation operation time

Inactive Publication Date: 2013-11-20
BEI ZHENG STEM CELLS BIOLOGICAL TECH CO LTD BEIJING
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The purpose of the present invention is to provide a human bone marrow, umbilical cord blood, peripheral blood stem cell separation kit and stem cell separation method with strong operability, high clinical safety and easy clinical promotion, which fundamentally solves the problem of existing cell separation. There are problems in the technology such as high cost, low cell activity, unknown reaction of markers in the human body, the need to mobilize agents to cause pain to patients, long acquisition time, cumbersome and unsuitable for clinical use, etc.

Method used

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  • Kit for processing human marrow, cord blood and peripheral blood cells and cell processing method
  • Kit for processing human marrow, cord blood and peripheral blood cells and cell processing method
  • Kit for processing human marrow, cord blood and peripheral blood cells and cell processing method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1: Isolation of bone marrow

[0027] The composition of the kit is as follows:

[0028] No. 1 solution: diluent: PBS solution is selected, and the preparation process is to dissolve the following reagents with a small amount of water: 4g sodium chloride, 0.1g potassium chloride, 1.445g disodium hydrogen phosphate 12 water, 0.1g potassium dihydrogen phosphate , and then add water to 500ml, that is. The pH was adjusted to 7.2 with 5.6% sodium carbonate before use.

[0029] No. 2 solution: Precipitating agent: 6% hydroxyethyl starch (commercially available).

[0030] No. 3 liquid: Layering liquid: Polysucrose and diatrizoate are prepared into a layering liquid with a specific gravity of 1.075.

[0031] The above-mentioned No. 1 solution was sterilized at 10 pounds for 10 minutes, and its endotoxin content was detected to be ≤0.5 EU / ml, bottled, and stored at 4°C. Liquid No. 3 was sterilized at 10 pounds for 10 minutes, and its endotoxin content was detected to be...

Embodiment 2

[0036] Example 2: Isolation of umbilical cord blood

[0037] The composition of the kit is as follows:

[0038] Liquid No. 1: diluent: 0.9% sodium chloride injection (ie physiological saline) is commercially available.

[0039] No. 2 solution: Precipitating agent: 6% hydroxyethyl starch (commercially available).

[0040] No. 3 liquid: layering liquid: polysucrose and meglumine diatrizoate are prepared into a layering liquid with a specific gravity of 1.076.

[0041] The above-mentioned No. 3 liquid was sterilized at 10 pounds for 10 minutes, and its endotoxin content was detected to be less than or equal to 5 EU / ml, and bottled.

[0042] Use the above kit to separate stem cells: Add 100ml of human cord blood containing sodium citrate anticoagulant solution to reagent No. 1 solution containing 100ml, then add 100ml reagent No. 2 solution, shake well for 3 minutes, and place for 30 Minutes, after stratification, absorb the upper layer of cell fluid, divide into 50ml centrifug...

Embodiment 3

[0045] Embodiment 3: Separation of peripheral blood

[0046] The composition of the kit is as follows:

[0047] Liquid No. 1: diluent: 0.9% sodium chloride injection (ie physiological saline) is commercially available.

[0048] Liquid No. 2: Precipitating agent: 2.5 grams of methylcellulose (imported), added to 500 ml of 4°C normal saline, and shaken well. That is, 0.5% methylcellulose was obtained.

[0049] No. 3 liquid: Layering liquid: Polysucrose and diatrizoate are prepared into a layering liquid with a specific gravity of 1.074.

[0050] The above-mentioned No. 2 solution was sterilized at 10 pounds for 10 minutes, and its endotoxin content was detected to be ≤0.5 EU / ml, bottled, and stored at 4°C. Liquid No. 3 was sterilized at 10 pounds for 10 minutes, and its endotoxin content was detected to be ≤5EU / ml before being bottled.

[0051] Use the above kit to isolate stem cells: add 200ml of human peripheral blood containing sodium citrate anticoagulant to 200ml of rea...

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Abstract

The invention relates to a kit for processing human marrow, cord blood and peripheral blood cells and a cell processing method. The kit radically solves the problems of high manufacture cost, low cellactivity, indefinite reaction of markers in a human body, mobilizing agent injection causing patient pain, long obtainment time, complication, clinical unsuitability, and the like in the prior cell separation technology. The invention has the technical scheme that the kit comprises the following three reagents: (1) a diluent comprising 0.9 percent of sodium chloride injection or PBS solution, (2)a precipitator comprising 6 percent of hydroxyethyl starch or 0.2-1 percent of methyl cellulose and (3) a separating solution which has the density of 1.074-1.076 and is prepared by saccharosan and diatrizoate. The kit is easy to store and transport and convenient and rapid to use and can be produced in industrialization.

Description

technical field [0001] The invention relates to a method for separating stem cells from human bone marrow, umbilical cord blood, etc. in vitro, in particular to a human bone marrow, umbilical cord blood, peripheral blood stem cell separation kit and stem cell separation method, belonging to the field of biomedical technology applications. Background technique [0002] With the continuous development of modern science and technology, the separation technology for human blood cells has also been greatly developed. At present, there are mainly the following processing methods: [0003] 1. Immunomagnetic bead method: the known antibody is coated on the magnetic bead particles, and the magnetic bead particles are mixed with human blood. Antigen-antibody-positive cells adhere to the magnetic beads, pass through a magnetic tube, and the magnetic beads are adsorbed on the tube wall; after other cells that are not bound to the magnetic beads flow away, remove the magnetic properties ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/08
Inventor 唐明淇
Owner BEI ZHENG STEM CELLS BIOLOGICAL TECH CO LTD BEIJING
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