Kit for in-vitro separation of karyocytes and removal of erythrocytes and use method thereof

A technology of nucleated cells and kits, which is applied in the field of separating nucleated cells in vitro and can remove red blood cells and kits, and can solve the problems of inability to lyse red blood cells, low stem cell viability and recovery rate, etc.

Inactive Publication Date: 2012-10-24
BEIJING TSCELL BIOTECH
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  • Abstract
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AI Technical Summary

Problems solved by technology

[0003] The method for separating and purifying stem cells from bone marrow, umbilical cord blood and peripheral blood in the prior art has disadvantages such as low stem cell viability and recovery rate, and cannot lyse red blood cells therein
[0004] In our experiments, we believe that the use of 6% hydroxyethyl starch combined with Ficoll for the separation of monocytes in blood improves the recovery rate of monocytes, but it is still mixed with red blood cells.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Preparation method of the cell treatment kit:

[0037] Preparation of precipitation solution: Dissolve 2.40 g of hydroxyethyl starch in 40 ml of normal saline with a concentration of 6%, put it in a water bath at 37°C, and start counting for 8 hours from the temperature of the water bath at 37°C. After 8 hours in the water bath at 37°C, remove from the water bath Take it out, store at room temperature for half an hour.

[0038] Preparation of the purified solution: 60ml of HISTOPAQUE1077 solution (polysucrose sodium diatrizoate), and counted for 8 hours since the temperature of the refrigerator was stabilized at 4°C. After 8 hours at 4°C, take it out of the refrigerator at 4°C, and store it at room temperature for half an hour.

[0039] Preparation of the suspension: Measure 20ml of normal saline with a graduated cylinder and save it.

[0040] Preparation of erythrocyte lysate: 3.735g of ammonium chloride, 1.3g of tris, dissolved in water and diluted to 500ml. Steril...

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PUM

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Abstract

The invention discloses a kit for in-vitro separation of karyocytes and removal of erythrocytes and a use method thereof. The kit comprises a precipitation solution, a purification solution, a suspension and an erythrocyte lysis solution, wherein the precipitation solution, the purification solution, the suspension and the erythrocyte lysis solution are packaged separately. The precipitation solution is composed of normal saline and hydroxyethyl starch dissolved in the normal saline and has a weight volume concentration of 6%. The purification solution is a mixture of polysucrose and sodium diatrizoate. The suspension is normal saline. The erythrocyte lysis solution comprises ammonium chloride having a concentration of 7.47g / L and trihydroxymethylaminomethane having a concentration of 2.6g / L. The invention also discloses a use method of the kit.

Description

technical field [0001] The invention relates to a method and kit for separating and purifying nucleated cells in bone marrow, umbilical cord blood or peripheral blood in vitro and capable of removing red blood cells. Background technique [0002] Stem cells are primitive cells with the potential of self-replication and multi-directional differentiation. They are the origin cells of the body and the primitive cells that form various tissues and organs of the human body. Stem cells can be isolated and purified from bone marrow, umbilical cord blood, and peripheral blood. The commonly used isolation methods for mononuclear cells are as follows: 6% hydroxyethyl starch sedimentation method, methylcellulose sedimentation method, Ficoll separation method, Percoll separation method method, 3% gelatin separation method, ammonium chloride lysed red blood cell method, direct centrifugation method, etc. The commonly used isolation method of monocytes is Ficoll or Percoll direct separat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0786
Inventor 迟明国蔡云松
Owner BEIJING TSCELL BIOTECH
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