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Kit and method for separating mononuclear cells from umbilical cord blood

A kit and the technology of umbilical cord blood, which are applied in the field of separation of mononuclear cells, can solve the problems of unsatisfactory separation effect and achieve good effect, high efficiency and good application prospect

Inactive Publication Date: 2016-07-13
SICHUAN NEO LIFE STEM CELL BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, density gradient centrifugation is the most commonly used method because of its simplicity and effectiveness. At present, there are also many commercial mononuclear cell isolation kits, but the separation effect is still not satisfactory and needs further improvement.

Method used

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  • Kit and method for separating mononuclear cells from umbilical cord blood
  • Kit and method for separating mononuclear cells from umbilical cord blood
  • Kit and method for separating mononuclear cells from umbilical cord blood

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Embodiment 1 Kit of the present invention and separation method

[0045] 1. Kit composition

[0046] Diluent: 0.9% sodium chloride solution by mass percentage.

[0047] Precipitating agent: 6% hydroxyethyl starch solution by mass percentage.

[0048] Separation liquid: a suspension of silica gel particles treated with polyvinylpyrrolidone with a density of 1.078 g / ml.

[0049] Resuspension: 20% human AB plasma.

[0050] 2. Separation method

[0051] 1. Collect human cord blood in a sterile environment and store it in anticoagulant blood bags. In the ultra-clean bench, divide the collected human cord blood into 50ml centrifuge tubes, and put 20ml cord blood into each tube;

[0052] 2. Pour an equal volume of diluent into the centrifuge tube containing 20ml of umbilical cord blood for dilution, then add 10ml of precipitating agent, gently invert and mix, and let stand for 30-50 minutes;

[0053] 3. Place the supernatant obtained from the above sedimentation into a ne...

Embodiment 2

[0061] Embodiment 2 Kit of the present invention and separation method

[0062] 1. Kit composition

[0063] Diluent: diluted with 0.1% sodium chloride solution by mass percentage.

[0064] Precipitating agent: 2.4% hydroxyethyl starch solution in mass percentage.

[0065] Separation liquid: a suspension of silica gel particles treated with polyvinylpyrrolidone with a density of 1.072 g / ml.

[0066] Resuspension: 10% human AB plasma.

[0067] 2. Separation method

[0068] 1. Collect human cord blood in a sterile environment and store it in anticoagulant blood bags. In the ultra-clean bench, divide the collected human cord blood into 50ml centrifuge tubes, and put 20ml cord blood into each tube;

[0069] 2. Inject an equal volume of diluent into the centrifuge tube containing 20ml of umbilical cord blood to dilute, then add 10ml of precipitant, gently invert and mix, and centrifuge for 5-10min under 50g centrifugal force;

[0070] 3. Place the supernatant obtained from the ...

Embodiment 3

[0078] Embodiment 3 Kit of the present invention and separation method

[0079] 1. Kit composition

[0080] Diluent: the mass percentage is diluted with 5% sodium chloride solution.

[0081] Precipitating agent: 12% hydroxyethyl starch solution by mass percentage.

[0082] Separation liquid: a suspension of silica gel particles treated with polyvinylpyrrolidone with a density of 1.086 g / ml.

[0083] Resuspension: 30% human AB plasma.

[0084] 2. Separation method

[0085] 1. Collect human cord blood in a sterile environment and store it in anticoagulant blood bags. In the ultra-clean bench, divide the collected human cord blood into 50ml centrifuge tubes, and put 20ml cord blood into each tube;

[0086] 2. Pour an equal volume of diluent into the centrifuge tube containing 20ml of umbilical cord blood for dilution, then add 10ml of precipitating agent, gently invert and mix, and let stand for 30-50 minutes;

[0087] 3. Place the supernatant obtained from the above sedimen...

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Abstract

The present invention discloses a kit and a method for separating mononuclear cells from umbilical cord blood. The kit for separating the mononuclear cells from the umbilical cord blood comprises the following components: a dilution solution which is a sodium chloride solution with the mass percentage of 0.1% to 5% or a phosphate buffer with pH of 7 to 7.5; a precipitating agent which is a hydroxyethyl starch solution with the mass percentage of 2.4%-12%, a hydroxymethyl starch solution with the mass percentage of 2.4%-12% or a methyl cellulose solution with the mass percentage of 2.4%-12%; a resuspension solution which is an albumin solution with the concentration of 5 to 10% or human AB blood plasma with the concentration of 10-30%; and a separation liquid which is a silica gel particle suspension treated by polyvinylpyrrolidone, and the separation liquid has a density of 1.072-1.086g / ml. The kit and the method can effectively separate and save the mononuclear cells, and have good application prospect. The effect is better than that of commercially available kits.

Description

technical field [0001] The invention relates to a kit and method for isolating mononuclear cells from umbilical cord blood. Background technique [0002] Umbilical cord blood (Umbilical Cord Blood Stem Cells) is the blood remaining in the ligated umbilical cord and placenta after fetal delivery. It contains all the components of normal blood, such as red blood cells, white blood cells, platelets and plasma, as well as umbilical cord blood stem cells (Umbilical Cord Blood Stem Cells), endothelial precursor cells, etc. . [0003] The cell population with a single nucleus in umbilical cord blood mainly includes stem / progenitor cells represented by hematopoietic stem cells, mesenchymal stem cells, and vascular endothelial progenitor cells, as well as lymphocytes and monocytes. Among them, the monocytes and lymphocytes in the white blood cells are treated in vitro and then reinfused into the patient's body. They have broad-spectrum anti-tumor effects and can be used for the trea...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/078
Inventor 李文娴杨超毛松钟立武张博陈强
Owner SICHUAN NEO LIFE STEM CELL BIOTECH
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