Method for separating lymphocytes

A separation method and liquid separation technology, applied in the biological field, can solve the problems of cumbersome steps, easy fatigue, time-consuming and labor-intensive, etc., and achieve the effect of reasonable ratio and small uncertainty of raw materials.

Inactive Publication Date: 2016-05-11
北京弘润天源基因生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Moreover, the current gradient centrifugation method has cumbersome steps, strong skills, and relatively high requirements for the operator's experience: gradient centrifugation requires three steps, especially the second step. Density of the cell suspension without mixing the interface, which greatly tests the operator's experience and patience, time-

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] In an environment with local 100-level laminar flow purification conditions, at 20°C, dextran 3g, trehalose 4g, ultra-low viscosity sodium alginate 0.8g, diatrizoate meglumine 1g, polydiallyl 0.15 g of dimethyl ammonium chloride was fully dissolved in sterile water for injection to prepare 100 mL of a physiological solution, and then filtered through a 0.22 μm filter membrane into a sterile, endotoxin-free container.

Embodiment 2

[0027] In an environment with local 100-level laminar flow purification conditions, at 20°C, dextran 5g, trehalose 2g, ultra-low viscosity sodium alginate 0.3g, diatrizoate meglumine 3g, polydiallyl 0.1 g of dimethyl ammonium chloride was fully dissolved in sterile water for injection to prepare 100 mL of a physiological solution, and then filtered through a 0.22 μm filter membrane into a sterile, endotoxin-free container.

Embodiment 3

[0029] In an environment with local 100-level laminar flow purification conditions, at 20°C, 3.5g of dextran, 3.5g of trehalose, 0.4g of ultra-low viscosity sodium alginate, 2.5g of diatrizoate, poly 0.15 g of diallyl dimethyl ammonium chloride was fully dissolved in sterile water for injection to prepare 100 mL of a physiological solution, and then filtered through a 0.22 μm filter membrane into a sterile, endotoxin-free container.

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Abstract

The invention discloses a method for separating lymphocytes and belongs to the technical field of biology. The method comprises the following steps: fully dissolving the following components that meet the medical standards: 3 to 5 parts by weight of dextran, 2 to 4 parts by weight of mycose, 0.3 to 0.8 part by weight of sodium alginate with an ultralow viscosity, 1 to 3 parts by weight of meglumine diatrizoate, and 0.1 to 0.15 part by weight of poly(diallyl dimethyl ammonium chloride) into sterile water to prepare 100 parts by weight of physiological solution; filtering the physiological solution by a filter membrane to remove the bacteria to obtain a separated liquid; adding the separated liquid into a centrifuge tube, wherein the volume of the separated liquid is not more than 1/2 of the volume of the centrifuge tube; paving a sample, which has been processed by an anticoagulant and diluted by a balanced salt solution or serum-free culture medium, on the separated liquid, carrying out centrifugation for 10 to 30 minutes at a temperature of 8 to 25 DEG C under a gravity of 200 to 800 g; wherein the sample is derived from peripheral blood, umbilical cord blood or marrow. The obtained lymphocytes have a good tumor killing activity.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a separation method for separating lymphocytes. Background technique [0002] During the standard separation process, the cells must be in contact with the separation medium for 15-45 minutes to achieve the best separation effect. During this period, the components in the separation solution may accumulate inside the cells through passive diffusion, active cell transport, endocytosis / pinocytosis, surface adhesion, etc., and this part of the accumulated components is difficult to pass through conventional methods Washing completely removes the isolated target cells (eg, lymphocytes). [0003] Moreover, the current gradient centrifugation method has cumbersome steps, strong skills, and relatively high requirements for the operator's experience: gradient centrifugation requires three steps, especially the second step. Density of the cell suspension without confusing the interface will...

Claims

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Application Information

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IPC IPC(8): C12N5/0781C12N5/0783
CPCC12N5/0635C12N5/0636C12N2500/30C12N2500/34C12N2500/46C12N2509/10
Inventor 宫喜魁李若鲲马艳马艳玲郝丽敏
Owner 北京弘润天源基因生物技术有限公司
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