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Method for obtaining human OPCs and OPCs medium

A medium and fresh medium technology, applied in the field of obtaining human OPCs, can solve the problems of increased cost of culture, limited cell yield, etc., and achieve the effect of easy operation

Active Publication Date: 2014-10-01
栾佐
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the induction process of these two types of cells requires a long time and the joint action of various cytokines, which will inevitably lead to an increase in the cost of culture and limit the production of cells.

Method used

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  • Method for obtaining human OPCs and OPCs medium
  • Method for obtaining human OPCs and OPCs medium
  • Method for obtaining human OPCs and OPCs medium

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] Example 1: Sorting Human OPCs from the Third Generation Human Neural Stem Cells

[0091] 1 Digestion of third-generation neural stem cells into single cells

[0092] ①Centrifuge at 100g for 5 minutes to collect neural stem cells, discard the supernatant;

[0093] ②Add 1ml of 0.025% trypsin (diluted in 0.01M dPBS), mix well, and incubate in a 37°C incubator for 5-7min until the cell balls become loose;

[0094] ③ Add 100 μl of 1.2 mg / ml trypsin inhibitor, and pipette gently with a 200 μl pipette to form a single-cell suspension;

[0095] Neural stem cell culture medium:

[0096]

[0097] 2 Separation of OPCs from human neural stem cells using magnetic beads

[0098] ①The single-cell suspension of neural stem cells was passed through a 40 μm nylon mesh to filter out cell clumps, and trypan blue was used to calculate the total number of cells;

[0099] ② take 10 7 Cells were collected by centrifugation at 300g for 10 minutes, and the supernatant was removed;

[01...

Embodiment 2

[0128] Example 2: Sorting human OPCs from the thirtieth passage of human neural stem cells

[0129] 1 The thirtieth generation neural stem cells were digested into single cells (the method is the same as in Example 1)

[0130] 2 Use magnetic beads to sort OPCs from human neural stem cells (method is the same as in Example 1)

[0131] 3 OPCs in vitro expansion ability and differentiation ability detection (method is the same as embodiment 1)

[0132] Under the microscope, it was found that the OPCs sorted from P30 neural stem cells also had typical bipolar or tripolar morphology, and highly expressed OPCs-specific markers O4, PDGFαR and Sox10, and only a few cells expressed astrocyte markers After induction of GFAP and neuron marker Tuj-1, some cell synapses formed a longer, dendritic multi-branched complex network morphology, which could be recognized by multi-branched O4 and Galc.

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Abstract

The invention relates to a method for obtaining human OPCs (Oligodendrocyte Progenitor Cells) and an OPCs medium. According to the method, neural stem cells are used as a sorting source, the human OPCs can be obtained by separation through an immunomagnetic bead method, and a more economical medium suitable for OPCs in vitro culture and passage, so that a great amount of high-purity human OPCs can be obtained in a short time, and a new choice is provided for clinical application.

Description

technical field [0001] The invention relates to a method for obtaining human OPCs and an OPCs culture medium. Background technique [0002] Myelin-related diseases such as white matter injury, spinal cord injury, and multiple sclerosis are caused by the myelinization disorder or demyelination of neurons, which leads to the inability of neurons to transmit electrical excitation normally, resulting in functional disability such as body movement. Since there is currently no drug that can effectively treat these myelin-related diseases, it has brought great harm to patients and families of these diseases, which makes effective treatment of myelin-related diseases an urgent social and social issue to be solved today. medical conundrum. [0003] Whether it is a myelinating disorder or a demyelinating lesion, the root cause is the reduction or loss of the myelin sheath of the neuron axon. Therefore, if the degree of myelinization of the neuron axon can be restored, the neuron curr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0797
Inventor 栾佐
Owner 栾佐
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