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Kit for separating culture of olfactory ensheathing cells

A technology for olfactory ensheathing cells, separation and culture, applied in the field of rapid separation of olfactory ensheathing cells and kits for rapid separation of olfactory ensheathing cells, which can solve the problems of the purity, quantity and time of olfactory ensheathing cell separation

Inactive Publication Date: 2013-05-08
北京清美联创干细胞科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve the problems of the purity, quantity and time of olfactory ensheathing cell separation, the invention provides a kit for rapidly separating olfactory ensheathing cells

Method used

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  • Kit for separating culture of olfactory ensheathing cells
  • Kit for separating culture of olfactory ensheathing cells

Examples

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Embodiment 1

[0017] According to one aspect of the present invention, the method for quickly isolating olfactory ensheathing cells can be realized through the following steps: since fibroblasts are mainly affected on purification, adding cytarabine after 24-48 hours of culture can preferentially inhibit and kill fibroblasts entering the division phase, but OECs are also inhibited to a certain extent. Using BPE (bovine pituitary extract) to promote the proliferation of OECs, OECs in good growth state can be obtained on the 8th to 16th day, and the purity can reach 80%. The OECs cell surface has a specific low-affinity neurotrophic factor receptor (L-NGFR), which can specifically bind to the LNGFR antibody, providing a basis for immunopurification, and the purity can be increased to 98% after immunopurification.

[0018] The culture method of the present invention adopts the first half-quantity liquid replacement method after 24 hours of culture, and utilizes the addition of cytarabine after...

Embodiment 2

[0020] According to one aspect of the present invention, the method for rapidly separating olfactory ensheathing cells may include:

[0021] Primary isolation and culture of olfactory ensheathing cells

[0022] Wash the cells with the washing solution, collect the cells after digesting the digestive solution, centrifuge, discard the supernatant, blow and beat the cell culture mixture into a single cell suspension, and inoculate and culture. After 24 hours of inoculation, half of the medium was changed, and the culture was continued, and the cell culture mixture was changed every 3 days thereafter. The primary culture can be completed in 7 days.

[0023] Subculture of olfactory ensheathing cells

[0024] Discard the culture mixture of the primary culture, wash the cells fully with the washing solution, remove the round cells suspended on the surface of the adherent cells, add the cell digestion solution, and add the cell culture mixture when the cells retract and the intercel...

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Abstract

The invention provides a kit for rapidly separating olfactory ensheathing cells, and a method thereof. The kit comprises a washing liquid, a cell culture liquid A, a cell culture liquid B and a cell digestion liquid, wherein the washing liquid is an artificial cerebrospinal fluid, the cell culture liquid A is a DMEM culture medium, the cell culture liquid B is fetal calf serum, and the cell digestion liquid comprises 89% of pancreatin, 10% of a DNA enzyme and 1% of heparin. Compared with traditional separating culture methods, the method provided by the invention allows the cell purity to be high, the propagation to be fast and the primary cell culture to be generally completed 7 days later through adopting the kit to culture for 24h and then culture and purifying in a half-amount liquid replacement manner. Primary cells undergo passage and are cultured by the culture liquids, and the cell purity can reach 90% 2-3 days later, so an abundant cell source is provided for the further medical and biological researches on the olfactory ensheathing cells.

Description

technical field [0001] The invention belongs to the field of tissue engineering, and in particular relates to a kit for rapidly isolating olfactory ensheathing cells, an application of the kit and a method for rapidly isolating olfactory ensheathing cells. Background technique [0002] OECs are one of the very few cells found in the central nervous system that can regenerate. It is characterized by its lifelong nerve regeneration function and the ability to release a variety of neurotrophic factors and nerve adhesion molecules. It is considered to be the glial cell with the strongest myelinating ability, and is gradually being used to treat spinal cord injuries. OECs share phenotypes with glial cells and Schwann cells. They can both promote axon regeneration. The main difference is that OECs exist not only in the central nervous system, but also in the peripheral nerves. Neurons in the olfactory mucosa are the only neurons that grow after birth and continue to differentiate...

Claims

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Application Information

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IPC IPC(8): C12N5/079
Inventor 田杰
Owner 北京清美联创干细胞科技有限公司
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