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Method for separating and culturing mouse cerebellar granule cells

A granulosa cell and culture method technology, applied in the field of mouse cerebellar granulosa cell separation and culture, can solve the problems of reduced number and survival rate of cultured neurons, uncertainty of trypsin digestion intensity, low purity of cerebellar granulosa cells, etc. The effect of saving tedious steps and reducing dosage and improving survival rate

Inactive Publication Date: 2017-03-15
JIANGYIN CHI SCI
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Problems solved by technology

At present, there are many methods for the in vitro isolation and culture of cerebellar granule cells at home and abroad, but the purity of the obtained cerebellar granule cells is not high due to the pollution of non-neuronal cells, and the separation steps are complicated and the strength of trypsin digestion is uncertain. Result in decreased number and survival of cultured neurons

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  • Method for separating and culturing mouse cerebellar granule cells
  • Method for separating and culturing mouse cerebellar granule cells
  • Method for separating and culturing mouse cerebellar granule cells

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Embodiment Construction

[0021] In order to present the purpose and advantages of the present invention more clearly, specific embodiments will now be further described. The specific embodiments described here are only for explaining the present invention, and are not intended to limit the present invention.

[0022] The present invention selects newborn 8-day-old SD mice, and separates cerebellar granule cells. The specific operation is as follows:

[0023] 1. Take 8-day-old newborn mice and kill them by neck dislocation. Fix the mouse, soak it in 75% alcohol for 5 minutes, quickly cut the skin and skull from both sides of the mouse head, lift the back skull of the mouse, expose the cerebellum and part of the brain, gently peel off the whole cerebellum with tweezers and place it in a pre-cooled In PBS buffer;

[0024] 2. Quickly separate the meninges and blood vessels attached to the cerebellum under a microscope on ice, and transfer the stripped cerebellum to a new PBS buffer to wash twice;

[0...

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Abstract

The invention provides a method for separating and culturing mouse cerebellar granule cells. The method comprises (a) killing a mouse through breaking the neck, disinfecting the mouse through alcohol, separating mouse cerebellum, putting the cerebellum in a pre-cooled sterile dual antibody-containing PBS buffer, and fast and carefully stripping a meninx and blood vessels on the cerebellum through a dissecting microscope, (b) digesting the cerebellum through a preheated mixed enzyme solution in an incubator at a temperature of 37 DEG C for 10-15min, (c) acquiring cerebellar granule cells through combination of a differential sedimentation method and a differential attachment method, and (d) during the culture process, adding an appropriate amount of cytarabine at different time points to inhibit growth of non-neuronal cells. The method provided by the invention can acquire mouse cerebellar granule cells with high yield, a high survival rate and high purity.

Description

technical field [0001] The invention belongs to the field of cell biology, in particular to a method for separating and culturing mouse cerebellar granule cells. Background technique [0002] Neurons are the most basic structural and functional units of the nervous system, and cerebellar granule cells are a type of neurons with abundant content and small size in the central nervous system. In view of the large number of cerebellar neurons, easy to obtain materials, and the characteristics of growth and differentiation similar to cerebral cortical neurons, cerebellar granule neurons are often used in in vitro neuron growth and development, nerve axon regeneration, nervous system damage repair and clinical neuropharmacology. Research. At present, there are many methods for the in vitro isolation and culture of cerebellar granule cells at home and abroad, but the purity of the obtained cerebellar granule cells is not high due to the pollution of non-neuronal cells, and the sep...

Claims

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Application Information

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IPC IPC(8): C12N5/071
Inventor 张清齐来俊蒋敏张亚洲
Owner JIANGYIN CHI SCI
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