Neural stem cell medium change system and medium change method for neural stem cell culture

A technology for neural stem cells and cell culture, applied in the field of replacing cell culture medium, can solve the problems of time-consuming and laborious, loss of neural stem cells, affecting the long-term cultivation of neural stem cells, etc., to save manpower, prevent the loss of neural stem cells, and facilitate industrialized production.

Inactive Publication Date: 2015-12-09
ZHEJIANG ORIGIN BIOTECH
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

It is understood that one of the most common methods at home and abroad is the direct aspiration method and the centrifugation method, that is, the cell culture flask is erected first, and after the neural stem cell spheres are precipitated, the supernatant is gently aspirated, and then an equal amount of fresh culture medium is added. It was determined that the removed supernatant contained some single neural stem cells and small neural stem cell spheres, which directly caused the loss of the number of neural stem cells during the culture process.
The other method is the centrifugation method. First, the cell culture bottle is erected. After the neural stem cell spheres settle, use a pipette to transfer the supernatant to a centrifuge tube, centrifuge at 1500 rpm for 20 minutes, and discard the supernatant. Single neural stem cells and small neural stem cell spheres were collected by centrifugation, and returned to the culture bottle, and fresh culture medium was added to culture. The disadvantage of this method is that it is time-consuming and laborious, and the centrifuged precipitation liquid contains long-term cultured cell fragments. Tissues containing these debris affect long-term culture of neural stem cells

Method used

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  • Neural stem cell medium change system and medium change method for neural stem cell culture
  • Neural stem cell medium change system and medium change method for neural stem cell culture
  • Neural stem cell medium change system and medium change method for neural stem cell culture

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Experimental program
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Effect test

Embodiment 1

[0023] A neural stem cell replacement system, such as figure 1 and figure 2 As shown, the system includes a sequentially connected neural stem cell replacement cell blocker A5, a waste liquid storage bottle 3, a sterile air bottle 2, and an air pump 1. The main body of the cell blocker A is a titanium rod filter head and a stainless steel tube 6 Connect, and then connect with the pipe, titanium rod filter head specifications: the titanium rod filter head is flat-topped and the lower end is arc-shaped, and the flat top is connected to the stainless steel pipe. About 4.5 μm), the length of the stainless steel tube is 215 mm, the outer diameter is 6 mm, and the inner diameter is 4 mm. The stainless steel tube is connected to the waste liquid storage bottle 3 through the connecting tube, and the end of the stainless steel tube connected to the connecting tube 7 is provided with a pagoda-shaped threaded port. The material of titanium rod filter head is titanium alloy.

[0024] A...

Embodiment 2

[0029] A neural stem cell replacement system such as image 3 As shown, the system includes a syringe 10, a cell arrester B9 and a cell culture bottle 4 connected in sequence. The cell arrester B9 is a stainless steel tube with a built-in stainless steel filter 8, and the stainless steel filter 8 has a pore size of 4.5 μm. The volume of the syringe 10 is 100-200ml. One end of the cell blocker B is connected to the syringe head of the syringe, and the other end of the cell blocker B is connected to the pipette through a leather tube, and the pipette is directly inserted into the cell culture bottle 4 to be changed, and manually The pulling force of the syringe 10, when sucking out 100ml of liquid, makes the floating neural stem cells be stopped outside the stainless steel filter. Therefore, the liquid treated by the neural stem cell replacement system does not contain any neural stem cells or floating cells, but may contain cell fragments, and these liquids are used as waste l...

Embodiment 3

[0032] Effects of direct discarding liquid method and liquid exchange system in this method on cell loss

[0033] The direct discarding method is to stand the neural stem cell culture bottle to be changed for 20 minutes. After all the neural stem cell spheres have settled to the bottom of the bottle with the naked eye, take out the supernatant, and check 100 fields of view under a microscope with a magnification of 100 times. RESULTS: 1-7 NSCs and suspension cells per field of view, up to 100%. The liquid exchange system of this method is to use the waste liquid extracted from the neural stem cell liquid exchange system, and check 100 fields of view under the condition of 100 times magnification of the microscope. Results: Among 100 visual fields, only one neural stem cell was found in one visual field, 1%. Only a small amount of cell debris tissue. It shows that the liquid exchange system of this method prevents the loss of valuable cells, and also removes the debris tissue...

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Abstract

The invention relates to a cell culture medium change method, in particular to a neural stem cell medium change system and a medium change method for neural stem cell culture. The neural stem cell medium change system comprises a cell interceptor, a waste storage bottle, a sterile air bottle and an air pump in sequential connection, the cell interceptor is formed by connection of a titanium rod type filter head and a stainless steel pipe, and aperture of the titanium rod type filter is 4.0-5.0 micrometers. Alternatively, the neural stem cell medium change system structurally comprises the cell interceptor, a syringe and a connection pipe, the cell interceptor is made of the stainless steel pipe with a stainless steel filter screen inside, and mesh diameter of the stainless steel filter screen is 4.0-5.0 micrometers. The neural stem cell medium change system and the medium change method for neural stem cell culture have the advantages that medium change time in a neural stem cell culture process can be shortened, loss of precious neural stem cells in a medium change process is prevented, residual tissue fragments in a culture medium can be cleared away, lives of cells cultured in vitro are prolonged, and labor saving, productivity improvement and convenience for industrial production are realized.

Description

technical field [0001] The invention relates to a method for replacing cell culture fluid, in particular to a neural stem cell fluid replacement system and a fluid replacement method for neural stem cell culture. Background technique [0002] Mass culture of human neural stem cells is mainly used to prevent and treat neurodegenerative diseases, such as Parkinson's disease, Alzheimer's disease, spinal cord injury and other diseases. Usually, during the culture of neural stem cells, the medium needs to be changed regularly to remove part of the metabolic waste fluid, and then add an equal amount of fresh nutrient solution, so as to ensure the continued division and growth of neural stem cells. It is understood that one of the most common methods at home and abroad is the direct aspiration method and the centrifugation method, that is, the cell culture flask is erected first, and after the neural stem cell spheres are precipitated, the supernatant is gently aspirated, and then ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0797C12M3/00C12M1/24
Inventor 陈世明江克光
Owner ZHEJIANG ORIGIN BIOTECH
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