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Multi-functional-region cell three-dimensional co-culture method based on micro-fluidic chip

A microfluidic chip and multi-functional technology, applied in three-dimensional co-culture of cells and its application fields, can solve problems such as inability to construct, unsuitable conditions for two-dimensional culture, complex co-culture system, etc., and achieve the effect of short time consumption

Active Publication Date: 2016-06-29
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a new method of three-dimensional co-cultivation of cells in a multi-functional area based on a microfluidic chip, to solve the problems that the complex co-cultivation system cannot be constructed in previous cell experiments and the two-dimensional culture does not meet the actual conditions in the body.

Method used

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  • Multi-functional-region cell three-dimensional co-culture method based on micro-fluidic chip
  • Multi-functional-region cell three-dimensional co-culture method based on micro-fluidic chip
  • Multi-functional-region cell three-dimensional co-culture method based on micro-fluidic chip

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Embodiment 1

[0032] After the microfluidic chip of the present invention is prepared, it is sterilized overnight by ultraviolet irradiation in a clean bench. Prepare 30 microliters of 4mg / ml collagen solution according to the conventional ratio and divide it into three equal parts (10 microliters each) and store them on ice packs for later use. Cells were resuspended with a portion of collagen solution after passage. Add the above-mentioned collagen solution containing cells to the middle liquid pool of the central channel; add the other two blank collagen solutions to the middle and two side liquid pools of the central channel respectively; vacuumize at the other end of the central channel, and the three liquids flow in laminar flow In the channel, the collagen is solidified at 37 degrees for half an hour to form a collagen distribution structure with a cell area in the middle and blank collagen areas on both sides. Add medium to the side channels on both sides for cell culture ( figure...

Embodiment 2

[0034] Based on the method for forming the multifunctional region in Example 1, collagen containing endothelial cells was added to the middle liquid pool of the central channel, and a blank collagen solution was added to the liquid pools on both sides to remain unchanged. After the collagen was solidified, culture medium was added to the left channel; tumor cells and endothelial cells were co-cultured with endothelial cells in the right channel. Such as image 3 As shown, after 48 hours, the endothelial cells migrated and sprouted to the right under the induction of tumor cells, while the control group added to the medium channel on the left did not see endothelial cells sprouting in this direction. This experiment was completed within one time. Co-cultured cell interaction research and blank control experiments have been carried out, reflecting the superiority of the present invention. In addition, after the system was cultured for a longer period of time, the endothelial ce...

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Abstract

The invention provides a multi-functional-region cell three-dimensional co-culture method based on a micro-fluidic chip. The chip is composed of a central main channel which is relatively low and side channels which are relatively high and are arranged on two sides, wherein the main channel and the side channels are connected by virtue of trumpet-shaped grid structures. Specifically, the method comprises the following steps: (1) adding collagen solutions different in composition to the sample inlet of the main channel of the chip and applying negative pressure to the outlet of the main channel, wherein the three collagen solutions flow into the main channel and functional regions, which keep distinct boundaries, are formed on the basis of a lamina flow principle; and (2) sterilizing the chip for a whole night through ultraviolet radiation, and directly blending cells to the collagen solutions which are injected into the main channel, so that cell inoculation can be achieved. By virtue of the micro-fluidic chip designed by the invention, a cell three-dimensional culture system, which has a plurality of independent functional regions, can be constructed in one step; and the method has an important application potential in the construction of in vitro complex cell micro-environments.

Description

technical field [0001] The invention relates to the field of three-dimensional co-cultivation of cells and its application, in particular to a method for three-dimensional co-cultivation of cells in a multifunctional area based on a microfluidic chip. Background technique [0002] The three-dimensional co-culture system of cells is an important platform for the interaction between cells and the construction of cell microenvironment. Compared with the traditional two-dimensional culture system, three-dimensional cell culture is closer to the growth state of cells in vivo, so it is more conducive to the maintenance of cell phenotype and the simulation of cell microenvironment, and the research results can more truly reflect the situation in vivo. The cell co-culture system is an important means to study the interaction between cells, including various behaviors such as chemotaxis, recognition, and penetration between cells and their related substrates. [0003] At present, th...

Claims

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Application Information

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IPC IPC(8): C12M3/00C12N5/09C12N5/071
Inventor 秦建华张旭许慧
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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