CN115678778A - Micro-fluidic chip device for culturing three-dimensional cell clusters - Google Patents
Micro-fluidic chip device for culturing three-dimensional cell clusters Download PDFInfo
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- CN115678778A CN115678778A CN202211461434.2A CN202211461434A CN115678778A CN 115678778 A CN115678778 A CN 115678778A CN 202211461434 A CN202211461434 A CN 202211461434A CN 115678778 A CN115678778 A CN 115678778A
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Abstract
The invention relates to a microfluidic chip device for culturing a three-dimensional cell mass, which is a microfluidic chip combining a micro-channel made of silicon-based organic polymer Polydimethylsiloxane (PDMS) and a stainless steel pore plate with an array structure, and is used for culturing the cell mass in a hanging drop mode. The microfluidic chip manufactured by the invention can generate a large amount of cell clusters and form an array structure, thereby facilitating the operation and research of the cell clusters. The microfluidic chip manufactured by the invention has the characteristics of low cost, easy operation, high yield, array and the like.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a microfluidic chip device which has high flux and is provided with an array structure and used for culturing a three-dimensional cell mass.
Background
Cancer is always a major problem affecting the global human health, the incidence and mortality of cancer are continuously increasing, the treatment means is also a problem in the research in the fields of medicine, biology and the like, the early symptoms are not obvious, and the treatment difficulty is high, so that the establishment of a solid tumor model in vitro is very important. Understanding the processes of tumor formation, metastasis and the microenvironment in which they reside is crucial to achieving early diagnosis and treatment of tumors. The in vitro construction of tumor models becomes an important means for researches on canceration mechanism, screening and toxicity evaluation of anti-cancer drugs, drug resistance mechanism of cancer cells and the like, and plays an important role in explaining and understanding the complex canceration process. On one hand, the method can be used for drug detection and research and development of new anti-cancer drugs, and on the other hand, the method can be used for designing personalized treatment through a tumor model and implementing an optimal treatment scheme.
The culture technique of simulating the environment suitable for the growth of cells in vivo in vitro, culturing cells taken from organs or tissues in vivo in vitro to grow, proliferate and maintain the morphology and function of the cells is called cell culture. Cell culture techniques are continuously developed from single cell culture to multi-cell co-culture, from monolayer cell culture to three-dimensional cell culture and even organ chips which are rising in recent years.
At present, the common methods for culturing cell clusters include bioreactors, supports, hanging drop technology and microfluidic technology-based cell cluster construction. The micro-fluidic technology mainly uses a micro-scale channel to process micro liquid, can realize accurate control of mass transmission, and is very suitable for controlling a cell microenvironment to perform dynamic culture of cells. Meanwhile, cells can grow on the microfluidic chip, so that the microfluidic platform can be widely used for constructing three-dimensional cell groups.
Disclosure of Invention
Aiming at the defects in the prior art, the invention provides a microfluidic chip device for culturing three-dimensional cell clusters, which can be used for manufacturing high-flux three-dimensional cell clusters and enabling the cell clusters to have an array structure by means of a hanging drop method and a microfluidic technology. The device can be used for rapidly culturing a large number of various fine groups, is convenient for establishing an in vitro microenvironment model and relevant research, has great significance for researching cancer models and inflammation models, and can also be used for simulation experiments. In addition, the device is also very convenient for detecting the influence of the medicine on the cell activity.
In order to achieve the purpose, the invention adopts the following technical scheme:
a microfluidic chip device for culturing three-dimensional cell clusters is composed of an upper layer and a lower layer, wherein the upper layer is a micro-channel for liquid flowing and replacement, the lower layer is a stainless steel pore plate with an array structure, and the upper and lower layers are bonded and fixed through Polydimethylsiloxane (PDMS).
The upper-layer structure micro-channel is made of PDMS, a micro-channel with a snake-shaped structure is arranged on the upper-layer structure micro-channel, and round holes are formed at two ends of the micro-channel and are used as an inlet and an outlet of liquid, so that the liquid can flow and be replaced conveniently.
The lower layer structure is a stainless steel pore plate, and round micropores with an array structure are arranged on the lower layer structure and are convenient for the growth of cell clusters.
The invention injects cell suspension with certain concentration into the micro-channel through a liquid transfer device, the cells can fall into micropores of a stainless steel pore plate to form a hanging drop state under the action of gravity, the cells can grow into cell clusters in the hanging drop, and the cell clusters can also form a corresponding array structure after growing due to the pore plate with the array structure, and the cell clusters can be used for experiments.
The experimental procedure using this device was as follows:
1) The upper microchannel and the lower stainless steel well plate of the device were bonded with PDMS.
2) Injecting epithelial cell suspension containing a certain concentration into the micro flow channel through the inlet of the upper layer structure by using a syringe pump, injecting a proper amount of culture solution, and standing for culture.
3) After standing, the cells which do not fall into the holes are washed for a plurality of times by using the culture solution in a solution changing mode, and the cells are ensured to fall into the holes.
4) After the culture, the growth of the cell mass was observed with a microscope.
Compared with the prior art, the invention has the following advantages:
the device can be manufactured by only utilizing PDMS and a stainless steel pore plate with lower cost without using various devices for various complicated operations, thereby greatly reducing the cost and simplifying the operation mode of the experiment. Moreover, the device provides convenience for microenvironment, inflammation model, cancer model and tumor model. Meanwhile, the device is low in preparation cost, simple in structure and capable of being repeatedly utilized, and can be used for cell mass construction experiments of various cells.
Drawings
Fig. 1 is a structural diagram of a microfluidic chip device according to an embodiment of the present invention.
FIG. 2 is a drawing of a lower stainless steel orifice plate of the present invention and its dimensional parameters.
FIG. 3 is a schematic diagram of an upper layer PDMS structure according to the present invention.
FIG. 4 is a graph showing the results of the experiment according to the present invention.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described below by using preferred embodiments in conjunction with the accompanying drawings. The embodiments and features of the embodiments in the present application may be combined with each other arbitrarily.
As shown in fig. 1, a microfluidic chip device for culturing three-dimensional cell clusters is composed of an upper layer and a lower layer, wherein the upper layer is a micro-channel for liquid flowing and replacement, the lower layer is a stainless steel pore plate with an array structure, and the upper and lower layers are bonded and fixed by Polydimethylsiloxane (PDMS).
As shown in FIG. 3, the upper micro flow channel is made of PDMS, has a depth of 2mm, and has a micro flow channel with a snake-shaped structure, and round holes with a diameter of 2mm are formed at two ends of the micro flow channel and are used as an inlet and an outlet of liquid, so as to facilitate the flow and replacement of the liquid.
As shown in FIG. 2, the lower layer structure is a stainless steel pore plate, which has round micropores in an array structure, wherein the pore plate has a length of 14cm, a width of 10cm and a thickness of 1mm, and the diameter of the micropores is 1mm, and the micropores facilitate the growth of cell clusters.
In this example, the present apparatus was used to perform an epithelial cell clustering experiment:
firstly, an upper micro-channel is manufactured, a graph is designed through CAD software, then processing is carried out, and the upper micro-channel is obtained through mask and rollover processing. And selecting a stainless steel orifice plate with a proper array, and after the two orifice plates are prepared, coating the edge of the micro-channel with the prepared PDMS liquid, and fixing the micro-channel on the stainless steel orifice plate. Then the product is placed on a heating table at 55 ℃, covered by an article with a certain weight and heated for 3 hours, and then the product can be used.
Spraying alcohol on the whole device, placing under ultraviolet light for 1 hour, then slowly injecting epithelial cell suspension containing a certain concentration through the inlet of the micro-channel by using an injection pump, injecting a proper amount of culture solution, and then standing for culture. Culturing for 15min, flushing with culture solution to replace the liquid so that cells not falling into the hole fall into the hole, and standing in an incubator. After 24 hours of culture, the medium was changed and the growth of the cell mass was observed with a microscope, as shown in FIG. 4. After a period of incubation, cell aggregation was observed.
In the above experiment, the target drug may be added to a culture solution containing an epithelial cell mass, followed by observation to observe the state of cell activity, and further, the effect of the drug on the cell activity may be analyzed.
The microfluidic chip device for culturing the three-dimensional cell mass in the above embodiment is a microfluidic chip combining a micro channel made of silicon-based organic polymer Polydimethylsiloxane (PDMS) with a stainless steel pore plate having an array structure, and culturing the cell mass in a hanging drop manner. The microfluidic chip manufactured by the embodiment of the invention can generate a large number of cell clusters and form an array structure, thereby facilitating the operation and research of the cell clusters. The microfluidic chip manufactured by the invention has the characteristics of low cost, easy operation, high yield and arraying.
The embodiments of the present invention have been described with reference to the accompanying drawings, but the present invention is not limited to the embodiments, and various changes and modifications can be made according to the purpose of the invention, and any changes, modifications, substitutions, combinations or simplifications made according to the spirit and principle of the technical solution of the present invention shall be equivalent substitutions, as long as the purpose of the present invention is met, and the present invention shall fall within the protection scope of the present invention without departing from the technical principle and inventive concept of the present invention.
Claims (3)
1. A micro-fluidic chip device for culturing three-dimensional cell clusters is characterized in that: the micro-flow channel structure is composed of an upper layer and a lower layer, wherein the upper layer is a micro-flow channel for liquid flowing and replacement, the lower layer is a stainless steel pore plate with an array structure, and the upper layer and the lower layer are bonded and fixed through Polydimethylsiloxane (PDMS).
2. The microfluidic chip device for culturing a three-dimensional cell mass according to claim 1, wherein: the upper-layer structure micro-channel is made of PDMS, a micro-channel with a snake-shaped structure is arranged on the upper-layer structure micro-channel, and round holes are formed at two ends of the micro-channel and are used as an inlet and an outlet of liquid, so that the liquid can flow and be replaced conveniently.
3. The microfluidic chip device for culturing three-dimensional cell masses according to claim 1, wherein: the lower layer structure is a stainless steel pore plate, and round micropores of an array structure are formed in the lower layer structure and are convenient for growth of cell clusters.
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| Application Number | Priority Date | Filing Date | Title |
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| CN202211461434.2A CN115678778A (en) | 2022-11-21 | 2022-11-21 | Micro-fluidic chip device for culturing three-dimensional cell clusters |
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| CN202211461434.2A CN115678778A (en) | 2022-11-21 | 2022-11-21 | Micro-fluidic chip device for culturing three-dimensional cell clusters |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116855366A (en) * | 2023-08-19 | 2023-10-10 | 北京航空航天大学 | Cell chip for metabolic fingerprint spectrum visualization and application method |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116855366A (en) * | 2023-08-19 | 2023-10-10 | 北京航空航天大学 | Cell chip for metabolic fingerprint spectrum visualization and application method |
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