KR20230120487A - Modular microfluidic chip with individual flow rate control and oxygen concentration control - Google Patents

Abstract

The present invention is a microfluidic chip which simulates human organs, and a modular microfluidic chip which allows adjustment of the oxygen concentration and fluid flow rate required for cell culture, and provides a highly efficient hypoxic target disease screening or drug testing platform by modularizing the microfluidic chip's channels and organ chips. To solve this purpose, the present invention comprises: a chip module (100) in which cells are cultured using a cell culture medium and a gas permeable membrane (112) formed on the bottom; a flow rate control module (200) which allows cell culture medium to flow into the chip module (100); an oxygen control module (300) which provides hypoxic conditions to the chip module (100) while flowing an oxygen absorption solution and a diluent toward the gas permeable membrane (112); and a module port layer (400) which is laminated on the upper surface of the oxygen control module (300) and allowing the chip module (100) and a flow rate control module (200) to be detachably assembled.

Description

Modular microfluidic chip with individual flow rate control and oxygen concentration control}

The present invention is a microfluidic chip that mimics human organs, which enables control of the oxygen concentration and flow rate of fluids required for cell culture, and makes the channels and organ chips of the microfluidic chip modular, enabling very efficient hypoxia target disease screening or drug testing It relates to a modular microfluidic chip that provides a platform.

In general, drug testing is performed through animal testing, and drug testing using animals requires the development of a test platform that can replace animal testing due to ethical issues. In particular, in the case of animal testing for hypoxic diseases, artificial This can cause serious ethical problems in the process of inducing hypoxic conditions in animals. Recently, a microfluidic chip has been developed that can simulate the physiological function of the organ and its response to drugs by culturing cells constituting a specific organ. there is.

For example, looking at Korean Patent Registration No. 10-2279961, a cell culture layer; membrane layer; an oxygen absorbing layer containing an oxygen absorbing solution; an optical oxygen sensor located above the membrane layer facing the oxygen absorbing solution; and an optical detector for measuring a change in fluorescence of the oxygen sensor according to the oxygen concentration, wherein the oxygen absorbing layer controls the preeclampsia environment caused by hypoxia by controlling the flow rate of the upper and lower layers of the oxygen absorbing solution. An implemented placenta-like organ chip is presented.

However, in the prior art, since it has a structure that induces hypoxia while simply controlling the flow rate of the oxygen absorbing solution, the oxygen concentration in the controlled state is not constant due to the flow characteristics of the fluid, and in the case of cell culture media, the flow rate and the shear stress applied to the cells I had a problem that I couldn't control.

In addition, conventionally, microfluidic chips can only simulate hypoxia for specific organs, and cells constituting human organs often have different culture conditions, so it is cumbersome to remanufacture the channels of the microfluidic chip in response to the cultured cells. However, there was a problem in that various organ cells could not be cultured on a single microfluidic chip.

Korean Registered Patent Publication No. 10-2279961 (registration date: 2021.07.15)

The present invention is to solve the conventional problems, and is configured to adjust the oxygen concentration by determining the ratio of the oxygen absorbing solution and the dilution solution corresponding to various organ chips, while adjusting the oxygen concentration respectively corresponding to the organ chips. Its purpose is to keep the oxygen concentration constant.

In addition, according to the present invention, channels for supplying cell culture fluid are formed in different sizes in correspondence with various organ chips, so that the flow rate of the cell culture fluid and the shear stress applied to the cells are different for each corresponding organ chip. Its purpose is to be controlled.

In addition, the present invention is composed of a modular structure in which a channel for supplying cell culture medium and an organ chip are selectively combined and coupled to a module port, so that human organs with different culture conditions can be easily cultured on a single microfluidic chip. But it has a purpose.

The present invention to solve this object;

a chip module in which cells are cultured by a cell culture medium and a gas permeable membrane is formed on a bottom surface;

a flow control module for allowing the cell culture medium to flow through the chip module;

an oxygen control module providing a hypoxic condition to the chip module while flowing an oxygen absorbing solution and a diluting solution toward the gas permeable membrane; and

A modular microfluidic chip capable of adjusting the flow rate and oxygen concentration, characterized in that it is configured to include; a module port layer that is laminated on the upper surface of the oxygen control module and allows the chip module and the flow control module to be detachably assembled, respectively. provides

According to the present invention, there is an effect of maintaining a constant oxygen concentration corresponding to each corresponding organ chip, an effect of controlling the flow rate of the cell culture medium and the shear stress applied to the cells, and a microfluidic chip This modularization has the effect of easily culturing human organs under different culture conditions on a single microfluidic chip.

1 and 2 are diagrams of the overall configuration of a modular microfluidic chip capable of controlling flow rate and oxygen concentration according to the present invention.
Figure 3 is a view showing a chip module of the modular microfluidic chip of Figure 1;
4 is a view showing a flow control module of the modular microfluidic chip of FIG. 1;
5 is a view showing an oxygen control module in the modular microfluidic chip of FIG. 1;
6 is a measurement graph for oxygen concentration control of the modular microfluidic chip of FIG. 1 .

The characteristics of the modular microfluidic chip capable of controlling the flow rate and oxygen concentration according to the present invention will be understood by the embodiments described in detail below with reference to the accompanying drawings.

On the other hand, in describing the embodiments, detailed descriptions of components that are widely known and used in the technical field to which the present invention belongs or does not belong will be omitted, and unnecessary descriptions will be omitted, and the present invention according to this will be omitted. This is to make the point more clear.

1 is a perspective view of a modular microfluidic chip, FIG. 2 is a cross-sectional view of main parts of a modular microfluidic chip, FIG. 3 is a perspective view of a chip module, FIG. 4 is a plan view of a flow control module, FIG. 5 is a plan view of an oxygen control module, 6 is a graph for an embodiment of a modular microfluidic chip according to the present invention.

Schematically looking at the modular microfluidic chip 1 capable of adjusting the flow rate and oxygen concentration accordingly, the chip module 100 in which cells are cultured; a flow control module 200 to flow the cell culture medium through the chip module 100; An oxygen control module 300 providing hypoxic conditions to the chip module 100; A module port layer 400 enabling the chip module 100 and the flow control module 200 to be attached or detached, respectively; is configured to include.

Hereinafter, the configuration of the modular microfluidic chip capable of controlling the flow rate and oxygen concentration according to the present invention will be described in detail.

First, the chip module 100;

Cells are cultured by the cell culture medium, and a gas permeable membrane 112 is formed on the bottom surface, and the chip module 100 corresponds to the section in which the cells are cultured, and is connected to the flow control module 200 through a tube to generate cells. As the culture medium is supplied and flowing, cells to simulate human organs can be cultured.

Meanwhile, the chip module 100 further includes a first organ chip 110a, a second organ chip 110b, and a third organ chip 110c that simulate human organs.

As shown in FIG. 2, the first chamber 111, which is a space for culturing cells, is provided in the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c, respectively. The chamber 111 has an inlet and an outlet so that the cell culture medium continuously flows.

Here, the first chambers 111 of the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c have different sizes corresponding to the simulated human organs. (110a) is for culturing liver cells, and the first chamber (111) is made in a straight shape, and the second organ chip (110b) is for culturing kidney cells, and the first chamber (111) is made in an arc shape. The third organ chip 110c is for culturing arterial cells, and the first chamber 111 is formed in a zigzag shape.

At this time, the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c have different shapes of the first chamber 111 in consideration of the shear stress and volume of the corresponding human organ. In the case of internal shear stress, the first organ chip (110a) is 0.1 ~ 0.5 dyne/cm^2, the second organ chip (110b) is 0.1 ~ 0.5 dyne/cm^2, and the third organ chip (110c) is 10 to 15 dyne/cm^2, and in the case of in vitro shear stress is 0.3 dyne/cm^2, the second organ chip 110b is 0.25 dyne/cm^2, and the third organ chip 110c is 9.25 dyne /cm^2.

In addition, the first long-term chip 110a, the second long-term chip 110b, and the third long-term chip 110c are made of polymethyl methacrylate (PMMA), which has low gas permeability to minimize the influence of external gases. But it has a purpose. A gas permeable membrane 112 is formed at the bottom of the chip module 100, and the gas permeable membrane 112 is made of Polydimethylsiloxane (PDMS), a transparent high molecular material capable of gas transmission.

The gas-permeable membrane 112 is formed to a thickness of 50 μm on the bottom surface of the first chamber 111, and when the oxygen absorbing solution flows toward the gas-permeable membrane 112, the oxygen concentration of the first chamber 111 is controlled to reduce hypoxia to the cultured cells. conditions will be provided.

In addition, the first long-term device chip 110a, the second long-term device chip 110b, and the third long-term device chip 110c each have a first inclined portion 113 having a downwardly contracted circumferential surface, and the first inclined portion Corresponding to (113), a second inclined portion 423 extending upward is formed on the module port layer 400.

The first inclined portion 113 is formed on the peripheral surfaces of the first long-term device chip 110a, the second long-term chip 110b, and the third long-term device chip 110c, respectively, and the second inclined portion 423 is a module port described later. It is formed on the inner circumferential surface of the second porthole 422 of the layer 400, and the first long-term chip 110a, second long-term chip 110b, and third long-term chip 110c are inserted into the second porthole 422, respectively. When assembled, the first inclined part 113 and the second inclined part 423 come into contact so that they are assembled in a luar-lock method. In this case, the first long-term chip 110a and the second long-term chip 110b ), it prevents the oxygen absorbing solution from leaking into the assembled part of the third organ chip 110c.

And, the flow control module 200;

The cell culture fluid flows through the chip module 100, and the flow control module 200 is connected to the inlet side and the outlet side of the chip module 100 through tubes, respectively, to continuously supply the cell culture fluid to the chip module 100. and the flow control module 200 is connected to a metering pump so that the cell culture medium is supplied from the outside.

On the other hand, the flow control module 200 includes a first flow control module 210a into which the cell culture medium is injected; And a second flow rate control module (210b) for discharging the cell culture solution that has passed through the chip module (100);

The first flow control module 210a and the second flow control module 210b are arranged in a symmetrical manner in both directions around the chip module 100, and the first flow control module 210a and the second flow control module 210a The flow control module 210b is detachably assembled to the first port hole 421 of the module port layer 400 so that the flow control module 200 can be modularized.

Here, the first flow control module 210a and the second flow control module 210b are provided with a first channel 211 through which the cell culture medium flows, and the first channel 211 constitutes the chip module 100. One or more first passages 211a are provided to supply the cell culture medium to the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c, respectively, and the one or more first passages 211a ) further includes those made of different sizes corresponding to the human organs to be simulated.

The first channel 211 has an inlet through which the cell culture medium is introduced and an outlet through which the inflowed cell culture medium is discharged, and one or more first passages 211a are connected to each other between the inlet and the outlet of the first channel 211. .

One or more first passages 211a are for supplying cell culture fluids differently in response to human organ-like cells cultured in the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c. The first passage 211a connected to the first long-term chip 110a has a width of 130 μm and a height of 200 μm, and a flow rate of 6 μl/min. The first passage 211a connected to the second long-term chip 110b has a width of 233 μm, The height is 200 μm, the flow rate is 20 μl/min, and the first passage 211a connected to the third organ chip 110c has a width of 493 μm, a height of 200 μm, and a flow rate of 74 μl/min.

And, the oxygen control module 300;

A low-oxygen condition is provided to the chip module 100 while the oxygen absorbing solution and the diluent flow toward the gas permeable membrane 112. It has an outlet through which the oxygen absorbing solution and the diluted solution are discharged, and the oxygen control module 300 is also connected to the metering pump so that the oxygen absorbing solution and the diluted solution are continuously supplied and flowed.

On the other hand, the oxygen control module 300 is provided with a second channel 310 into which the oxygen absorbing solution and the diluent are respectively injected, and the second channel 310 mixes the oxygen absorbing solution and the diluent and supplies one or more of them. It further includes that the second passage 311 is provided.

The second channel 310 has an inlet through which the oxygen absorbing solution and the diluent are introduced, an outlet through which the oxygen absorbing solution and the dilution solution are discharged, and one or more channels are provided between the inlet and the outlet of the second channel 310. Two passages 311 are provided connected to each other.

At this time, the chip module 100 is seated in the second channel 310 to form a second chamber 312 through which the oxygen absorbing solution and the diluent flow through the gas permeable membrane 112, and the second chamber 312 It is connected to the side where the chip module 100 is assembled.

The one or more second passages 311 pass the oxygen absorbing solution and dilution solution differently in response to human organ-like cells cultured in the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c. The one or more second passages 311 correspond to the positions of the first long-term chip 110a, the second long-term chip 110b, and the third long-term chip 110c. ), and while one or more second passages 311 are connected to each other, the oxygen absorbing solution and the diluent are mixed and flowed through all the second passages 311.

At this time, the second passage 311 is provided with an inlet through which the oxygen absorbing solution flows and an inlet through which the diluted solution flows, respectively, so that the oxygen absorbing solution and the diluted solution are generally supplied to one or more second passages 311.

Here, the oxygen control module 300 may adjust the oxygen concentration corresponding to the dilution rate of the dilution solution in the oxygen absorption solution, and the oxygen control module 300 controls the oxygen absorption solution at 100 μl/min as shown in the measurement graph of FIG. After flowing for more than 10 minutes at a flow rate of , the oxygen concentration at the outlets of the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c were measured. The dilution ratio of the diluent to the absorption solution was determined.

Meanwhile, the dilution solution can be diluted differently according to the human organ to be simulated according to the measurement value as described above, and the dilution rate of the oxygen absorption solution is 10% for the first organ chip 110a. , The second organ chip 110b has a ratio of 60%, and the third organ chip 110c has a ratio of 40%.

Here, the oxygen absorbing solution uses a mixture of 0.5 g of cobalt nitrate, 25 μl of sodium sulfite, and 50 ml of DW, and DW can be used as the diluted solution.

And, the module port layer 400;

It is laminated on the upper surface of the oxygen control module 300 and allows the chip module 100 and the flow control module 200 to be detachably assembled, respectively, and the module port layer 400 is the first part of the chip module 100. The 1 long-term chip 110a, the 2nd long-term chip 110b, the 3rd long-term chip 110c, and the first flow control module 210a and the second flow control module 210b of the flow control module 200 are attached and detached, respectively. Possibly assembled to provide a microfluidic chip (1) of modular structure.

On the other hand, the module port layer 400 is a lower plate 410 laminated on the upper surface of the oxygen control module 300; and an upper plate 420 stacked on an upper surface of the lower plate 410.

The lower plate 410 is stacked on the upper surface of the oxygen control module 300 to form a flow path while covering the second channel 310 of the oxygen control module 300, and is opened to the side where the chip module 100 is assembled. A through hole 411 is formed, and the through hole 411 corresponds to a position where the first long-term chip 110a, the second long-term chip 110b, and the third long-term chip 110c are assembled.

The upper plate 420 is stacked on the upper surface of the lower plate 410, and the first flow control module 210a and the second flow control module 210b of the flow control module 200 are assembled on both sides of the upper plate 420, respectively. A first porthole 421 is formed, and the first long-term chip 110a, the second long-term chip 110b, and the third long-term chip 110c are assembled between the first portholes 421 on both sides, respectively. 2 portholes 422 are formed.

In this case, the top plate 420 allows the chip module 100 and the flow control module 200 to be assembled in a detachable structure through the first porthole 421 and the second porthole 422, thereby providing a module structure. .

At this time, the second port hole 422 corresponds to the first inclined portion 113 provided on the first long-term chip 110a, the second long-term chip 110b, and the third long-term chip 110c, respectively, so that the module port layer 400 ) is formed with a second inclined portion 423 extending upwardly, and the second inclined portion 423 is a surface on which the first inclined portion 113 abuts and is fitted.

For example, in the present invention according to the above structure, the oxygen concentration can be adjusted by determining the ratio of the oxygen absorbing solution and the dilution solution corresponding to the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c. It is possible to keep the oxygen concentration constant while adjusting the oxygen concentration in response to each organ chip.

In addition, according to the present invention, the shape of one or more second passages 311 for supplying the cell culture medium is formed differently in correspondence to the first organ chip 110a, the second organ chip 110b, and the third organ chip 110c. Accordingly, the flow rate of the cell culture medium and the shear stress applied to the cells can be differently adjusted according to each corresponding organ chip.

In addition, the present invention has a module structure in which the flow control module 200 and the chip module 100 are selectively combined and coupled to the module port layer 400, so that human organs under different culture conditions can be easily cultured on one microfluidic chip. You can do it.

As described above, the present invention has been expressed in the description and drawings illustrating specific preferred embodiments, but the terms used herein are for easily describing the present invention, and the meaning of these terms is limited, but the claims It is not intended to limit the scope described in

The present invention can be variously changed, modified, modified, etc. by those skilled in the art to which the present invention pertains within the scope without departing from the spirit and scope of the invention indicated by the claims according to the above embodiments. Anyone can easily see that it is possible.

One; microfluidic chip 100; chip module
110a, 110b, 110b; 1st long-term chip, 2nd long-term chip, 3rd long-term chip
111; first chamber 112; gas permeable membrane
113; a first inclined portion 200; Flow control module
210a, 210b; The first flow control module and the second flow control module
211; first channel 211a; 2nd aisle
300; oxygen control module 310; 2nd channel
311; second passage 312; 2nd chamber
400; module port 410; lower plate
420; top plate 411; through hole
421; a first port hole 422; 2nd porthole
423; 2nd slope

Claims (11)

a chip module in which cells are cultured by a cell culture medium and a gas permeable membrane is formed on a bottom surface;
a flow control module for allowing the cell culture medium to flow through the chip module;
an oxygen control module providing a hypoxic condition to the chip module while flowing an oxygen absorbing solution and a diluting solution toward the gas permeable membrane; and
A modular microfluidic chip capable of adjusting the flow rate and oxygen concentration, characterized in that it is configured to include; a module port layer that is laminated on the upper surface of the oxygen control module and allows the chip module and the flow control module to be detachably assembled, respectively. . According to claim 1,
The chip module further includes a first organ chip, a second organ chip, and a third organ chip simulating a human organ, and a modular microfluidic chip capable of controlling flow rate and oxygen concentration. According to claim 2,
The first organ chip, the second organ chip, and the third organ chip are each provided with a first chamber in which cells are cultured, and the first chamber of the first organ chip, the second organ chip, and the third organ chip are copied. A modular microfluidic chip capable of controlling the flow rate and oxygen concentration, further including those of different sizes corresponding to human organs. According to claim 2,
The first long-term chip, the second long-term chip, and the third long-term chip have a first inclined portion having a downwardly contracted circumferential surface, and a second inclined portion extending upward to the module port layer corresponding to the first inclined portion. A modular microfluidic chip capable of controlling the flow rate and oxygen concentration, further including a portion formed thereon. According to claim 1,
The flow control module includes a first flow control module into which the cell culture medium is injected; And a second flow rate control module for discharging the cell culture fluid that has passed through the chip module. According to claim 5,
The first flow control module and the second flow control module are provided with a first channel through which the cell culture medium flows, and the first channel is provided with cells in the first organ chip, the second organ chip, and the third organ chip constituting the chip module. One or more first passages for supplying the culture medium are provided, and the one or more first passages have different sizes corresponding to the human organs to be simulated. A modular microfluidic chip capable of adjusting the flow rate and oxygen concentration. According to claim 1,
The oxygen control module is a modular microfluidic chip capable of adjusting the flow rate and oxygen concentration, further including adjusting the oxygen concentration in response to the dilution rate of the diluent in the oxygen absorbing solution. According to claim 1,
The diluted solution is capable of adjusting the flow rate and oxygen concentration, further including dilution at different dilution ratios corresponding to the human organs simulated by the first organ chip, the second organ chip, and the third organ chip constituting the chip module. Modular microfluidic chip. According to claim 1,
The oxygen absorbing solution is a modular microfluidic chip capable of adjusting the flow rate and oxygen concentration, further including using a mixed solution of 0.5 g of cobalt nitrate, 25 μl of sodium sulfite, and 50 ml of DW. According to claim 1,
The oxygen control module is provided with a second channel through which an oxygen absorbing solution and a diluent are respectively injected, and the second channel is provided with one or more second passages for mixing and supplying the oxygen absorbing solution and the diluent, respectively. Modular microfluidic chip with controllable concentration. According to claim 1,
The module port layer further includes a first porthole into which the flow control module is assembled and a second porthole through which the chip module is assembled. Modular microfluidic chip capable of controlling the flow rate and oxygen concentration.