KR102577768B1 - Microfluidic Device Capable Of Trapping Cells - Google Patents

Abstract

The present invention relates to a microfluidic device capable of capturing cells, including a lower jig; an upper jig capable of horizontal movement within a predetermined range on the lower jig; a first channel layer fixed to the lower jig; and a second channel layer fixed to the upper jig and including a fluid inlet, a fluid passage connected to the fluid inlet, and a plurality of cell trapping holes formed along the longitudinal direction of the fluid passage. A plurality of cell fixation grooves connected to the plurality of cell capture holes are formed, and a microfluidic device capable of cell capture can be provided by moving the upper jig horizontally to position the cell capture holes in the cell fixation groove. You can.

Description

Microfluidic device capable of trapping cells {Microfluidic Device Capable Of Trapping Cells}

The present invention relates to a microfluidic device capable of capturing multiple single cells using fluid resistance.

Cells maintain life by performing various biological functions such as gene expression, cell growth, cell cycle, metabolism, and signal transmission through diverse and complex protein-protein interactions. In the case of research methodologies targeting cell populations used in most biological research, data are analyzed under the assumption that individual cells within the cell population produce uniform cellular responses. However, in reality, if individual cells do not produce a uniform response, result analysis based on average measured values may lead to errors. In reality, there are very few cases where it has been experimentally proven whether individual cells produce a uniform reaction due to various technical problems, so the development of technology for single cell analysis is essential.

Existing methods of using microarrays for single cell analysis include drilling a hole similar in size to the cell and allowing a single cell to enter it, and using dielectrophoresis to capture the single cell at a specific location. There was a way. In the case of dielectric electrophoresis, there is a problem in that when the applied voltage is cut off, the cells move to an undesirable location.

In addition, there is a method called electrorotation to measure the dielectric properties of a single cell, which measures the rotation rate by applying a rotating alternating current signal with a phase difference of 360°/n to n electrodes and based on the single-shell dielectric model. This is a method of deriving the dielectric properties of a cell by comparing and analyzing the rotational speed of the cell with the spectral theoretical results.

Registered Patent Publication No. 10-1996847 (2019.07.01.)

Most conventional microfluidic devices have complex fluid path structures and have the problem of using multiple valves and separate systems to control the flow of fluid.

Accordingly, the present invention provides a microfluidic device that can capture multiple single cells using fluid resistance without multiple valves in order to minimize manual work to capture single cells and increase the amount of cells captured for analysis. The purpose is to

In order to achieve the above purpose, a lower jig; an upper jig capable of horizontal movement within a predetermined range on the lower jig; a first channel layer fixed to the lower jig; and a second channel layer fixed to the upper jig and including a fluid inlet, a fluid passage connected to the fluid inlet, and a plurality of cell trapping holes formed along the longitudinal direction of the fluid passage. A plurality of cell fixing grooves connected to the plurality of cell capture holes are formed, and the cell capture hole can be positioned in the cell fixation groove by horizontally moving the upper jig. A microfluidic device capable of cell capture is provided.

In addition, the microfluidic device capable of capturing cells according to the present invention further includes a pressing part that presses the upper jig, and the pressing part includes a pressure plate, a connecting member connecting the pressure plate and the lower jig, and a connection member disposed on one surface of the pressure plate to press the upper jig. Provided is a microfluidic device capable of cell capture that includes one or more pressurizing members that apply pressure to a jig.

In addition, the microfluidic device capable of cell capture according to the present invention further includes a manipulation unit capable of horizontally moving the upper jig.

In addition, the manipulation unit provides a microfluidic device capable of capturing cells, including a first manipulation unit that slides the upper jig in a first direction and a second manipulation unit that slides the upper jig in a direction perpendicular to the first direction.

In addition, a microfluidic device capable of capturing cells is provided in which a plurality of first through holes are formed in the upper channel layer to match the arrangement of the plurality of cell capture holes.

In addition, a microfluidic device capable of capturing cells is provided in which a plurality of second through holes matching the arrangement of the first through holes are formed in the upper jig.

The microfluidic device capable of cell capture according to the present invention can capture multiple single cells at once, minimizing manual work by operators and improving processing speed and throughput for analyzing cells.

In addition, the configuration is relatively simple as it is a capture technology using fluid resistance without the need for multiple valves.

In addition, after capturing cells, the upper jig can be moved to create an open space, and the genome of a single cell can be extracted immediately, improving the processing speed of the work.

Figure 1 shows a perspective view of a microfluidic device capable of capturing cells according to the present invention.
Figure 2 shows a perspective view of the lower jig of the present invention.
Figure 3 shows the shape of the upper jig of the present invention in a perspective view.
Figure 4 shows the configuration of the first channel layer of the present invention.
Figure 5 shows the configuration of the second channel layer of the present invention.
Figure 6 is a photograph showing the cell fixing hole of the first channel layer of the present invention aligned with the cell passing hole of the second channel layer and the appearance of a single cell captured in the cell fixing hole.
Figure 7 shows a state in which the second channel layer of the present invention is moved horizontally and the cell fixing hole of the first channel layer is positioned in the through hole of the second channel layer.
Figure 8 shows a perspective view of the connecting member of the present invention.
Figure 9 is a side view of a microfluidic device capable of cell capture according to the present invention.
Figure 10 is a photograph showing the actual appearance of a microfluidic device capable of cell capture according to the present invention.

The invention described below may be subject to various changes and may have various embodiments, and specific embodiments will be illustrated in the drawings and described in detail. However, this is not intended to limit the invention described below to specific embodiments, and should be understood to include all changes, equivalents, and substitutes included in the spirit and scope of the technology described below.

1 to 10 show data explaining the microfluidic device capable of capturing cells according to the present invention. Hereinafter, the present invention will be described in more detail with reference to the accompanying drawings to aid understanding of the present invention. However, the following examples are provided only to make the present invention easier to understand, and the content of the present invention is not limited by the following examples.

Figure 1 schematically shows a microfluidic device capable of cell capture according to the present invention. Referring to FIG. 1, the microfluidic device 10 capable of capturing cells includes a lower jig 100, an upper jig 200, a first channel layer 300, an upper channel layer 400, and a pressurizing unit 500. ) and an operating unit 600.

When described with reference to FIG. 2, the lower jig 100 may be formed with a flow groove 110 and a first channel layer groove 120. The position of the lower jig 100 is fixed, and the upper jig 100 can move horizontally in the flow groove 110 of the lower jig 100. The horizontal cross-sectional shape of the flow groove 110 may be a square shape or a square with rounded corners, and a side wall 111 is formed around the flow groove 110 to limit the horizontal movement range of the upper jig 100. .

The first channel layer groove 120 may be a groove formed in the flow groove 100 into which the first channel layer 300 is inserted and fixed. The first channel layer 300 may be smaller than the horizontal cross-sectional area of the flow groove 110. The horizontal cross-sectional shape of the first channel layer 300 may be a square shape or a square shape with rounded corners. The first channel layer groove 120 is the same or similar to the area of the horizontal cross section of the first channel layer 300, so that when the first channel layer 300 is inserted into the first channel layer groove 120, the lower jig does not flow. It can be fixed at (100).

3, the upper jig 200 horizontally moves the inner space of the flow groove 110 and horizontally moves the second channel layer 400, and the second channel layer 400 and the first channel layer ( 400) may be a member for adjusting the relative position. A fixing wall 210 may be protruding from one surface of the upper jig 200. The fixing wall 210 may be formed to protrude along the perimeter of the second channel layer so that the second channel layer 400 can be inserted and fixed. The fixing wall 210 may limit the horizontal movement of the upper jig 200 by the side wall 111 when the upper jig 200 moves horizontally.

A plurality of connection holes 220, 230, and 240 connected to the second channel layer 400 may be formed in the upper jig 200. The first connection hole 220 is a hole connected to the fluid inlet 410, the second connection hole 230 is a hole connected to the fluid outlet 440, and the third connection hole 240 is a through hole 450. ) may be a hole connected to

In particular, the third connection hole 240 is connected to communicate with the through hole 450 to capture cells in the cell fixing hole 310 of the first channel layer 300, and then move the upper jig 200 horizontally to capture cells. The third connection hole 240 and the through hole 450 are located on the fixing hole 310 to secure an open partition space, and then the next dielectric extraction operation can be performed immediately. The arrangement and number of the plurality of third connection holes 240 may match the arrangement and number of the plurality of through holes 450.

Referring to FIG. 4, the second channel layer 400 is a member inserted and fixed into the internal space formed by the fixing wall 210, and includes a fluid inlet 410 through which fluid containing cells flows, and a fluid inlet. A fluid passage 420 through which the fluid introduced through 410 can flow, a plurality of cell passage holes 430 formed at predetermined intervals along the longitudinal direction on one side of the fluid passage 420, and a fluid remaining after the cells are captured. A fluid outlet 440 through which fluid flows out and a plurality of through holes 450 may be formed.

The cell passage hole 430 may be a hole for capturing cells included in the introduced fluid. A plurality of cell passage holes 430 may be arranged along the longitudinal direction on one side of the fluid passage 420. The diameter of the cell passage hole 430 may be the same or similar to the size of the cell. As the fluid flows through the fluid flow path 420, a single cell may pass through the cell passage hole 430. At this time, the cell passing through the cell passage hole 430 may be connected to the cell fixation hole 310 of the first channel layer 300. It may flow in and block the cell passage hole 430. If the cell passage hole 430 is clogged with a single cell, fluid resistance occurs, and the fluid can flow to the other cell passage hole 430 adjacent to the cell passage hole 430, and the other adjacent cell passage hole 430 is also operated in the same manner. Cells can be captured.

A plurality of cell passage holes 430 may be arranged in a row at regular intervals along the fluid passage 420.

A fluid outlet 440 may be formed at the end of the fluid passage 420 to allow cells to be captured and the remaining fluid to be discharged.

5 and 6, the first channel layer 300 is in close contact with the second channel layer 400 and forms a layer, capturing single cells that have passed through the second channel layer 400. It could be a place to do it. A cell anchoring hole 310 may be formed in the first channel layer 300. The cell fixation holes 310 can be formed in the same number and arrangement as the cell passage holes 430. The distance between the plurality of cell passage holes 430 may also be identical to the cell fixation hole 310. By horizontally moving the upper jig 200, the position of the first channel layer 300 can be manipulated so that the cell passage hole 430 and the cell fixation hole 310 are connected.

Referring to FIG. 4 , a plurality of cell passage holes 430 may be formed in the second channel layer 400 and a plurality of first through holes 440 may be formed at positions spaced apart from each other by a predetermined distance. The plurality of first through-holes 440 may be formed in the same number as the plurality of cell through-holes 430, and the arrangement of the plurality of through-holes 440 is consistent with the arrangement of the plurality of cell through-holes 430. can do. That is, the number and arrangement of the cell passing holes 430, cell fixing holes 310, and first through holes 440 may be identical.

Referring to FIG. 7 , when a cell is captured in the cell fixing hole 310 of the first channel layer 300, the upper jig 200 is horizontally moved to open a plurality of through holes 440 in the cell fixing hole 310. ), the cells can be exposed, and the genome can be manually extracted from the exposed cells.

When a single cell is captured in the cell fixing hole 310, fluid can no longer pass through the cell fixing hole 310 due to fluid resistance. When fluid resistance occurs, the fluid flows through the cell fixing hole 310 and the cell passing hole 430 to another adjacent cell fixing hole 310 and the cell passing hole 430, and flows to the adjacent cell fixing hole and the cell passing hole. Another single cell can be captured at (430). In this way, a single cell can be captured in the cell fixation hole 310.

A cell anchoring hole 310, a first guide 320, and a second guide 330 may be formed in the second channel layer 400. The first guide 320 is connected to the cell passing hole 430 and can guide a single cell to the cell fixing hole 310.

The second guide 330 may guide the fluid that has passed through the cell fixation hole 310. When cells are captured in the cell fixing hole 310, the cells block the cell processing hole 310, so the second guide 330 can no longer guide fluid.

It is important that the first channel layer 300 and the second channel layer 400 are in close contact to prevent fluid from leaking through the gap between the first channel layer 300 and the second channel layer 400. Therefore, the microfluidic device capable of capturing cells according to the present invention includes a pressurizing portion 500 at the top of the upper jig 200 to pressurize the upper jig 200. When the upper jig 200 is pressed, the first channel layer 300 and the second channel layer 400 come into closer contact, thereby minimizing fluid leakage.

When described with reference to FIG. 9 , the pressing unit 500 may include a pressing plate 510, a connecting member 520, and a pressing member 530. The pressing unit 500 may be a place that presses the upper surface of the upper jig 200 to bring the first channel layer 300 and the second channel layer 400 into close contact.

The pressure plate 510 is connected to the lower jig 100 and the connecting member 520 and can be positioned at a certain distance upward from the upper jig 200. The pressure plate 510 supports a plurality of pressure members 530 and may be a place where the plurality of pressure members 530 uniformly apply pressure to the upper jig 200. The pressure plate 510 may be located on a plurality of connecting members 520.

1 and 8, the connecting member 520 is located at the center of each side of the lower jig 100 and can be coupled to the lower jig 100, and the plurality of connecting members 520 are connected to a pressure plate ( It may be a member to distribute and support the weight of 510 and to connect the lower jig 100 and the pressure plate 510. The connecting member 520 may be formed in a ‘ㄷ’ shape.

An upper jig coupling hole 521, an operation unit coupling hole 522, and a lower jig coupling hole 523 may be formed in the connecting member 520.

The pressing member 530 may have one end coupled to the pressing plate 510 and the other end pressing the upper jig 200. A plurality of pressing members 530 are provided to uniformly pressurize the upper jig 200, and the plurality of pressing members 530 may be arranged in positions symmetrical to each other. It is preferable that the plurality of pressing members 530 be in an even number so that they can be disposed in symmetrical positions and apply uniform pressure.

A ball roller 531 may be provided at the end of the pressing member 530. Since the ball roller 531 includes a spherical ball, when the upper jig 200 moves in the horizontal direction, the ball of the ball roller 531 is in rolling contact with the upper jig 200, so the ball is in contact with the upper jig 200. ) can roll, so the upper jig 200 can move precisely because only rolling friction, which is much less resistant than the sliding friction generated when in contact with the ball, is generated.

Since the channels formed in the first channel layer 300 and the second channel layer 400 are only a few micrometers in size, the arrangement of the cell passing hole 430 and the cell fixing hole 310 must be aligned or the cell fixing hole ( Fine manipulation is required to place 310) in the first through hole. The manipulation unit 600 may enable fine movement of the second channel layer 400 by slightly horizontally moving the upper jig 200.

The manipulation unit 600 is for horizontally moving the upper jig 200, and includes a first manipulation unit that horizontally moves the upper jig 200 in a first direction and a second manipulation unit that horizontally moves the upper jig 200 in a direction perpendicular to the first direction. It can be included.

The manipulation unit 600 may include a rotation handle 610, a screw member 620 connected to the rotation handle 610, and a push member 630 connected to the screw member. When the rotation handle 610 is rotated, the screw member 620 can also rotate and move forward or backward, and the push member 630 connected to the screw member 620 can also move forward or backward. On the outer surface of the screw member 620, a screw thread may be formed in a spiral shape in the longitudinal direction of the screw member 620, and one rotation of the screw member 620 can advance or retreat by the distance between the screw thread and other adjacent screw threads. So, you can finely adjust the distance.

The manipulation unit 600 can perform finer manipulation as the length of the cross-sectional circumference of the screw member 620 increases compared to the distance between the screw threads.

There may be a plurality of operating units 600, and each operating unit 600 is coupled to each connecting member 520 by penetrating through the connecting member 520, and each operating unit 620 is connected to each side of the upper jig 200. The upper jig 200 can be moved horizontally by pushing the center.

The push member 630 may have a flat end so that it can accurately push the center of the side of the upper jig 200, and a buffer member is attached to the end of the push member 630 to strengthen adhesion to the upper jig 200. It can be.

Figure 10 shows the actual appearance of a microfluidic device capable of cell capture according to the present invention.

Although the present technology has been described through the above examples, the present technology is not limited thereto. The above embodiments may be modified or changed without departing from the spirit and scope of the present technology, and those skilled in the art will recognize that such modifications and changes also belong to the present technology.

10: Microfluidic device 100: Lower jig
110: flow groove 120: first channel layer groove
200: upper jig 210: fixed wall
220: first connection hole 230: second connection hole
240: third connection hole 300: first channel layer
310: Cell fixation hole 400: Second channel layer
410: fluid inlet 420: fluid flow path
430: cell passage hole 440: fluid outlet
450: through hole 500: pressurized part
510: pressure plate 520: connection member
530: Pressure member 600: Control unit
610: Rotation handle 620: Screw member
630: Push member

Claims (6)

Lower jig;
an upper jig capable of horizontal movement within a predetermined range on the lower jig;
a first channel layer fixed to the lower jig; and
A second channel layer fixed to the upper jig, the second channel layer including a fluid inlet, a fluid passage connected to the fluid inlet, and a plurality of cell passage holes formed along the longitudinal direction in the fluid passage. ,
A plurality of cell anchoring holes in which cells are captured are formed in the first channel layer,
A microfluidic device capable of capturing cells that can move the upper jig horizontally to position the cell passage hole in the cell fixation hole. According to claim 1,
The microfluidic device capable of capturing cells according to the present invention further includes a pressurizing part that pressurizes the upper jig,
The pressing unit includes a pressing plate, a connecting member connecting the pressing plate and the lower jig, and one or more pressing members disposed on one surface of the pressing plate to apply pressure to the upper jig. A microfluidic device capable of capturing cells. According to claim 1,
The microfluidic device capable of cell capture according to the present invention further includes a manipulation unit capable of horizontally moving the upper jig. According to claim 3,
The manipulation unit is a microfluidic device capable of cell capture including a first manipulation unit that horizontally moves the upper jig in a first direction and a second manipulation unit that horizontally moves the upper jig in a direction perpendicular to the first direction. According to claim 1,
A microfluidic device capable of capturing cells, wherein a plurality of first through holes are formed in the second channel layer to match the arrangement of the plurality of cell through holes. According to claim 5,
A microfluidic device capable of capturing cells in which a plurality of connection holes are formed in the upper jig that matches the arrangement of the first through hole.