CN107603850A - Micro fluidic device for cell sorting and preparation method thereof - Google Patents

Abstract

The invention provides a microfluidic device for cell sorting and a preparation method thereof, and relates to the technical field of cell sorting. The microfluidic device for cell sorting includes a lower base body and an upper base connected to the lower base body. The substrate, the lower substrate is provided with a microcolumn array for sorting cells; along the direction of cell flow, the angle between the arrangement direction of the columns in the microcolumn array and the direction of cell flow is 2°-10°; along the direction perpendicular to the direction of cell flow, the microcolumn The spacing between the columns in the column array is 8-13 microns, and the microfluidic device can alleviate the technical problems of complex structure and high equipment cost of the cell sorting device in the prior art.

Description

Microfluidic device for cell sorting and preparation method thereof

technical field

The invention relates to the technical field of cell sorting, in particular to a microfluidic device for cell sorting and a preparation method thereof.

Background technique

Cell sorting is a necessary means to obtain target cells with uniform properties from complex environments. Cell sorting is a key link in the study of cell physiology and pathology, and it is of great significance for the diagnosis and treatment of major diseases. The research on cell sorting methods and related technologies has always been a research hotspot in the medical field and related fields.

The existing cell sorting methods mainly use different fluorescently labeled cells for sorting with a flow cytometer with a sorting function. Flow cytometry uses high-energy laser light to irradiate single cells or particles stained with fluorescent pigments under high-speed flow, and measures the intensity of scattered light and emitted fluorescence to separate cells. However, the flow cytometer with sorting function is bulky, complex in structure, expensive, and the processing method of cells is cumbersome, and the experimental cost is high.

Contents of the invention

The first object of the present invention is to provide a microfluidic device for cell sorting, so as to alleviate the technical problems of complex structure and high equipment cost of the cell sorting device in the prior art.

The second object of the present invention is to provide a method for preparing a microfluidic device for cell sorting, which has the advantage of low manufacturing cost.

In order to realize the above-mentioned purpose of the present invention, special adopt following technical scheme:

A microfluidic device for cell sorting, comprising a lower substrate and an upper substrate connected to the lower substrate, the lower substrate is provided with a microcolumn array for sorting cells; along the cell flow direction, the microcolumns The included angle between the arrangement direction of the pillars in the pillar array and the cell flow direction is 2°-10°; along the direction perpendicular to the cell flow direction, the spacing between the pillars in the micro-pillar array is 8-13 microns.

Further, along the cell flow direction, the angle between the column arrangement direction in the micropillar array and the cell flow direction is 2°-9°, more preferably 2°-8°; along the direction perpendicular to the cell flow direction, the The spacing between the pillars in the micropillar array is 8-12 microns, more preferably 10-12 microns.

Further, the pillars in the micropillar array include triangular pillars, circular pillars or trapezoidal pillars.

Further, the triangular column is an isosceles triangular column; preferably, the isosceles triangular column is a right-angled isosceles triangular column or an obtuse-angled isosceles triangular column.

Further, when the cells flow along the cell flow direction, they collide with the side where the base of the isosceles triangular column is located.

Further, along the direction perpendicular to the flow direction of the cells, magnetic control devices for applying a magnetic field to the cells are provided on both sides of the lower base.

Further, electrodes for applying an electric field to the cells are provided on both sides of the lower substrate along a direction perpendicular to the flow direction of the cells.

Further, along the direction of cell flow, one end of the lower substrate is provided with a first injection port for injecting cells and a second injection port for injecting medium;

The other end of the lower base body is provided with sample outlets, and the sample outlets are not less than two and distributed along a direction perpendicular to the cell flow direction.

A preparation method of the above-mentioned microfluidic device for cell sorting includes firstly preparing a lower substrate by a casting method, and then connecting the lower substrate with an upper substrate to obtain the microfluidic device for cell sorting.

Further, the casting method includes pouring liquid raw materials into a mold, and separating to obtain the lower matrix after solidification and molding;

Preferably, the liquid raw material includes a prepolymer composed of a curing agent and PDMS;

Preferably, the weight ratio of the curing agent to the PDMS is 1: (8-12);

Preferably, after pouring, first perform degassing treatment and then solidify and form;

Preferably, the temperature during the curing molding process is 80-100° C., and the time is 1.5-2.5 hours.

Further, the preparation method of the mold includes the following steps: coating a photoresist on the surface of the mold base, and obtaining a cavity structure corresponding to the micro-column array in the lower base after exposure and development;

Preferably, the coated photoresist has a thickness of 8-14 microns.

Compared with the prior art, the present invention has the following beneficial effects:

In the microfluidic device used for cell sorting provided by the present invention, since there is a certain angle between the arrangement direction of the columns in the microcolumn array and the cell flow direction, the angle and distance at which cells of different sizes are bounced when passing through the microcolumn array All of them are different. When the flow rate is constant, the rebound angle and distance of large-sized cells will be larger. The cells are bounced back and the space between the pillars can be used to further increase the flow difference between the large and small cells (including the flow angle and the offset distance from the incident direction of the cells), so in the present invention, the arrangement of the pillars in the microcolumn array Orientation and alignment spacing allow cells to decisively choose flow paths according to their size, enabling the sorting of cells of various sizes ingeniously.

Compared with the existing flow cytometer, the microfluidic device provided by the present invention has the advantages of simple structure, low cost, and the sorting of cells can be realized without fluorescent treatment of cells.

Description of drawings

In order to more clearly illustrate the specific implementation of the present invention or the technical solutions in the prior art, the following will briefly introduce the accompanying drawings that need to be used in the specific implementation or description of the prior art. Obviously, the accompanying drawings in the following description The drawings show some implementations of the present invention, and those skilled in the art can obtain other drawings based on these drawings without any creative work.

FIG. 1 is a schematic structural diagram of a microfluidic device for cell sorting provided in Example 1 of the present invention;

2 is a schematic structural diagram of a microfluidic device for cell sorting provided in Example 2 of the present invention;

3 is a schematic structural diagram of a microfluidic device for cell sorting provided in Example 3 of the present invention;

Fig. 4 is a schematic structural diagram of a microfluidic device for cell sorting provided by Example 4 of the present invention.

Icons: 10-lower substrate; 11-microcolumn array; 12-column; 13-first injection port; 14-second injection port; 15-sample outlet; 20-magnetic control device; 30-electrode.

Detailed ways

The technical solutions of the present invention will be clearly and completely described below in conjunction with the accompanying drawings. Apparently, the described embodiments are some of the embodiments of the present invention, but not all of them. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the protection scope of the present invention.

In the description of the present invention, it should be noted that the terms "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inner", "outer" etc. The indicated orientation or positional relationship is based on the orientation or positional relationship shown in the drawings, and is only for the convenience of describing the present invention and simplifying the description, rather than indicating or implying that the referred device or element must have a specific orientation, or in a specific orientation. construction and operation, therefore, should not be construed as limiting the invention. In addition, the terms "first", "second", and "third" are used for descriptive purposes only, and should not be construed as indicating or implying relative importance.

In the description of the present invention, it should be noted that unless otherwise specified and limited, the terms "installation", "connection" and "connection" should be understood in a broad sense, for example, it can be a fixed connection or a detachable connection. Connected, or integrally connected; it can be mechanically connected or electrically connected; it can be directly connected or indirectly connected through an intermediary, and it can be the internal communication of two components. Those of ordinary skill in the art can understand the specific meanings of the above terms in the present invention according to specific situations.

One aspect of the present invention provides a microfluidic device for cell sorting, comprising a lower base and an upper base connected to the lower base, the lower base is provided with a microcolumn array for sorting cells; Along the cell flow direction, the angle between the arrangement direction of the columns in the microcolumn array and the cell flow direction is 2°-10°; along the direction perpendicular to the cell flow direction, the spacing between the columns in the microcolumn array is 8-13 microns.

In the microfluidic device used for cell sorting provided by the present invention, since there is a certain angle between the arrangement direction of the columns in the microcolumn array and the cell flow direction, the angle and distance at which cells of different sizes are bounced when passing through the microcolumn array All of them are different. When the flow rate is constant, the rebound angle and distance of large-sized cells will be larger. The cells are bounced back and the space between the pillars can be used to further increase the flow difference between the large and small cells (including the flow angle and the offset distance from the incident direction of the cells), so in the present invention, the arrangement of the pillars in the microcolumn array Orientation and alignment spacing allow cells to decisively choose flow paths according to their size, enabling the sorting of cells of various sizes ingeniously.

In addition, compared with the existing flow cytometer, the microfluidic device provided by the present invention has the advantages of simple structure, low cost, and the ability to sort cells without fluorescent treatment of cells.

The middle part of the lower substrate is a sorting area composed of microcolumn arrays, wherein the microcolumn array is composed of columns arranged in horizontal and vertical directions, the vertical direction represents the flow direction of cells, and the horizontal direction is the direction perpendicular to the flow direction of cells. And the height direction of the column represents the depth scale of the sorting area.

The microcolumn array in the present invention can be one group or multiple groups, and the sorting accuracy can be further improved when there are multiple groups.

The angle between the arrangement direction of the pillars in the micropillar array and the flow direction of the cells is in the range of 2°-10°. When cells of different sizes need to be sorted, the angle can be set to different sizes. In addition, when there are multiple sets of micropillar arrays, different sets of micropillar arrays can also be set to have different included angles, and different included angles can also be set in the same set of micropillar arrays, which can further increase the separation effect.

Similarly, the setting range of the spacing between columns in the micropillar array is 8-13 microns, and when cells of different sizes need to be sorted, the spacing can be set to different sizes. In addition, when there are multiple sets of micropillar arrays, different sets of micropillar arrays can also be set at different pitches, and the same set of micropillar arrays can also be set with different pitches, which can further increase the separation effect.

The lower substrate is preferably made of PDMS material, while the upper substrate is preferably made of glass sheet, and the two are preferably combined by bonding to form a microfluidic device for cell sorting.

In the present invention, the included angle between the column arrangement direction and the cell flow direction is typically but not limited to, for example: 2°, 3°, 4°, 5°, 6°, 7°, 8°, 9° or 10° °; Typical but non-limiting examples of the spacing between the pillars in the micropillar array are: 8 microns, 9 microns, 10 microns, 11 microns, 12 microns or 13 microns.

As a preferred embodiment of the present invention, along the cell flow direction, the angle between the column arrangement direction in the microcolumn array and the cell flow direction is 2°-9°, more preferably 2°-8°; In the direction of cell flow, the spacing between columns in the microcolumn array is 8-12 microns, more preferably 10-12 microns. By further optimizing the setting angle and spacing, the sorting accuracy can be improved.

As a preferred embodiment of the present invention, the pillars in the micropillar array include triangular pillars, circular pillars or trapezoidal pillars. The pillars in the micropillar array can be pillars of various shapes and structures, and other shapes besides the above-mentioned shapes can also be used.

As a further preferred embodiment of the present invention, the triangular column is an isosceles triangular column; further preferably, the isosceles triangular column is a right-angled isosceles triangular column or an obtuse-angled isosceles triangular column. With an isosceles triangular column, cells can collide and launch on each side as they flow towards the column. The use of right-angled isosceles triangles or obtuse-angled isosceles triangles can reduce the secondary reflection of cells between the columns, so that they can pass between the two columns smoothly after a collision and reflection. At the same time, the use of right-angled isosceles triangles is also convenient for molds. preparation.

As a preferred embodiment of the present invention, when the cells flow along the cell flow direction, they collide with the side where the base of the isosceles triangular column is located. At this time, the side of the base of the isosceles triangular column is set against the flow direction of the cells, so that the cells flow to the column and collide with the side of the base and reflect them.

When an isosceles triangular column is used, the side where the bottom edge is located forms an angle of 45° with the cell flow direction, so that the emission of cells is more regular.

As a preferred embodiment of the present invention, along the direction perpendicular to the flow direction of the cells, magnetic control devices for applying a magnetic field to the cells are provided on both sides of the lower base. In a preferred mode of the present invention, the magnetic control device includes magnetic rods arranged on both sides of the lower substrate, and energizing the magnetic rods can generate a magnetic field, and the generated magnetic field can control the cells to deviate from the flow direction of the cells. At this time, cells of different sizes or different types are subjected to different magnetic forces, and the deviation effect produced by cells with a large force is more obvious. Therefore, the effect of further separating cells can be achieved by increasing the magnetic field.

As a preferred embodiment of the present invention, electrodes for applying an electric field to the cells are provided on both sides of the lower substrate along a direction perpendicular to the flow direction of the cells. When the cells to be separated contain charged cells, the charged cells in the cells can be separated by applying electricity to the cells.

As a preferred embodiment of the present invention, along the cell flow direction, one end of the lower base is provided with a first injection port for injecting cells and a second injection port for injecting medium; the other end of the lower base is provided with a sample There are no less than two sample outlets and they are distributed along the direction perpendicular to the flow direction of the cells.

The cell sample inlet is used to connect to the cell tank, the medium injection port is used to connect to the medium tank tank, and the sample outlet is used to connect to the cell collection tank. Because cells of different sizes or types can be obtained after cell separation, multiple sample outlets can be set. , and the sample outlets are distributed along the direction perpendicular to the cell flow direction, that is, the direction in which the columns in the microcolumn array are arranged laterally.

Another aspect of the present invention provides a method for preparing the above-mentioned microfluidic device for cell sorting. Firstly, the lower substrate is prepared by casting method, and then the lower substrate is bonded with the upper substrate to obtain the microfluidic device for cell sorting. Microfluidic devices for cell sorting.

The lower substrate in the present invention is prepared by a casting method, and the process is simple and easy to realize. After the lower substrate is obtained, it can be connected with the upper substrate to obtain the above-mentioned microfluidic device for cell sorting. In the present invention, the connection between the lower substrate and the upper substrate is preferably bonded.

As a preferred embodiment of the present invention, the casting method includes pouring a liquid raw material into a mold, and separating to obtain the lower substrate after curing and molding; optionally, the liquid raw material includes a curing agent and polydimethylsiloxane A prepolymer composed of polydimethylsiloxane (Polydimethylsiloxane, PDMS for short); optionally, the weight ratio of the curing agent to the PDMS is 1:(8-12). In a preferred embodiment of the present invention, the lower substrate is made of PDMS material, and the upper substrate is flat glass.

In the above-mentioned preferred embodiment, the weight ratio of the curing agent to the PDMS is typically but not limited to, for example, 1:8, 1:9, 1:10, 1:11 or 1:12; wherein the The curing agent is preferably a dealcohol-type curing agent or a deacidification-type curing agent.

As a preferred embodiment of the present invention, after pouring, degassing treatment is performed before solidification and molding; more preferably, the temperature during the solidification and molding process is 80-100° C., and the time is 1.5-2.5 hours. The first degassing treatment can eliminate the air bubbles in the PDMS, and ensure that there are no residual air bubbles in the lower matrix, so as to prevent the air bubbles from adversely affecting the separation of cells.

In the above preferred embodiment, the typical but non-limiting temperature during the curing molding process is, for example: 80°C, 85°C, 90°C, 95°C or 100°C; the typical but non-limiting time during the curing molding process is For example: 1.5h, 1.7h, 2.0h, 2.2h or 2.5h.

As a preferred embodiment of the present invention, the preparation method of the mold includes the following steps: coating a photoresist on the surface of the mold base, and obtaining a cavity structure corresponding to the lower base structure after exposure and development; optionally, coating The thickness of the overlying photoresist is 8-14 microns. In a preferred embodiment of the present invention, the mold is obtained by coating photoresist on a silicon wafer, exposing and developing, and the method is simple in operation and easy to implement.

In the above preferred embodiments, the thickness of the coated photoresist is typically but not limited to, for example: 8 microns, 10 microns, 11 microns, 12 microns, 13 microns or 14 microns.

As a preferred embodiment of the present invention, after the lower substrate is fabricated, use a puncher to punch holes in the lower substrate to prepare cell sample inlets, medium injection ports and sample outlets, and finally bond with the upper substrate to obtain the microfluidic device.

The present invention will be described in further detail below in conjunction with examples, comparative examples and accompanying drawings.

Example 1

As shown in Figure 1, the present embodiment is a microfluidic device for cell sorting, including a lower substrate 10 and an upper substrate connected to the lower substrate 10, the upper substrate is a glass slide, and the lower substrate 10 is provided with a For the microcolumn array 11 for sorting cells, along the cell flow direction, the angle between the arrangement direction of the columns 12 in the microcolumn array 11 and the cell flow direction is 2.8°; along the direction perpendicular to the cell flow direction, the column 12 in the microcolumn array 11 The spacing between them is 10 microns. The micropillar array 11 in this embodiment is a group, and the pillars 12 in the micropillar array 11 are circular pillars. Along the cell flow direction, one end of the lower base body 10 is provided with a cell sample inlet 13 and a medium injection port 14, the cell sample inlet 13 is used to connect the cell tank, and the medium injection port 14 is used to connect the medium tank; the other end of the lower base body 10 One end is provided with a sample outlet 15, and the sample outlet 15 is used to connect three cell collection tanks, and there are no less than two sample outlets 15, which are distributed along the direction perpendicular to the cell flow direction.

Wherein, the dotted lines between the pillars 12 in FIG. 1 show the arrangement direction of the pillars 12 in the micropillar array 11 .

Example 2

This embodiment is a microfluidic device for cell sorting. The difference from Embodiment 1 is that along the direction of cell flow, the angle between the arrangement direction of the columns in the microcolumn array and the direction of cell flow is 9°; In the direction of cell flow, the distance between the pillars in the micropillar array is 12 micrometers.

Example 3

This embodiment is a microfluidic device for cell sorting. The difference from Embodiment 1 is that, along the cell flow direction, the angle between the arrangement direction of the columns in the microcolumn array and the cell flow direction is 10°; In the direction of cell flow, the distance between the pillars in the micropillar array is 13 microns.

Example 4

As shown in Figure 2, the present embodiment is a microfluidic device for cell sorting, including a lower substrate 10 and an upper substrate connected to the lower substrate 10, and the lower substrate 10 is provided with micropillars for sorting cells Array 11, along the cell flow direction, the angle between the arrangement direction of the columns 12 in the microcolumn array 11 and the cell flow direction is 5.6°; along the direction perpendicular to the cell flow direction, the spacing between the columns 12 in the microcolumn array 11 is 10 microns . The micropillar array 11 in this embodiment is a group, and the columns 12 in the micropillar array 11 are right-angled isosceles triangular columns. Along the cell flow direction, one end of the lower substrate 10 is provided with a cell sample inlet 13 and a medium injection port 14, the cell sample inlet 13 is used to connect the cell tank, and the medium injection port 14 is used to connect the medium tank; The other end is provided with a sample outlet 15, the sample outlet 15 is used to connect 3 cell collection tanks, there are no less than two sample outlets 15 and they are distributed along the direction perpendicular to the cell flow direction. Wherein, the dotted lines between the pillars 12 in FIG. 2 show the arrangement direction of the pillars 12 in the micropillar array 11 .

Example 5

As shown in Figure 3, this embodiment is a microfluidic device for cell sorting. The difference from Embodiment 2 is that there are three groups of microcolumn arrays 11 in this embodiment, and the first group of microcolumn arrays The angle between the arrangement direction of the column 12 in 11 and the cell flow direction is 2.8°, the arrangement direction of the column 12 in the second group and the third group of micro-pillar arrays 11 and the angle between the cell flow direction are 5.6°; In the direction of cell flow, the distance between the pillars 12 in the first group of micropillar arrays 11 is 10 microns, and the distance between the pillars 12 in the second and third groups of micropillar arrays 11 is both 12 micrometers. Wherein, the dotted lines between the pillars 12 in FIG. 3 show the arrangement direction of the pillars 12 in the micropillar array 11 .

Example 6

As shown in Figure 4, this embodiment is a microfluidic device for cell sorting. The difference from Embodiment 3 is that in this embodiment, along the direction perpendicular to the direction of cell flow, the two sides of the lower substrate 10 are provided with The magnetic control device 20 for applying a magnetic field to the cells and the electrode 30 for applying an electric field to the cells. Wherein, the dotted lines between the pillars 12 in FIG. 4 show the arrangement direction of the pillars 12 in the micropillar array 11 .

Comparative example 1

This comparative example is a microfluidic device for cell sorting. The difference from Example 1 is that along the direction of cell flow, the angle between the arrangement direction of the columns in the microcolumn array and the direction of cell flow is 1°; In the direction of cell flow, the distance between the pillars in the micropillar array is 3 microns.

Comparative example 2

This comparative example is a microfluidic device for cell sorting. The difference from Comparative Example 1 is that along the direction of cell flow, the angle between the arrangement direction of the columns in the microcolumn array and the direction of cell flow is 1°; In the direction of cell flow, the distance between the pillars in the micropillar array is 10 microns.

Comparative example 3

This comparative example is a microfluidic device for cell sorting. The difference from Comparative Example 1 is that along the direction of cell flow, the angle between the arrangement direction of the columns in the microcolumn array and the direction of cell flow is 2.8°; In the direction of cell flow, the distance between the pillars in the micropillar array is 3 microns.

Comparative example 4

This comparative example is a microfluidic device for cell sorting. The difference from Example 4 is that along the direction of cell flow, the angle between the arrangement direction of the columns in the microcolumn array and the direction of cell flow is 12°; In the direction of cell flow, the distance between the pillars in the micropillar array is 15 micrometers.

Comparative example 5

This comparative example is a microfluidic device for cell sorting. The difference from Example 5 is that, along the cell flow direction, the angle between the arrangement direction of the columns in the first group of microcolumn arrays and the cell flow direction is 1.5° , the angles between the arrangement direction of the pillars in the second group and the third group of micropillar arrays and the cell flow direction are both 12°; along the direction perpendicular to the cell flow direction, the spacing between the pillars in the first group of micropillar arrays is 10° micron, the spacing between the columns in the second group and the third group of micro-pillar arrays is 12 microns.

Comparative example 6

This comparative example is a microfluidic device for cell sorting. The difference from Example 6 is that the angle between the arrangement direction of the pillars in the first group of micropillar arrays and the direction of cell flow is 1°, and the second group The included angles between the arrangement direction of the pillars in the third group of micropillar arrays and the cell flow direction are both 1.2°; along the direction perpendicular to the cell flow direction, the spacing between the pillars in the first group of micropillar arrays is 4 microns, and the second The spacing between the pillars in the first and third sets of micropillar arrays is 5 micrometers.

Verification experiment: use the microfluidic devices provided in Examples 1-6 and Comparative Examples 1-6 to sort the same cell population, the specific test operation process is as follows: the bacteria to be separated are cultured and transferred twice in shake flasks After growing to the logarithmic phase, when OD600=0.2, use the first syringe to draw the bacterial sample and enter and exit from the cell sample inlet, and at the same time use the second syringe to absorb the medium and then inject it from the medium injection port, and the bacteria and The medium is injected at the same time and the flow rate is the same; the bacteria flow through the microfluidic device and then enter the cell collection tank connected to the sample outlet through the sample outlet, and concentrate in the collection tank for microscope observation and tracking. Through image processing and analysis methods such as matlab, Data on differences in the motility of the cells are available.

Tests have proved that the microfluidic devices in Examples 1-6 can well separate the 2-4um bacteria into three parts with an average length of 2μm, 3μm and 4μm. The effect of adding an electric field is more obvious, and the bacteria collected separately The bacterial length discrimination can reach about 0.9 μm. When using the microfluidic device in Comparative Examples 1-6 to sort Escherichia coli by size, the bacteria could not be separated well because the inclination angle was too small or the spacing was too wide.

Finally, it should be noted that: the above embodiments are only used to illustrate the technical solutions of the present invention, rather than limiting them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that: It is still possible to modify the technical solutions described in the foregoing embodiments, or perform equivalent replacements for some or all of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the technical solutions of the various embodiments of the present invention. scope.

Claims (10)

1. a kind of micro fluidic device for cell sorting, it is characterised in that be connected including lower substrate and with the lower substrate Upper matrix, the lower substrate are provided with the micro-pillar array for being used for sorting cell；Flowed to along cell, the column in the micro-pillar array The angle of orientation and the cell flow direction be 2 ° -10 °；Direction is flowed to along perpendicular to cell, in the micro-pillar array Spacing between column is 8-13 microns.

2. the micro fluidic device according to claim 1 for cell sorting, it is characterised in that flowed to along cell, it is described The orientation of column in micro-pillar array and the angle of cell flow direction are 2 ° -9 °, more preferably 2 ° -8 °；Hang down on edge Directly direction is flowed in cell, the spacing between column in the micro-pillar array is 8-12 microns, and more preferably 10-12 is micro- Rice.

3. the micro fluidic device according to claim 1 for cell sorting, it is characterised in that in the micro-pillar array Column includes triangular stud, circular abutment or trapezoid stand column；

Preferably, the triangular stud is isosceles triangle column；

Preferably, the isosceles triangle column is right angled isosceles triangle column or obtuse angle isosceles triangle column.

4. the micro fluidic device according to claim 3 for cell sorting, it is characterised in that cell is along the cell stream To sideways collisions where base during flowing and in the isosceles triangle column.

5. according to the micro fluidic device for cell sorting described in claim any one of 1-4, it is characterised in that along perpendicular to Cell flows to direction, and the both sides of the lower substrate are provided with the magnetic control means for being used for applying magnetic field to cell.

6. according to the micro fluidic device for cell sorting described in claim any one of 1-4, it is characterised in that along perpendicular to Cell flows to direction, and the both sides of the lower substrate are provided with the electrode for being used for applying electric field to cell.

7. according to the micro fluidic device for cell sorting described in claim any one of 1-4, it is characterised in that along cell stream To one end of the lower substrate, which is provided with, is used for the first inlet for injecting cell and the second inlet for injecting culture medium；

The other end of the lower substrate is provided with sample export, and the sample export is no less than two and along perpendicular to cell flow direction side To dispersed placement.

8. a kind of preparation method of the micro fluidic device for cell sorting described in any one of claim 1-7, its feature exist In first passing through casting method and be prepared lower substrate, then the lower substrate is connected with upper matrix obtain and described is used for cell sorting Micro fluidic device.

9. preparation method according to claim 8, it is characterised in that the casting method includes pouring liquid charging stock in mould In tool, the isolated lower substrate after cured shaping；

Preferably, the liquid charging stock includes the prepolymer of curing agent and PDMS compositions；

Preferably, the curing agent and the PDMS weight ratio are 1：(8-12)；

Preferably, degassing process resolidification shaping is first carried out after pouring；

Preferably, the temperature during curing molding is 80-100 DEG C, time 1.5-2.5h.

10. preparation method according to claim 9, it is characterised in that the preparation method of the mould comprises the following steps： Photoresist is coated on die matrix surface, it is corresponding with the micro-pillar array in the lower substrate by being obtained after exposing, developing Cavity structure；

Preferably, the thickness of the photoresist of coating is 8-14 microns.