A DLD microfluidic chip and method based on time-of-flight sorting

Abstract

The present invention discloses a DLD microfluidic chip and method based on time-of-passage sorting, including a microfluidic chip channel layer, which comprises a sample inlet channel, a sample outlet channel, a DLD channel based on time-of-passage sorting, a flushing inlet channel, and a flushing outlet channel. Combining the traditional DLD sorting principle and the phenomenon that small particles are easily blocked behind the array columns, the array columns are designed so that the time for small particles to pass through the DLD sorting channel is much longer than that of large particles, and even small particles are completely trapped in the DLD sorting channel, thereby realizing high-efficiency separation of samples based on time of passage. Compared with traditional DLD, the present invention does not require particle focusing and does not require buffer assistance, and has the advantages of high chip space utilization rate, high separation efficiency, low chip manufacturing difficulty, and long service life.

Description

Technical Field

[0001] The present invention relates to the field of passive sorting of biological particles by deterministic lateral displacement (DLD) in a microfluidic chip, and particularly to a microfluidic chip and method for sorting cells based on transit time by utilizing the phenomenon that small particles are blocked in the flow channel or the displacement speed is lower than that of large particles due to factors such as fluid vortices behind the array columns in a traditional DLD chip. Background Art

[0002] Cell separation and purification are the basis for many biomedical researches. Microfluidic technology is a technology capable of manipulating cell-scale particles on a chip and has developed rapidly in the field of cell separation in recent years. Its advantages include small sample and reagent volumes, good portability, high sensitivity, low cost, and great development potential and wide application prospects in the fields of detection and sorting of rare cells.

[0003] Currently, in the field of passive sorting of DLD microfluidic biological particles, in order to obtain higher separation accuracy, throughput (i.e., separation efficiency) and lower blockage rate, topological optimization, shape optimization, etc. of the array columns for separation have become research hotspots in recent years and some remarkable results have been achieved. However, no matter what type of DLD chip, it sorts cells based on the spatial displacement gap of particles. When the sample enters the DLD separation flow channel, the particles need to be pre-focused or a buffer solution needs to be used as a sheath flow in advance. This makes the proportion of the array columns that actually play a separation role in all the array columns relatively low, resulting in a waste of working efficiency. Affected by the introduction of the sheath flow, the throughput of the sample flow is limited. And the present invention sorts based on transit time without the need for pre-focusing of particles or buffer assistance. The sample flow is directly injected into the chip, making full use of all the column arrays in the chip, improving the sorting efficiency, and having the advantages of reducing the manufacturing difficulty and increasing the service life. Summary of the Invention

[0004] Object of the Invention: In order to overcome the waste of working efficiency and the limitation of sample throughput in the prior art, the present invention provides a DLD chip and method based on transit time sorting. By changing the shape of the traditional DLD array columns and adjusting the gap between the column arrays, small particles are trapped behind the array columns or the transit time is much longer than that of large particles, so as to realize the sorting of different cells based on their size characteristics in this flow channel according to transit time, with high space utilization rate and sorting efficiency, and being easier to manufacture and having a long service life.

[0005] Technical Solution: To achieve the above object, the technical solution adopted by the present invention is as follows:

[0006] A DLD microfluidic chip based on time-of-flight sorting, comprising a microfluidic chip channel layer, wherein the microfluidic chip channel layer includes a sample inlet channel, a flushing inlet channel, a sample outlet channel, a flushing outlet channel, and a DLD channel based on time-of-flight sorting, and:

[0007] The sample inlet channel is composed of a number of sample inlet channel guiding bars, the right end shape of the sample inlet channel guiding bar is the same as the right end shape of the DLD channel array column, its distance from the adjacent DLD channel array column is the same as the array distance in the DLD channel based on time-of-flight sorting, and the number of the sample inlet channel guiding bars is the same as the number of array rows in the DLD channel based on time-of-flight sorting;

[0008] The flushing inlet channel is composed of a number of flushing inlet channel guiding bars, the flushing inlet channel guiding bars are evenly distributed in the flushing inlet channel, and the lower end thereof maintains a certain distance from the upper end of the DLD channel based on time-of-flight sorting;

[0009] The sample outlet channel is composed of a number of sample outlet channel guiding bars, the left end shape of the sample inlet channel guiding bar is the same as the left end shape of the DLD channel array column, its distance from the adjacent DLD channel array column is the same as the array distance in the DLD channel based on time-of-flight sorting, and the number of the sample outlet channel guiding bars is the same as the number of array rows in the DLD channel based on time-of-flight sorting;

[0010] The DLD channel based on time-of-flight sorting is composed of a large number of DLD channel array columns, and its array pattern is the same as that of the traditional DLD array: a certain distance D is maintained horizontally between every two columns of array columns x , and there is a difference of ε vertically; a certain distance D is maintained between every two rows of array columns y . Wherein the array angle range is 5° to 60°; D x ranges from 10 μm to 300 μm; D y ranges from 20 μm to 600 μm; ε ranges from 2 μm to 300 μm.

[0011] Preferably: the sample inlet channel guiding bars and the sample outlet channel guiding bars have the same structure and arrangement, their widths are the same as the height of the DLD channel array column, the distance between two adjacent guiding bars is the same as the array row spacing in the DLD channel based on time-of-flight sorting, and the two kinds of guiding bars are arranged horizontally.

[0012] Preferably: the right end of the sample inlet channel guiding bar and the left end of the sample outlet channel guiding bar together with the DLD channel array column will form a sorting array.

[0013] Preferably, the flushing inlet flow channel guide strips are arranged vertically and have a width much greater than the width of the DLD flow channel array column.

[0014] Preferably, the shape of the DLD channel array column is a rectangle with semi-elliptical pits on both sides, with the sharp corners rounded and the long side perpendicular to the direction of the fluid flowing into the channel.

[0015] Preferably: the DLD flow channel array channels based on time sorting are divided into row channels and column channels, the row channels are wider and are composed of the upper and lower ends of the DLD flow channel array columns, forming a certain angle (i.e., array angle) with the horizontal line, and the column channels are narrower and are composed of the left and right end pits of the DLD flow channel array columns, and the column channels are vertical.

[0016] A DLD microfluidic method based on time sorting comprises the following steps:

[0017] Step 1: injecting a sample flow containing sample particles from an upper inlet at a sample flow inlet channel;

[0018] Step 2: After the sample flow passes through the flow channel formed by the flow channel guide strips at the sample flow inlet, it has a uniform horizontal flow velocity and is aligned with the channel inlet.

[0019] In step 3, after the sample flow enters the row channel entrance, the small particles in the sample flow follow the streamlines and flow through several rows of DLD channel array columns before flowing into the row channel composed of two elliptical pits behind a certain array column. At this time, the flow rate of small particles in this area drops significantly or even approaches 0, while the large particles in the sample flow collide with the DLD channel array columns and quickly flow into the sample flow outlet channel along the row channel, thereby leaving the separation device.

[0020] In step 4, after a certain amount of sample flow has passed through, the large particles in the sample flow have essentially all flowed out of the time-sorted DLD channel, while the small particles remain trapped within the DLD channel. At this point, the sample inlet and outlet channels are closed, the flushing inlet and outlet channels are opened, and the flushing flow is rapidly introduced through the flushing inlet channel.

[0021] In step 5, the flushing flow passes through the DLD flow channel for sorting based on the passing time and flushes all the small particles blocked inside through the column channel to the flushing flow outlet flow channel, thereby leaving the separation device for the next sorting.

[0022] Preferably, the flow rate of the flushing flow in step 5 should be much greater than the flow rate of the sample flow.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] (1) Since the array columns in the DLD chip based on time-of-flight sorting do not need to consider the problem of small particle blockage, larger array columns are required when separating particles or cells of the same size. Therefore, it is easier to manufacture, less likely to be damaged and worn during use, and can extend its service life.

[0025] (2) In existing DLDs, in order to prevent excessive sample dispersion, the particles need to be pre-focused or a buffer solution needs to be used as a sheath flow when the sample enters the DLD separation channel. This makes the chip manufacturing more complex, and the throughput of the sample flow is limited by the introduction of the sheath flow. In contrast, the present invention based on time-of-flight sorting does not require pre-focusing of particles or buffer assistance. The sample flow is directly injected into the chip, fully utilizing all the column arrays in the chip and improving the sorting efficiency.

[0026] (3) Compared with traditional DLDs, at the same sorting efficiency, the sample flow velocity in this device is lower, which is more cell-friendly.

[0027] (4) When the concentration of small particles in the required sample is relatively high or the chip is relatively small, since the present invention device has the same structure at the inlet and outlet, after the sample flow sorts and removes a part of the small particles from left to right, the sample flow can be sorted from right to left to wash away another part of the small particles. By repeating this process, large particles can be continuously purified on the same chip. This is impossible for traditional DLDs based on spatial sorting. Description of the Drawings

[0028] Figure 1 Schematic diagram of the microfluidic chip of the present invention;

[0029] Figure 2 Schematic diagram of the channel layer structure of the microfluidic chip of the present invention;

[0030] Figure 3 Schematic diagram of the DLD array structure of the present invention;

[0031] Figure 4 Schematic diagram of the separation principle of large and small particles of the present invention;

[0032] In the figure: 1. Sample flow inlet channel, 11. Sample flow inlet channel guide bar, 2. Flushing flow inlet channel, 21. Flushing flow inlet channel guide bar, 3. Sample flow outlet channel, 31. Sample flow outlet channel guide bar, 4. Flushing flow outlet channel, 5. DLD channel based on time-of-flight sorting, 51. DLD channel array column, 52. Row channel, 53. Column channel, 6. Sample flow, 61. Small particle, 62. Large particle, 7. Flushing flow. Detailed Description of the Invention

[0033] The present invention will be further illustrated below in conjunction with the accompanying drawings and specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. After reading the present invention, those skilled in the art's various equivalent forms of modification of the present invention fall within the scope defined by the appended claims of this application.

[0034] A DLD microfluidic chip based on time-of-passage sorting, as Figure 1 , shown in FIG. 2, includes a microfluidic chip channel layer, and the microfluidic chip channel layer includes a sample inlet channel 1, a flushing inlet channel 2, a sample outlet channel 3, a flushing outlet channel 4, and a DLD channel 5 based on time-of-passage sorting, wherein:

[0035] As Figure 2 shown, the sample inlet channel 1 is composed of a plurality of sample inlet channel guiding strips 11. The right end shape of the sample inlet channel guiding strip 11 is the same as the right end shape of the DLD channel array column 51. Its distance from the adjacent DLD channel array column 51 is the same as the array distance in the DLD channel 5 based on time-of-passage sorting, and the number of the sample inlet channel guiding strips 11 is the same as the number of array rows in the DLD channel 5 based on time-of-passage sorting;

[0036] The flushing inlet channel 2 is composed of a plurality of flushing inlet channel guiding strips 21. The flushing inlet channel guiding strips 21 are evenly distributed in the flushing inlet channel 2, and the lower end thereof keeps a certain distance from the upper end of the DLD channel 5 based on time-of-passage sorting. The flushing channel is mainly used to quickly flush the residual small particles 61 in the chip, so there is no need for too small guiding strips to avoid increasing the manufacturing difficulty;

[0037] The sample outlet channel 3 is composed of a plurality of sample outlet channel guiding strips 31. The left end shape of the sample outlet channel guiding strip 31 is the same as the left end shape of the DLD channel array column 51. Its distance from the adjacent DLD channel array column 51 is the same as the array distance in the DLD channel 5 based on time-of-passage sorting, and the number of the sample outlet channel guiding strips 31 is the same as the number of array rows in the DLD channel 5 based on time-of-passage sorting;

[0038] As Figure 3 shown, the DLD channel 5 based on time-of-passage sorting is composed of a large number of DLD channel array columns 51. The shape of the DLD channel array column 51 is a vertically placed rectangle with an oval pit opened on both sides and the sharp corners rounded. The column channels 53 composed of such array columns have good obstruction to small particles and can accommodate more small particles. Its array method is the same as the traditional DLD array method: a certain distance D is maintained horizontally between every two columns of array columns x, with a vertical difference of ε; there is a certain distance D between every two rows of array columns y . Among them, D x ranges from 10 μm to 300 μm; D y ranges from 20 μm to 600 μm; the range of ε is from 2 μm to 300 μm, specifically related to the cell size, sorting time, and the size of the DLD flow channel array columns 51.

[0039] The DLD flow channel 5 array channels based on time-of-flight sorting are divided into row channels 52 and column channels 53. The row channels 52 are wider and composed of the upper and lower ends of the DLD flow channel array columns, forming a certain angle (i.e., the array angle) with the horizontal line, and the range of the array angle is from 5° to 60°; the column channels 53 are narrower and composed of the concave pits at the left and right ends of the DLD flow channel array columns, and the column channels 53 are vertical.

[0040] A DLD microfluidic method based on time-of-flight sorting includes the following steps:

[0041] Step 1, injecting the sample stream 6 with sample particles from the upper inlet at the sample stream inlet channel 1;

[0042] Step 2, after the sample stream 6 passes through the flow channel composed of the sample stream inlet channel guiding strips 11, it has a uniform horizontal flow rate and is aligned with the inlet of the row channel 52.

[0043] Step 3, after the sample stream 6 enters the inlet of the row channel 52, the small particles 61 in the sample stream 6 flow along the streamline and flow into the column channel 53 composed of two elliptical concave pits behind a certain array column after flowing through several columns of DLD flow channel array columns 51. At this time, the flow rate of the small particles 61 in this area drops significantly or even approaches 0, while the large particles 62 in the sample stream 6 collide with the DLD flow channel array columns 51 and quickly flow into the sample stream outlet channel 3 along the row channel 52 to leave the separation device.

[0044] As Figure 4 shown, the small particles 61 collide with the lower array column 51 after flowing a certain distance in the row channel 52. Under the combined action of the fluid and the wall surface, they can always flow into the column channel 53. Due to the special shape design of the array column, the flow rate of the small particles 61 decreases or even gets completely trapped in the column channel 53. While the large particles, due to their size, are subjected to a stronger fluid driving force, collide with the array column 51 but do not flow into the column channel due to size factors.

[0045] Step 4, after a certain amount of sample stream 6 has flowed through, substantially all of the large particles 62 in the sample stream 6 have flowed out of the DLD channel 5 sorted based on transit time, while the small particles 61 are still all trapped within the DLD channel 5. At this time, the sample stream inlet channel 1 and the sample stream outlet channel 3 are closed, the flushing stream inlet channel 2 and the flushing stream outlet channel 4 are opened, and the flushing stream 7 is quickly introduced from the flushing stream inlet channel 2.

[0046] Step 5, the flushing stream 7 passes through the DLD channel 5 sorted based on transit time and flushes all the small particles 61 blocked therein through the column channels 53 to the flushing stream outlet channel 4, thereby leaving the separation device for the next sorting.

[0047] The above are only the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and refinements can be made, and these improvements and refinements should also be regarded as the protection scope of the present invention.

Claims

1. A DLD microfluidic chip based on time-of-flight sorting, comprising a microfluidic chip channel layer, characterized in that: The microfluidic chip channel layer includes a sample inlet channel (1), a flushing inlet channel (2), a sample outlet channel (3), a flushing outlet channel (4), and a DLD channel (5) based on time-of-flight sorting, wherein: The sample inlet channel (1) is composed of a plurality of sample inlet channel guiding strips (11). The shape of the right end of the sample inlet channel guiding strip (11) is the same as the shape of the right end of the DLD channel array column (51). The distance between it and the adjacent DLD channel array column (51) is the same as the array distance in the DLD channel (5) based on time-of-flight sorting. And the number of the sample inlet channel guiding strips (11) is the same as the number of array rows in the DLD channel (5) based on time-of-flight sorting; The flushing inlet channel (2) is composed of a plurality of flushing inlet channel guiding strips (21). The flushing inlet channel guiding strips (21) are evenly distributed in the flushing inlet channel (2), and the lower end thereof maintains a certain distance from the upper end of the DLD channel (5) based on time-of-flight sorting; The sample outlet channel (3) is composed of a plurality of sample outlet channel guiding strips (31). The shape of the left end of the sample outlet channel guiding strip (31) is the same as the shape of the left end of the DLD channel array column (51). The distance between it and the adjacent DLD channel array column (51) is the same as the array distance in the DLD channel (5) based on time-of-flight sorting. And the number of the sample outlet channel guiding strips (31) is the same as the number of array rows in the DLD channel (5) based on time-of-flight sorting; The DLD flow channel (5) based on time sorting is composed of a large number of DLD flow channel array columns (51), and its array method is the same as the traditional DLD array method: a certain distance D is maintained between each two rows of array columns in the horizontal direction. x , the vertical direction differs by ε; a certain distance D is maintained between each two rows of array columns y ; The array channels of the DLD channel (5) based on time-of-flight sorting are divided into row channels (52) and column channels (53). The row channels are composed of the relative upper and lower ends of adjacent two rows of DLD channel array columns (51), and form a certain angle with the horizontal line, that is, the array angle. The column channels are narrower than the row channels and are composed of the relative left and right end pits of adjacent two columns of DLD channel array columns (51), and the column channels are vertical; Small particles in the sample flow (6) flow into the column channel (53) under the combined action of the fluid and the wall surface, while large particles do not flow into the column channel (53) due to size factors.

2. The DLD microfluidic chip based on time sorting according to claim 1, characterized in that: The sample inlet channel guiding strips (11) and the sample outlet channel guiding strips (31) have the same structure and arrangement. Their widths are the same as the height of the DLD channel array column (51), the distance between adjacent two guiding strips is the same as the array row spacing in the DLD channel (5) based on time-of-flight sorting, and the two kinds of guiding strips are both horizontally arranged.

3. The DLD microfluidic chip based on time-of-flight sorting according to claim 2, characterized in that: The right end of the sample inlet channel guiding strip (11) and the left end of the sample outlet channel guiding strip (31) together with the DLD channel array column (51) will form a sorting array.

4. The DLD microfluidic chip based on time-of-flight sorting according to claim 3, characterized in that: The flushing inlet channel guiding strips (21) are vertically arranged and the width is much larger than the width of the DLD channel array column (51).

5. The DLD microfluidic chip based on time-of-flight sorting according to claim 4, wherein: The DLD flow channel array column (51) is rectangular with semi-elliptical pits on both sides, and the sharp corners are rounded. The long side is perpendicular to the fluid inflow channel direction.

6. The DLD microfluidic chip based on time-of-flight sorting according to claim 5, characterized in that: The horizontal distance between every two columns of the DLD flow channel array columns (51) is maintained at D. x , and the vertical difference is ε; the distance between every two rows of array columns is maintained at D. y , where the array angle ranges from 5° to 60°; D x ranges from 10 μm to 300 μm; D y ranges from 20 μm to 600 μm; ε ranges from 2 μm to 300 μm.

7. A microfluidic method based on the DLD microfluidic chip sorted by time as claimed in claim 6, characterized in that, It includes the following steps: Step 1: Inject the sample flow (6) with sample particles from the upper inlet above the sample flow inlet channel (1). Step 2: After passing through the flow channel formed by the sample flow inlet channel guide bars (11), the sample flow (6) has a uniform horizontal flow rate and is aligned with the inlet of the row channel (52). Step 3: After the sample flow (6) enters the inlet of the row channel (52), the small particles (61) in the sample flow (6) flow along the streamline and flow into the column channel (53) composed of two elliptical pits behind a certain array column after flowing through several columns of the DLD flow channel array column (51). At this time, the flow rate of the small particles (61) in this area drops significantly or even approaches 0, while the large particles (62) in the sample flow (6) collide with the DLD flow channel array column (51) and quickly flow out of the sample flow outlet channel (3) along the row channel (52). Step 4: After a certain amount of the sample flow (6) has flowed through, almost all of the large particles (62) in the sample flow (6) have flowed out of the DLD flow channel (5) based on time-of-flight sorting, while all the small particles (61) are still trapped in the DLD flow channel (5). At this time, close the sample flow inlet channel (1) and the sample flow outlet channel (3), open the flushing flow inlet channel (2) and the flushing flow outlet channel (4), and quickly introduce the flushing flow (7) from the flushing flow inlet channel (2). Step 5: The flushing flow (7) passes through the DLD flow channel (5) based on time-of-flight sorting and flushes all the blocked small particles (61) out of the flushing flow outlet channel (4) through the column channel (53) for the next sorting.

8. The microfluidic method based on the DLD microfluidic chip sorted by passing time according to claim 7, characterized in that: In Step 5, the flow rate of the flushing flow (7) should be much greater than the flow rate of the sample flow (6).

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