Device for sorting and enriching and application thereof

Abstract

The invention provides a device for sorting and enriching and application of the device, and the device comprises a shell; screening holes are formed in the sorting chips, the N sorting chips which are sequentially arranged divide the inner space of the shell into N + 1 cavities, and N is larger than or equal to 1; wherein a first cavity is formed between one side of the shell and the sorting chip adjacent to the shell; a second cavity is formed between the other side of the shell and the sorting chip adjacent to the shell; a first outlet and a first inlet which are used for connecting the first cavity with the outside are formed in the cavity wall of the first cavity; and a second outlet for connecting the second cavity with the outside is formed in the cavity wall of the second cavity.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of biomedical technology, in particular to a device for sorting and enriching and application thereof. BACKGROUND

[0002] At present, the methods for sorting and / or enriching cells are generally divided into two categories: one is sorting and / or enriching cells according to their physical properties (cell size, density, motility, electrical properties, etc.), and the other is sorting and / or enriching cells according to their biochemical properties (surface antigens, etc.). The method for sorting and / or enriching cells according to their physical properties, for example, according to the size of the cells, a screen with a certain pore size is designed to filter and separate, and for example, according to the different densities of cells, the method of density gradient centrifugation is used for sorting and / or enriching. The method for sorting and / or enriching cells according to their biochemical properties, for example, according to the different surface antigens of cells, the antibody labeled with fluorescence is combined with the cells, and the flow cytometer is used for sorting and / or enriching.

[0003] The cell clusters formed by the aggregation of cells may be formed by the aggregation of the same type of cells, or may be formed by the aggregation of different types of cells. Different types of cells in the cell cluster may play different roles, so that the cell cluster has biological functions that individual cells do not have, and realizes biological processes that individual cells cannot complete. The size of the cell cluster is related to the number of cells and the type of cells.

[0004] Malignant tumors are a serious disease, and most tumor patients die of tumor metastasis. In the past, it was believed that tumors metastasize through single tumor cells circulating in the blood or lymph to form metastatic tumors, so various techniques for separating and detecting single circulating tumor cells from blood have been developed. Typical separation and detection techniques for single circulating tumor cells, such as Johnson's CellSearch product. Recent studies have shown that tumors mainly metastasize through tumor cell clusters. The tumor cell cluster may contain tumor cells of different properties, and may also contain non-tumor cells. The size of the tumor cell cluster also varies, and the content in the blood circulation system is very low. The number of tumor cell clusters in the blood is related to the prognosis of the patient. Because the tumor cell clusters in the blood circulation system are not easy to obtain, there are also technical solutions to obtain dispersed single cells by digesting tumor tissue, and then aggregate to form tumor cell clusters for scientific research and clinical detection. However, the tumor cell clusters formed by this technical solution have large size differences, and the tumor cell clusters containing a certain number of cells and uniform size need to be sorted and / or enriched to obtain stable and accurate detection results.

[0005] Existing cell sorting and / or enrichment methods are mainly applied to the sorting and / or enrichment of single cells, while methods and techniques for sorting and / or enriching cell clusters are severely lacking. Due to the differences in cell number and cell species, cell cluster sizes vary widely, making single-cell sorting and / or enrichment techniques largely unsuitable for cell cluster sorting and / or enrichment. Existing cell sieving and / or enrichment methods primarily use mesh polymer membranes for filtration, mostly used for filtering out small-volume solid tissues after mincing and digestion. The pore size of the mesh polymer membranes used in cell sieving is mostly greater than 40 μm, making them unsuitable for sorting and / or enriching free cell clusters present in fluid samples. Mesh polymer membranes are prone to clogging, making it difficult to process large-volume samples. Cell clusters trapped within the mesh polymer membrane are also difficult to recover, resulting in significant losses and making them unsuitable for separating sparse circulating tumor cell clusters. Summary of the Invention

[0006] To address the problems in the prior art, this application provides an apparatus for sorting and enriching, and its application. The technical solution of this application is as follows:

[0007] 1. An apparatus for sorting and enriching, comprising:

[0008] case;

[0009] N sorting chips, each with a screening hole, are arranged sequentially to divide the internal space of the housing into N+1 cavities, where N≥1;

[0010] in,

[0011] A first cavity is formed between one side of the housing and the adjacent sorting chip;

[0012] A second cavity is formed between the other side of the housing and the adjacent sorting chip;

[0013] The cavity wall of the first cavity has a first outlet and a first inlet that connect the first cavity to the outside.

[0014] A second outlet is formed on the cavity wall of the second cavity, connecting the second cavity to the outside.

[0015] 2. The device as described in item 1, wherein N≥2; a third cavity is formed between any adjacent sorting chips, and each third cavity has a third outlet on its cavity wall that connects the third cavity to the outside.

[0016] 3. The apparatus of claim 2, wherein at least one of the third cavities has a third inlet formed on its cavity wall connecting the third cavity to the outside.

[0017] 4. The apparatus of item 1, wherein the first cavity has two or more first inlets formed on the cavity wall.

[0018] 5. The apparatus of item 2, wherein the pore size of the screening holes of each of the sorting chips gradually increases from one side of the housing to the other side of the housing; and / or the pore size of the screening holes is equal to or greater than 8 μm.

[0019] 6. The apparatus of item 1, wherein the cross-sectional shape of the screening holes is circular, polygonal; and / or at least two of the screening holes in at least one of the sorting chips have different shapes.

[0020] 7. The apparatus of item 2, wherein the apparatus further comprises micro pillars, at least one micro pillar is disposed in at least one of the first cavity, the second cavity, and the third cavity.

[0021] 8. The apparatus of item 7, wherein the micro pillars are disposed on the housing; and / or the micro pillars are disposed on the surface of the sorting chips.

[0022] 9. The apparatus of item 7, wherein the cross-section of the micro pillars is circular, elliptical, or polygonal.

[0023] 10. The apparatus of item 1, wherein the sorting chips are disposed in the region of the screening holes to form a screening area; at least one surface of the screening area of at least one of the sorting chips is partially or entirely non-planar.

[0024] 11. The apparatus of item 10, wherein at least one surface of the screening area of at least one of the sorting chips facing the first cavity is partially or entirely non-planar.

[0025] 12. The apparatus of item 10, wherein the non-planar structure is a curved surface.

[0026] 13. An apparatus for sorting and enriching, comprising:

[0027] a housing;

[0028] N sorting chips, each of the sorting chips having a screening hole disposed thereon, N being an integer greater than or equal to 2, the N sorting chips being disposed in sequence to divide the interior space of the housing into N+1 cavities;

[0029] wherein,

[0030] a first cavity is formed between one side of the housing and the sorting chip adjacent thereto;

[0031] a second cavity is formed between the other side of the housing and the sorting chip adjacent thereto.

[0032] A third cavity is formed between any two adjacent sorting chips.

[0033] A first outlet is formed on the cavity wall of the first cavity to connect the first cavity with the outside world.

[0034] A second outlet is formed on the cavity wall of the second cavity to connect the second cavity with the outside world.

[0035] A third outlet is formed on the cavity wall of the third cavity to connect the third cavity with the outside world, and more than one third inlet is formed on the cavity wall of at least one third cavity to connect the third cavity with the outside world.

[0036] 14. The device of item 13, wherein N = 2; and / or, more than one third inlet is formed on the cavity wall of at least one third cavity to connect the third cavity with the outside world.

[0037] 15. The device of item 14, wherein N = 2; and, the aperture diameters of the screening holes 34b between the two sorting chips are the same.

[0038] 16. The device of item 13, wherein the aperture diameter of the screening holes is equal to or greater than 8 μm.

[0039] 17. The device of item 13, wherein the cross-sectional shape of the screening holes is circular, polygonal; and / or, in at least one sorting chip, at least two screening holes have different shapes.

[0040] 18. The device of item 13, wherein the device further comprises micro-pillars, and at least one micro-pillar is disposed in at least one of the first cavity, the second cavity, and the third cavity.

[0041] 19. The device of item 18, wherein the micro-pillars are disposed on the housing; and / or, the micro-pillars are disposed on the surface of the sorting chip.

[0042] 20. The device of item 19, wherein the cross-section of the micro-pillars is circular, elliptical, and / or polygonal.

[0043] 21. The device of item 13, wherein the sorting chip is provided with a screening area formed by the screening holes; and at least one surface of the screening area of at least one sorting chip is partially or entirely non-planar.

[0044] 22. The device of item 21, wherein at least one surface of the screening area of at least one sorting chip facing the third inlet is partially or entirely non-planar.

[0045] 23. The apparatus of item 21, wherein the non-planar structure is a curved surface.

[0046] 24. Use of the apparatus of any one of items 1 to 23 for sorting and enriching a fluid sample of cell clusters in a peripheral blood sample; cell clusters in a pleural effusion, ascites effusion, lymphatic fluid, urine or cerebrospinal fluid; cell clusters formed after enzymatic digestion of a solid tissue; cell clusters formed after digestion of a solid tissue into single cells and re-clustering of the single cells; or, liposomes, water-in-oil microdroplets or oil-in-water microdroplets.

[0047] The above described apparatus for sorting and enriching provided in the present application, by the "direction of movement 2", traps the cells and / or cell clusters larger than the screening hole 34a in the corresponding cavity, and collects the cells and / or cell clusters of the corresponding size range (larger size) through the outlet corresponding to the cavity, thereby screening and enriching the cells and / or cell clusters of the size range; by the "direction of movement 1", the cells and / or cell clusters smaller than the screening hole pass through the screening hole into the lower cavity, thereby further screening and enriching the cells and / or cell clusters of the corresponding size range (smaller size); when the inlet for providing the fluid such as buffer is provided on the cavity, the fluid can drive the flow of the fluid sample in the corresponding cavity on one hand, reducing / preventing the cells and / or cell clusters from blocking the screening hole, and on the other hand, directly collecting the cells and / or cell clusters suspended in the corresponding fluid, thereby directly obtaining the suspension of the cells and / or cell clusters which can be used for subsequent use; when the screening area of the micro-column and / or the sorting chip is set to be non-planar, the effect of changing the original flow direction of the fluid in the cavity can be achieved, the opportunity of the contact between the cell clusters and the screening hole of the sorting chip is increased, and the cell clusters smaller than the screening hole flow to the next cavity through the screening hole as much as possible, rather than directly flowing to the outlet corresponding to the cavity, thereby enhancing the screening and enrichment effect of the apparatus of the present application.

[0048] The above description is only a summary of the technical solutions of the present application, in order to make the technical means of the present application more clear and understandable, to the extent that the person skilled in the art can implement according to the content of the description, and in order to make the above and other purposes, features and advantages of the present application more obvious and easy to understand, the specific embodiments of the present application are illustrated as follows. BRIEF DESCRIPTION OF DRAWINGS

[0049] Figure 1 : Structure schematic diagram of the apparatus for sorting and enriching in an embodiment of the present application;

[0050] Figure 2 : Structure schematic diagram of the inner side of the upper shell in an embodiment of the present application;

[0051] Figure 3: In an embodiment of the present application, the structure diagram of the upper shell outer side;

[0052] Figure 4 : In an embodiment of the present application, the structure diagram of the first sorting chip upper side;

[0053] Figure 5 : In an embodiment of the present application, the structure diagram of the first sorting chip lower side;

[0054] Figure 6 : In an embodiment of the present application, the structure diagram of the lower shell inner side;

[0055] Figure 7 : In an embodiment of the present application, the structure diagram of the lower shell outer side;

[0056] Figure 8 : In an embodiment of the present application, the structure diagram of the sorting chip upper side not adjacent to the upper shell;

[0057] Figure 9 : In an embodiment of the present application, the structure diagram of the sorting chip lower side not adjacent to the upper shell;

[0058] Figure 10 : In an embodiment of the present application, the structure diagram of the sorting chip with hexagonal screening holes;

[0059] Figure 11 : In an embodiment of the present application, the structure diagram of the sorting chip with rectangular and circular combined screening holes;

[0060] Figure 12 : In an embodiment of the present application, the structure diagram of the upper shell with micro columns;

[0061] Figure 13 : In an embodiment of the present application, the structure diagram of the lower shell with micro columns;

[0062] Figure 14 : In an embodiment of the present application, the structure diagram of the sorting chip with micro columns;

[0063] Figure 15 : In an embodiment of the present application, the structure diagram of the sorting chip with wavy structure;

[0064] Figure 16 : In another embodiment of the present application, the structure diagram of the device for sorting and enriching.

[0065] Explanation of reference signs:

[0066] 10a, upper shell; 11a, upper shell side wall;

[0067] 20a, lower housing; 21a, lower housing sidewall;

[0068] 30a, sorting chip; 31a, first sorting chip; 32a, second sorting chip; 33a, third sorting chip; 34a, screening hole; 35a, chip sidewall;

[0069] 41a, first cavity; 42a, second cavity; 43a, third cavity; 44a, first outlet; 45a, second outlet; 46a, third outlet; 47a, first inlet; 48a, third inlet;

[0070] 50a, opening; 60a, microcolumn; 70a, cell cluster.

[0071] 10b, upper housing;

[0072] 20b, lower housing;

[0073] 31b, first sorting chip; 32b, second sorting chip; 34b, screening hole;

[0074] 41b, first cavity; 42b, second cavity; 43b, third cavity; 44b, first outlet; 45b, second outlet; 46b, third outlet; 48b, third inlet;

[0075] 70b, cell cluster. DETAILED DESCRIPTION

[0076] The following embodiments of the present application are only used to illustrate the specific embodiments of the present application, and these embodiments cannot be understood as the limitation of the present application. Any changes, modifications, substitutions, combinations, simplifications made without departing from the spirit and principles of the present application are considered as equivalent replacement methods, which fall within the protection scope of the present application.

[0077] The present embodiment provides a device for sorting and enrichment, as shown in Figures 1-11 The device comprises:

[0078] a housing;

[0079] N sorting chips, the sorting chips are provided with screening holes 34a, and the N sorting chips arranged in sequence divide the internal space of the housing into N+1 cavities, N≥1, as shown in Figure 1 In the present embodiment, the N sorting chips are three sorting chips arranged from top to bottom, i.e. the first sorting chip 31a, the second sorting chip 32a, and the third sorting chip 33a; wherein,

[0080] A first cavity 41a is formed between one side of the shell (such as the inner side of the upper shell 10a) and the sorting chip (such as the first sorting chip 31a) adjacent thereto;

[0081] A second cavity 42a is formed between the other side of the shell (such as the inner side of the lower shell 20a) and the sorting chip (such as the third sorting chip 33a) adjacent thereto;

[0082] A first outlet 44a and a first inlet 47a are formed on the cavity wall of the first cavity 41a, connecting the first cavity 41a with the outside world;

[0083] A second outlet 45a is formed on the cavity wall of the second cavity 42a, connecting the second cavity 42a with the outside world.

[0084] The shell of the present application is not particularly limited in terms of its composition structure, as long as it can form a closed internal space, such as a separate closed shell, in which the sorting chips are arranged in sequence, or as in the present embodiment, the shell is composed of the upper shell 10a, the lower shell 20a, and the chip side walls 35a of the N sorting chips 30a arranged therebetween.

[0085] The present application is not particularly limited in terms of the specific implementation of the N sorting chips arranged in sequence to divide the internal space of the shell into N+1 cavities. For example, if N sorting chips are arranged in sequence in a closed shell, the N sorting chips will divide the internal space of the shell into N+1 cavities, or as in the present embodiment, N+1 cavities are formed by the stacking of the upper shell 10a, the sorting chips 30a, and / or the lower shell 20a (see Figures 1-9 ).

[0086] The present application is not particularly limited in terms of the specific implementation of each outlet (such as the first outlet 44a, the second outlet 45a, and the third outlet 46a below) and / or inlet (the first inlet 47a, and the second inlet and the third inlet 48a below). For example, a channel is directly formed on the shell to communicate with each cavity and the outside world to obtain the corresponding inlet and / or outlet, or as in the present embodiment, openings 50a are provided on the side walls of the upper shell 10a, the sorting chips 30a, and / or the lower shell 20a, and the stacking of the upper shell 10a, the lower shell 20a, and the sorting chips 30a to obtain the corresponding inlet and / or outlet (see Figures 1-9 ).

[0087] The number and density of each screening hole 34a can be specifically set by those skilled in the art as needed, and the present application will not be repeated here. For example, a plurality of screening holes 34a are arranged in an array at a certain density.

[0088] As for the shape of the screening holes, the present application is not specifically limited, as long as it can be applied to the screening of the corresponding cell clusters, such as the cross-sectional shape of the screening holes being circular, polygonal, such as Figure 10 as shown, the cross-sectional shape of the screening holes is hexagonal, that is, the array is even a plurality of screening holes with hexagonal cross-section. In one sorting chip, the screening holes can be a combination of multiple shapes, such as Figure 11 In the present application, the screening holes are a combination of rectangular and circular shapes, that is, in at least one (such as one, part or all) of the sorting chips, at least two of the screening holes have different shapes and / or hole diameters.

[0089] As for the hole diameter of each screening hole 34a, those skilled in the art can make specific settings as needed. Specifically, in the present embodiment, the hole diameter of the screening holes is as small as 8 μm, that is, the hole diameter of the screening holes 34a in the present application is 8 μm or more, specifically, 9 μm, 10 μm, 12 μm, 15 μm, 18 μm, 20 μm, 25 μm, 30 μm, 35 μm, 50 μm, 75 μm, 100 μm, 200 μm or 500 μm or more. Further, when the hole diameter of the screening holes 34a is less than or equal to 40 μm, the sorting chip is suitable for sorting and / or enriching free cell clusters present in a bulk fluid sample. It should be noted that in the present application, the hole diameter refers to the diameter of the largest circle that can be accommodated by the cross-section of the screening hole.

[0090] As for the preparation method of the housing (such as the upper housing 10a, the lower housing 20a, etc.) and / or the sorting chip, it can be obtained by the existing and mature microfluidic chip processing technology such as etching, injection molding, etc. Then, the complete device for sorting and enriching of the present application is formed by bonding and packaging.

[0091] As for the material of the housing (such as the upper housing 10a, the lower housing 20a, etc.) and / or the sorting chip in the present application, it can be a metal material, an inorganic material and / or a high polymer material, preferably an inorganic material (such as glass, silicon wafer, ceramic) and / or a high polymer material (such as a polymer material).

[0092] In use of the device provided by the present embodiment, the fluid sample (here, the fluid sample containing cell clusters is taken as an example, and the principle is the same for other fluid samples) is supplied from the first inlet 47a, and the fluid sample is powered to flow by applying positive pressure at the first inlet 47a, negative pressure at the first outlet 44a, and / or negative pressure at the second outlet 45a. When the fluid sample flows in each cavity (such as the first cavity 41a, the second cavity 42a, etc.), the flow direction is mainly divided into: movement direction 1, for cells and / or cell clusters smaller than the pore size of the screening hole 34a of the sorting chip, which pass through the screening hole 34a to enter the lower cavity; movement direction 2, for cells and / or cell clusters larger than the pore size of the screening hole 34a of the sorting chip, which cannot pass through the screening hole 34a of the sorting chip and can only flow to the outlet corresponding to the cavity where it is located. Then, as shown in Figure 1 the fluid sample is supplied from the first inlet 47a, and the cells and / or cell clusters larger than the screening hole 34a of the first sorting chip 31a flow into the first outlet 44a through the above-mentioned "movement direction 2", so as to screen and enrich the cells and / or cell clusters in the corresponding size range (larger size); and the cells and / or cell clusters smaller than the screening hole 34a of the first sorting chip 31a pass through the screening hole 34a of the first sorting chip 31a to enter the lower cavity through the above-mentioned "movement direction 1", so as to further screen and enrich the cells and / or cell clusters in the corresponding size range (smaller size).

[0093] It should be noted that in the present application, "cell clusters" refer to cell aggregates formed by two or more cells combined together through covalent and / or non-covalent interaction, which move in a fluid as a whole.

[0094] Preferably, when N≥2 (i.e. when the sorting chip is more than two, such as 3, 4, 5, 6, 7, 8, 9 or 10 or more, hereinafter, the embodiment with three sorting chips is taken as an example), a third cavity 43a is formed between the sorting chips, and the third cavity 43a is formed with a third outlet 46a on the cavity wall, connecting the third cavity 43a with the outside world. The aperture of the sorting chip sieve hole gradually increases from one side of the shell (the upper shell 10a) to the other side of the shell (the lower shell 20a). At this time, the negative pressure can also be provided through the third outlet 46a to provide power for the fluid sample flow. Then, the fluid sample flowing into the third cavity 43a (hereinafter, the cavity between the first sorting chip 31a and the second sorting chip 32a) from the first cavity 41a also has two flow directions. Through the above-mentioned "movement direction 2", the cells and / or cell clusters larger than the second sorting chip 32a flow into the third outlet 46a corresponding to the cavity, thereby screening and enriching the cells and / or cell clusters of the corresponding size range. Through the above-mentioned "movement direction 1", the cells and / or cell clusters smaller than the second sorting chip 32a enter the lower cavity through the sieve hole 34a of the second sorting chip 32a, thereby screening and enriching the cells and / or cell clusters of the corresponding size range. According to the same principle, from top to bottom, each sorting chip realizes the step-by-step screening of cells and / or cell clusters of different sizes and corresponding enrichment.

[0095] Preferably, two or more (two in the embodiment) first inlets 44a are formed on the cavity wall of the first cavity 41a; and / or, at least one (such as one, part or all) third cavity is formed with a third inlet 48a on the cavity wall, connecting the third cavity 43a with the outside world. Generally, the screened and enriched cells and / or cell clusters need to be suspended in the corresponding fluid (such as a buffer solution and other components contained in the buffer solution) to obtain a cell and / or cell cluster suspension for subsequent use. In the preferred embodiment, the additional first inlet 44a and / or third inlet 48a can be used to introduce the corresponding fluid, or even provide positive pressure through the fluid. The introduced fluid can drive the flow of the fluid sample in the corresponding cavity, reduce / prevent the cells and / or cell clusters from blocking the sieve hole, and directly collect the cells and / or cell clusters suspended in the corresponding fluid, thereby directly obtaining the cell and / or cell cluster suspension that can be used for subsequent use.

[0096] Of course, a corresponding second inlet can also be provided on the second cavity 42a to achieve the above-mentioned effect of reducing / preventing the cells and / or cell clusters from blocking the sieve hole and directly collecting the cells and / or cell clusters suspended in the corresponding fluid.

[0097] In one embodiment of the above embodiment, as Figures 12-14As shown, the device further comprises micro-pillars 60a (may also be referred to as "protrusions") disposed in at least one (e.g., one, some or all) of the first cavity 41a, the second cavity 42a, and the third cavity 43a, more preferably, in all of the above-mentioned cavities.

[0098] For example, the micro-pillars 60a can be disposed on the inner surface of the housing so that the micro-pillars 60a extend into the first cavity 41a, the second cavity 42a, and / or the third cavity 43a. For example, Figure 12 As shown, a schematic view of the micro-pillars 60a disposed on the upper housing 10a is shown, which are disposed on the inner side of the upper housing 10a, specifically, on the inner side of the upper housing side wall 11a of the upper housing 10a and on the inner side of the upper housing 10a facing the adjacent sorting chip (the first sorting chip 31a). For example, Figure 13 As shown, a schematic view of the micro-pillars 60a disposed on the lower housing 20a is shown, which are disposed on the inner side of the lower housing 20a, specifically, on the inner side of the lower housing side wall 21a of the lower housing 20a and on the inner side of the lower housing 20a facing the adjacent sorting chip (the third sorting chip 33a). For example, Figure 14 As shown, a schematic view of the micro-pillars 60a disposed on the sorting chip (specifically, the first sorting chip 31a) is shown, which are disposed on the inner side of the chip side wall 35a.

[0099] The micro-pillars 60a can also be disposed on the side of the sorting chip facing the housing (e.g., facing the upper housing 10a) and / or on the other side of the sorting chip facing the housing (e.g., facing the lower housing 20a), for example, Figure 14 As shown, a schematic view of the micro-pillars 60a disposed on the sorting chip (specifically, the first sorting chip 31a) is shown, which can be disposed on the upper side of the sorting chip, of course, can also be disposed on the lower side of the sorting chip according to the need. That is, the micro-pillars 60a can be disposed on the region of the sorting chip for disposing the screening hole 34a (i.e., the screening region described below).

[0100] As for the number and density of the micro-pillars 60a, those skilled in the art can make specific arrangements according to the need, which will not be described herein. For example, a plurality of micro-pillars 60a can be arranged in an array in at least one (e.g., one, some or all) of the first cavity 41a, the second cavity 42a, and the third cavity 43a.

[0101] As for the cross-sectional shape of the micro-pillars 60a, it can be circular, elliptical, and / or polygonal, etc. according to the need. Figures 12-14 For example, the cross-section of the micro-pillars 60a is hexagonal and trapezoidal.

[0102] By setting the micro-column 60a in the cavity, the original flow direction of the fluid in the cavity can be changed, the opportunity of the cells and / or cell clusters 70a in the fluid sample to contact the screening holes 34a of the sorting chip 30a is increased, and the cell clusters with a size smaller than the screening holes 34a can flow to the next cavity through the screening holes 34a as much as possible instead of directly flowing to the outlet corresponding to the cavity, so that the screening and enrichment effects of the device are enhanced.

[0103] In one embodiment of the above embodiment, the sorting chip 30a (for example, the first sorting chip 31a, the second sorting chip 32a, and the third sorting chip 33a) is arranged to form a screening area in the region of the screening hole 34a; and at least one surface of at least one (for example, one, part, or all) of the sorting chip is partially or entirely in a non-planar structure, preferably, at least one surface of the sorting chip facing the first cavity is partially or entirely in a non-planar structure, and more preferably, both surfaces of the sorting chip are partially or entirely in a non-planar structure. Figure 15 Exemplarily, an example in which both surfaces of the screening area are curved surfaces (specifically, wavy structures) is given.

[0104] By arranging the surface of the sorting chip facing the first cavity to be in a non-planar structure, the original flow direction of the fluid in the cavity can be changed, the opportunity of the cell clusters 70a to contact the screening holes 34a of the sorting chip 30a is increased, and the cell clusters with a size smaller than the screening holes 34a can flow to the next cavity through the screening holes 34a as much as possible instead of directly flowing to the outlet corresponding to the cavity, so that the screening and enrichment effects of the device are enhanced.

[0105] In another embodiment of the present application, another device for sorting and enrichment is provided, as shown in FIG. 6, which comprises: Figure 16 as shown in FIG. 6, which comprises:

[0106] a housing;

[0107] N sorting chips, the sorting chips being arranged with screening holes 34b, and the N sorting chips arranged in sequence divide the internal space of the housing into N+1 cavities, N≥2, as shown in FIG. 6, in this embodiment, the N sorting chips are two sorting chips arranged from top to bottom, i.e., the first sorting chip 31b and the second sorting chip 32b; wherein, Figure 16

[0108] a first cavity 41b is formed between one side (for example, the inner side of the upper housing 10b) of the housing and the sorting chip (i.e., the first sorting chip 31b) adjacent to the one side;

[0109] ​A second cavity 42b is formed between the other side (the inner side of the lower shell 20b) of the shell and the sorting chip (the second sorting chip 32b) adjacent thereto;

[0110] A third cavity 43b is formed between two adjacent sorting chips (e.g., between the first sorting chip 31b and the second sorting chip 32b, etc.);

[0111] A first outlet 44b is formed on the cavity wall of the first cavity 41b to connect the first cavity 41b with the outside;

[0112] A second outlet 45b is formed on the cavity wall of the second cavity 42b to connect the second cavity 42b with the outside;

[0113] The third cavity 43b is formed on the cavity wall of the third cavity 43b to connect the third cavity 43b with the outside, and at least one (e.g., one, part or all) of the third cavities 43b is formed on the cavity wall of the third cavity 43b to connect the third cavity 43b with the outside.

[0114] In this embodiment, the structure of the shell, the specific implementation of the N sorting chips arranged in sequence to divide the internal space of the shell into N+1 cavities, the specific implementation of each outlet and / or inlet, the number and density of the screening holes, the shape and aperture of the screening holes, the preparation method and material of the shell and / or sorting chip, the position, size, density and shape of the micro-pillars, the surface form of the screening area of the sorting chip, etc. are all referred to the previous embodiment, and the person skilled in the art knows that the same can also bring corresponding effects, which will not be described here.

[0115] In the use of the device provided in this embodiment, the fluid sample (here, the fluid sample containing cell clusters is taken as an example, and the principle is the same for other fluid samples) is supplied through the third inlet 48b, a positive pressure is applied at the third inlet 48b, a negative pressure is applied at the first outlet 44b, a negative pressure is applied at the second outlet 45b, and / or a negative pressure is applied at the third outlet 46b, which can provide power for the flow of the fluid sample. When the fluid sample flows in the third cavity 43b, the flow direction mainly includes: movement direction 1, for cells and / or cell clusters smaller than the aperture of the screening hole 34b of the sorting chip, which can pass through the screening hole 34b to enter the adjacent cavity; movement direction 2, for cells and / or cell clusters larger than the aperture of the screening hole 34b of the sorting chip, which cannot pass through the screening hole 34b of the sorting chip and can only flow to the outlet (the third outlet 46b) corresponding to the cavity where it is located. Then, as shown in FIG. 6, the fluid sample can be sorted according to the size of the cells and / or cell clusters. Figure 16As shown, fluid sample is supplied by the third inlet 48b, and cells and / or cell clusters larger than the screening holes 34b of the sorting chips (such as the first sorting chip 31b and the second sorting chip 32b) on both sides of the third chamber 43b flow into the third outlet 46b through the above-mentioned "movement direction 2", so as to screen and enrich cells and / or cell clusters in the corresponding size range (larger size); and cells and / or cell clusters smaller than the screening holes 34b of the sorting chips (such as the first sorting chip 31b and the second sorting chip 32b) on both sides of the third chamber 43b flow into the chambers (such as the first chamber 41b or the second chamber 42b) on both sides through the screening holes 34b of the first sorting chip 31b or the second sorting chip 32b through the above-mentioned "movement direction 1", so as to further screen and enrich cells and / or cell clusters in the corresponding size range (smaller size).

[0116] When the number of sorting chips on either side or both sides of the third inlet 48b for introducing fluid sample is greater than 2, the pore diameter of the screening holes 34b between the sorting chips can gradually decrease in the direction away from the third inlet 48b for introducing fluid sample, so that cells and / or cell clusters of different sizes can be screened and enriched in stages, similar to the previous embodiment.

[0117] Preferably, the cavity wall of at least one of the third chambers 43b is formed with two or more third inlets 48b connecting the third chamber with the outside. Generally, cells and / or cell clusters after screening and enrichment need to be suspended in a corresponding fluid (such as a buffer containing other costs in the buffer) to obtain a cell and / or cell cluster suspension for subsequent use. In this preferred embodiment, among the two or more third inlets 48b, one is used to introduce fluid sample, and the extra third inlets 48b are connected to the corresponding fluid, or even connected to the corresponding fluid to provide positive pressure. The fluid introduced can drive the flow of fluid sample in the corresponding chamber, reduce / prevent cells and / or cell clusters from blocking the screening holes, and directly collect cells and / or cell clusters suspended in the corresponding fluid, so as to directly obtain a cell and / or cell cluster suspension that can be used for subsequent use. Of course, in addition to providing two or more third inlets 48b on one third chamber 43b, corresponding inlets (such as a first inlet on the first chamber, a second inlet on the second chamber, and / or a third inlet on at least one (such as one, part or all) of the other third chambers) can be provided on other chambers (such as the first chamber 41b, the second chamber 42b and / or other third chambers), so as to introduce corresponding fluid through the corresponding inlets, so as to achieve the effect of reducing / preventing cells and / or cell clusters from blocking the screening holes and directly collecting cells and / or cell clusters suspended in the corresponding fluid.

[0118] Preferably, N = 2, and the pore diameters of the screening holes 34b between the two sorting chips (the first sorting chip 31b and the second sorting chip 32b) on both sides of the third cavity 43b are the same. The cells and / or cell clusters in the fluid sample introduced through the third inlet 48b that are smaller than the pore diameter of the screening holes 34b pass through the screening holes 34b of the sorting chips into the first and second cavities and flow out through the corresponding first outlet 44b and second outlet 45b and are collected; the cells and / or cell clusters that are larger than the pore diameter of the screening holes 34b are retained by the sorting chips in the third cavity and flow out through the corresponding third outlet 46b and are collected. That is, after the cells and / or cell clusters in the fluid sample are sorted by the device, two cell and / or cell cluster components with different size ranges are obtained. When the device is used for cell and / or cell cluster sorting, the cells and / or cell clusters retained by the sorting chip are easy to flow out of the device through the third outlet along with the solvent, and are not easy to block the screening holes 34b of the sorting chip, thereby improving the sorting and / or enrichment efficiency of the device; and the device has two sorting chips with the same pore diameter of the screening holes 34b, which is also conducive to improving the sorting and enrichment efficiency of the device, so that the device is suitable for processing large-volume samples.

[0119] The device of any one of the above can be used for sorting and enriching the following fluid samples:

[0120] (1) cell clusters in peripheral blood samples;

[0121] (2) cell clusters in pleural effusion, ascitic effusion, lymph fluid, urine or cerebrospinal fluid;

[0122] (3) cell clusters formed after enzymatic digestion of solid tissues; and / or,

[0123] (4) cell clusters formed after single cells are re-aggregated after digestion of solid tissues into single cells.

[0124] In addition, it can also be used for sorting and enriching mixtures of particles of different sizes containing other biological molecules, such as (5) liposomes, water-in-oil microdroplets or oil-in-water microdroplets, etc.

[0125] That is, in addition to providing the above-mentioned device, the present application also provides the use of the above-mentioned device in sorting and enriching the above-mentioned fluid samples.

[0126] Those skilled in the art should understand that in the disclosure of the present application, the terms "first", "second", "third", "fourth", "fifth" and the like are only used to distinguish different structures, and do not limit the number, connection relationship and the like of the specific structure; in addition, the orientation or positional relationship indicated by "longitudinal", "transverse", "upper", "lower", "front", "rear", "left", "right", "vertical", "horizontal", "top", "bottom", "inner", "outer" and the like is based on the orientation or positional relationship shown in the drawings, which is only for the convenience of describing the present application and simplifying the description, and does not indicate or imply that the device or element referred to must have a particular orientation, be constructed and operated in a particular orientation, therefore the above terms cannot be understood as a limitation on the present application.

[0127] Although the embodiments of the present application are described above, the present application is not limited to the specific embodiments and application fields described above, and the specific embodiments described above are only illustrative and guiding, but not limiting. Those skilled in the art can also make many forms under the inspiration of the present application and without departing from the scope protected by the claims of the present application, which all belong to the present application claimed.

Claims

1. An apparatus for sorting and enriching, wherein, The device comprises: a housing; N sorting chips, wherein a screening hole is arranged on each sorting chip, and N sorting chips arranged in sequence divide the internal space of the housing into N+1 cavities, and N≥1; wherein, a first cavity is formed between one side of the housing and the sorting chip adjacent to the side; a second cavity is formed between the other side of the housing and the sorting chip adjacent to the side; a first outlet and a first inlet are formed on the cavity wall of the first cavity, and the first outlet and the first inlet connect the first cavity with the outside world; a second outlet is formed on the cavity wall of the second cavity, and the second outlet connects the second cavity with the outside world.

2. The device of claim 1, wherein, N≥2; a third cavity is formed between any adjacent sorting chips, and a third outlet is formed on the cavity wall of the third cavity, and the third outlet connects the third cavity with the outside world; preferably, at least one third cavity has a third inlet formed on the cavity wall, and the third inlet connects the third cavity with the outside world; more preferably, two or more first inlets are formed on the cavity wall of the first cavity; further preferably, the aperture of the screening hole of each sorting chip gradually increases from one side of the housing to the other side of the housing; and / or, the aperture of the screening hole is greater than or equal to 8 μm; more preferably, the cross-sectional shape of the screening hole is circular or polygonal; and / or, in at least one sorting chip, at least two screening holes have different shapes.

3. The device of claim 2, wherein, the device further comprises a microcolumn, and at least one microcolumn is arranged in at least one of the first cavity, the second cavity, and the third cavity; preferably, the microcolumn is arranged on the housing; and / or, the microcolumn is arranged on the surface of the sorting chip; more preferably, the cross-section of the microcolumn is circular, elliptical, or polygonal.

4. The device of claim 1, wherein, the region where the sorting chip is arranged with the screening hole forms a screening area; and / or, at least one surface of the screening area of at least one sorting chip is partially or entirely non-planar; preferably, at least one surface of the screening area of at least one sorting chip facing the first cavity is partially or entirely non-planar; more preferably, the non-planar structure is a curved surface.

5. An apparatus for sorting and enriching, wherein, The device comprises: a housing; N sorting chips, wherein a screening hole is arranged on each sorting chip, and N sorting chips arranged in sequence divide the internal space of the housing into N+1 cavities, and N≥2; wherein, a first cavity is formed between one side of the housing and the sorting chip adjacent to the side; a second cavity is formed between the other side of the housing and the sorting chip adjacent to the side; a third cavity is formed between any two adjacent sorting chips; a first outlet is formed on the cavity wall of the first cavity, and the first outlet connects the first cavity with the outside world; a second outlet is formed on the cavity wall of the second cavity, and the second outlet connects the second cavity with the outside world. The third cavity is formed with a third outlet on the cavity wall of the third cavity, and the cavity wall of at least one of the third cavities is formed with one or more third inlets connecting the third cavity with the outside world.

6. The device of claim 5, wherein, N = 2; and / or, the cavity wall of at least one of the third cavities is formed with two or more third inlets connecting the third cavity with the outside world.

7. The device of claim 6, wherein, N = 2; and the aperture diameters of the screening holes 34b of the two sorting chips are the same; Preferably, the aperture diameter of the screening hole is greater than or equal to 8 μm; More preferably, the cross-sectional shape of the screening hole is circular, polygonal; and / or, in at least one of the sorting chips, at least two of the screening holes have different shapes.

8. The device of claim 5, wherein, The device further comprises a micro-column, and at least one micro-column is arranged in at least one of the first cavity, the second cavity, and the third cavity; Preferably, the micro-column is arranged on the shell; and / or, the micro-column is arranged on the surface of the sorting chip; More preferably, the cross-section of the micro-column is circular, elliptical, and / or polygonal.

9. The device of claim 5, wherein, The sorting chip is arranged to form a screening area in the region of the screening hole; At least one surface of the screening area of at least one of the sorting chips is partially or entirely non-planar; Preferably, at least one surface of the screening area of at least one of the sorting chips is partially or entirely non-planar; More preferably, the non-planar structure is a curved surface.

10. Use of the device of any one of claims 1-9 in sorting and enriching the following fluid samples: cell clusters in peripheral blood samples; cell clusters in pleural effusion, ascitic effusion, lymphatic fluid, urine, or cerebrospinal fluid; cell clusters formed after enzymatic digestion of solid tissue; cell clusters formed after single cells are re-clustered after digestion of solid tissue into single cells; or, liposomes, water-in-oil microdroplets, or oil-in-water microdroplets.

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