A circulating tumor cell sorting chip integrating inertial and acoustic wave technologies

Abstract

The present invention discloses a circulating tumor cell sorting chip integrating inertial and acoustic wave technology, which relates to the field of microfluidic chip technology, solves the technical problem that the microfluidic chip has low flux and low precision, and the key points of its technical solution are to integrate inertial spiral, concentration module and acoustic sorting three-stage flow channel, combine the advantages of high flux of inertial sorting and high precision of acoustic sorting, and add a concentration module in the middle to further remove background cells and improve the accuracy of acoustic sorting, so as to achieve high-throughput and high-precision sorting of circulating tumor cells at the same time, and overcome the deficiency of the current single sorting technology that cannot achieve high-throughput and high-precision sorting at the same time. At the same time, the present application adopts non-labeling sorting technology, which has the advantages of low cost, easy integration, simple operation, etc., can automatically and quickly sort circulating tumor cells from whole blood, and has high activity, which can be used for subsequent clinical diagnosis, biochemical analysis, drug sensitivity testing and other research applications.

Description

Technical Field

[0001] The present application relates to the field of microfluidic chip technology, and in particular to a circulating tumor cell sorting chip that integrates inertial and acoustic wave technologies. Background Art

[0002] Since the 21st century, cancer has become one of the biggest threats to human health, and countless cancer patients die every year. According to the latest report, there are 18 million new cancer cases and 9.6 million cancer deaths each year in the world, of which 3.8 million new cases and 2.3 million deaths occur in my country. The number of new cancer patients and deaths in my country accounts for more than one-fifth of all cases, which shows that research on cancer is very important and has far-reaching significance.

[0003] Circulating tumor cells refer to various types of tumor cells that actively or passively fall off from the primary tumor lesion and successfully enter the peripheral blood. Through the circulation of peripheral blood, a small number of circulating tumor cells can metastasize and spread. The metastasis and spread of tumor cells is often the main cause of the patient's condition worsening or death. Therefore, the detection of circulating tumor cells in the blood is of great significance for the early diagnosis and prognosis evaluation of cancer patients. The first prerequisite for detecting circulating tumor cells is to remove a large number of background blood cells. Microfluidic technology accurately controls microliter and milliliter samples through micron-level flow channels. Due to its small size and high manipulation accuracy, it is used for cell sorting. Microfluidic sorting technology can be divided into passive sorting technology and active sorting technology according to whether it uses an external force field. Passive sorting technology includes inertial sorting, microstructure filtration, extrusion flow sorting, deterministic lateral displacement sorting, etc. Active sorting technology includes magnetic sorting, acoustic sorting, optical sorting, dielectrophoresis sorting, etc. Passive sorting technology can maintain a certain sorting accuracy under the condition of ensuring high throughput, while active sorting technology sacrifices flux to a certain extent under the condition of ensuring high sorting accuracy. Therefore, it is of great significance to use active and passive sorting technologies to achieve high-throughput and high-precision sorting of circulating tumor cells. Summary of the invention

[0004] The present application provides a circulating tumor cell sorting chip that integrates inertial and acoustic wave technologies, and its technical purpose is to improve the throughput and accuracy of a microfluidic chip used for sorting circulating tumor cells.

[0005] The above technical objectives of this application are achieved through the following technical solutions:

[0006] A circulating tumor cell sorting chip integrating inertial and acoustic wave technologies, comprising an upper cover plate, an upper guide layer, a primary sorting module, an inner outlet guide layer, an outer outlet guide layer, a concentration module, a side outlet guide layer, an intermediate outlet guide layer, an acoustic sorting module and a piezoelectric substrate, which are arranged in sequence from top to bottom;

[0007] The upper cover plate is provided with a sample liquid inlet A and four positioning holes A;

[0008] The upper guide layer is provided with a sample liquid inlet B, 4 straight flow channels, 4 curved flow channels, a sample liquid outlet and 4 positioning holes B; one end of the straight flow channel is connected to the sample liquid inlet B, and the other end is connected to the middle of the curved flow channel; both ends of the curved flow channel are connected to the sample liquid outlet;

[0009] The primary sorting module is provided with at least 8 identical inertial spiral units, each of which comprises a sample liquid inlet C, a spiral flow channel, a spiral unit inner outlet, a spiral unit outer outlet and 4 positioning holes C; one end of the spiral flow channel is connected to the sample liquid inlet C, and the other end is respectively connected to the spiral unit inner outlet and the spiral unit outer outlet;

[0010] The inner outlet guide layer is provided with 4 primary sorting collection liquid inlets A, 4 primary sorting waste liquid inlets A, a primary sorting collection liquid outlet A, a primary sorting collection liquid flow channel and 4 positioning holes D; the primary sorting collection liquid inlet A and the primary sorting collection liquid outlet A are connected through the primary sorting collection liquid flow channel;

[0011] The outer outlet guide layer is provided with 4 primary sorting waste liquid inlets B, a primary sorting waste liquid outlet, a primary sorting collection liquid outlet B, a primary sorting waste liquid flow channel and 4 positioning holes E; the primary sorting waste liquid inlet B and the primary sorting waste liquid outlet are connected through the primary sorting waste liquid flow channel;

[0012] The concentration module is provided with a primary sorting collection liquid inlet B, at least 4 concentration units, a primary sorting waste liquid inlet C and 4 positioning holes F, each concentration unit includes a concentration flow channel and a concentration unit waste liquid flow channel, one end of the concentration flow channel is connected to the primary sorting collection liquid inlet B, and the other end is connected to the concentration unit collection liquid outlet; both ends of the concentration unit waste liquid flow channel are connected to the concentration unit waste liquid outlet;

[0013] The side outlet guide layer is provided with 8 concentration unit waste liquid inlets, a concentration unit waste liquid collection outlet, 4 concentration unit collection liquid inlets A, a concentration unit waste liquid collection flow channel and 4 positioning holes G; the concentration unit waste liquid inlet and the concentration unit waste liquid collection outlet are connected through the concentration unit waste liquid collection flow channel;

[0014] The intermediate outlet guide layer is provided with four concentration unit collection liquid inlets B, a concentration unit collection liquid collection channel, a concentration unit collection liquid collection outlet, a separation and concentration waste liquid outlet and four positioning holes H; the concentration unit collection liquid inlet B and the concentration unit collection liquid collection outlet are connected through the concentration unit collection liquid collection channel;

[0015] The acoustic sorting module is provided with an acoustic sorting sample liquid inlet, a symmetrical sinusoidal flow channel, a direct flow channel, a secondary sorting collection liquid outlet A, a first waste liquid outlet A, a second waste liquid inlet, a second waste liquid flow channel, a second waste liquid outlet A and four positioning holes I; an empty slot structure is provided below the direct flow channel; one end of the symmetrical sinusoidal flow channel is connected to the acoustic sorting sample liquid inlet, and the other end is connected to the direct flow channel; one end of the direct flow channel is respectively connected to the secondary sorting collection liquid outlet A and the first waste liquid outlet A; one end of the second waste liquid flow channel is connected to the second waste liquid inlet, and the other end is connected to the second waste liquid outlet A;

[0016] The piezoelectric substrate is provided with an interdigital transducer, a secondary sorting collection liquid outlet B, a first waste liquid outlet B, a second waste liquid outlet B and four positioning holes J;

[0017] The sample liquid inlet A is connected to the sample liquid inlet B; the sample liquid outlet is connected to the sample liquid inlet C; the spiral unit inner outlet and the spiral unit outer outlet are respectively connected to the primary sorting collection liquid inlet A and the primary sorting waste liquid inlet A; the primary sorting collection liquid outlet A and the primary sorting waste liquid inlet A are respectively connected to the primary sorting collection liquid outlet B and the primary sorting waste liquid inlet B; the primary sorting collection liquid outlet B and the primary sorting waste liquid outlet are connected to the primary sorting collection liquid inlet B and the primary sorting waste liquid inlet C; the concentration unit collection liquid outlet and the concentration unit waste liquid outlet are respectively connected to the primary sorting collection liquid outlet B and the primary sorting waste liquid outlet. The two-stage sorting collecting liquid outlet A is connected to the second waste liquid inlet and the acoustic sorting sample liquid inlet respectively; the two-stage sorting collecting liquid outlet A, the first waste liquid outlet A and the second waste liquid outlet A are connected to the two-stage sorting collecting liquid outlet B, the first waste liquid outlet B and the second waste liquid outlet B respectively; the interdigital transducer is arranged in the empty slot structure.

[0018] The beneficial effects of the present application are as follows: the circulating tumor cell sorting chip integrating inertial and acoustic wave technologies described in the present application includes, from top to bottom, an upper cover plate, an upper guide layer, a primary sorting module, an inner outlet guide layer, an outer outlet guide layer, a concentration module, a side outlet guide layer, a middle outlet guide layer, an acoustic sorting module and a piezoelectric substrate. A sample liquid inlet A is provided at the center of the upper cover plate, and the sample liquid is injected into the primary sorting module through the upper guide layer; the primary sorting module includes 8 identical inertial spiral flow channels, and the sample liquid passes through the spiral flow channels to achieve the first-level sorting, a large number of blood cells contained in the sample liquid flow out from the outer outlet of the spiral flow channel and are removed, and circulating tumor cells and some blood cells flow into the next level from the inner outlet of the spiral flow channel; the inner outlet guide layer guides the liquid at the inner outlet of the spiral flow channel into the concentration module; the outer outlet guide layer collects the waste liquid at the outer outlet of the spiral flow channel; the concentration module includes 4 asymmetric sinusoidal flow channels, so that blood cells flow out from the two side outlets, and circulating tumor cells and some blood cells flow out from the middle outlet; the side outlet guide layer collects the waste liquid at the two side outlets of the concentration module and guides it downward; the middle outlet guide layer guides the sample liquid at the middle outlet of the concentration module into the acoustic sorting module; the acoustic sorting module first allows the sample liquid to flow into the sinusoidal flow channel to focus the cells, and then achieves the second-level sorting through acoustic sorting, thereby further improving the sorting accuracy of circulating tumor cells. This application combines active sorting technology with passive sorting technology to effectively improve the sorting accuracy of circulating tumor cells, and adopts multi-core sorting and chip vertical stacking methods to further improve the throughput of the chip. At the same time, the chip design of this application is flexible and novel in structure, and can be used for commercial mass production.

[0019] This application integrates the inertial spiral, concentration module and acoustic sorting three-stage flow channel, combines the advantages of high throughput of inertial sorting and high precision of acoustic sorting, and adds a concentration module in the middle to further remove background cells and improve the accuracy of acoustic sorting, so as to achieve high-throughput and high-precision sorting of circulating tumor cells at the same time, overcoming the shortcomings of the current single sorting technology that cannot achieve high-throughput and high-precision sorting at the same time. At the same time, this application adopts non-labeling sorting technology, which has the advantages of low cost, easy integration, simple operation, etc., and can automatically and quickly sort circulating tumor cells from whole blood, and has high activity, which can be used for subsequent clinical diagnosis, biochemical analysis, drug sensitivity testing and other research applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is an exploded diagram of the present application;

[0021] Figure 2 It is a structural schematic diagram of the upper cover;

[0022] Figure 3 Schematic diagram of the structure of the upper guide layer;

[0023] Figure 4It is a structural schematic diagram of the primary sorting module;

[0024] Figure 5 Schematic diagram of the structure of the inner outlet guide layer;

[0025] Figure 6 Schematic diagram of the structure of the external outlet guide layer;

[0026] Figure 7 It is a schematic diagram of the structure of the concentration module;

[0027] Figure 8 It is a schematic diagram of the structure of the side outlet guide layer;

[0028] Fig. 9 It is a schematic diagram of the structure of the middle outlet guide layer;

[0029] Fig.10 It is a structural diagram of the acoustic sorting module;

[0030] Fig.11 is a schematic diagram of the structure of a piezoelectric substrate;

[0031] In the figure: 1-upper cover plate; 2-upper guide layer; 3-primary sorting module; 4-inner outlet guide layer; 5-outer outlet guide layer; 6-concentration module; 7-side outlet guide layer; 8-middle outlet guide layer; 9-acoustic sorting module; 10-piezoelectric substrate; 11-sample liquid inlet A; 12-positioning hole A; 21-sample liquid inlet B; 22-straight channel; 23-curved channel; 24-sample liquid outlet; 25-positioning hole B; 31-sample liquid inlet C; 32-spiral channel; 33-spiral single channel Element inner outlet; 34-spiral unit outer outlet; 35-positioning hole C; 41-primary sorting collection liquid inlet A; 42-primary sorting waste liquid inlet A; 43-primary sorting collection liquid outlet A; 44-primary sorting collection liquid flow channel; 45-positioning hole D; 51-primary sorting waste liquid inlet B; 52-primary sorting waste liquid outlet; 53-fifth primary sorting collection liquid outlet B; 54-primary sorting waste liquid flow channel; 55-positioning hole E; 61-primary sorting collection liquid inlet B; 62-concentration flow channel ; 63-concentration unit collection liquid outlet; 64-concentration unit waste liquid flow channel; 65-concentration unit waste liquid outlet; 66-primary sorting waste liquid inlet C; 67-positioning hole F; 71-concentration unit waste liquid inlet; 72-concentration unit waste liquid collection outlet; 73-concentration unit collection liquid inlet A; 74-concentration unit waste liquid collection flow channel; 75-positioning hole G; 81-concentration unit collection liquid inlet B; 82-concentration unit collection liquid collection flow channel; 83-concentration unit collection liquid collection outlet; 84- Outlet for sorting and concentrating waste liquid; 85-positioning hole H; 91-inlet for acoustic sorting sample liquid; 92-symmetrical sinusoidal flow channel; 93-direct flow channel; 94-secondary sorting collection liquid outlet A; 95-first waste liquid outlet A; 96-second waste liquid inlet; 97-second waste liquid flow channel; 98-second waste liquid outlet A; 99-positioning hole I; 101-interdigital transducer; 102-secondary sorting collection liquid outlet B; 103-first waste liquid outlet B; 104-second waste liquid outlet B; 105-positioning hole J. DETAILED DESCRIPTION

[0032] The technical solution of the present application will be described in detail below with reference to the accompanying drawings.

[0033] like Figure 1 As shown, a circulating tumor cell sorting chip integrating inertial and acoustic wave technologies according to an embodiment of the present application includes an upper cover plate 1, an upper guide layer 2, a primary sorting module 3, an inner outlet guide layer 4, an outer outlet guide layer 5, a concentration module 6, a side outlet guide layer 7, an intermediate outlet guide layer 8, an acoustic sorting module 9 and a piezoelectric substrate 10, which are arranged in sequence from top to bottom.

[0034] As a specific embodiment, the upper cover plate 1, upper guide layer 2, primary sorting module 3, inner outlet guide layer 4, outer outlet guide layer 5, concentration module 6, side outlet guide layer 7, middle outlet guide layer 8, and acoustic sorting module 9 are all made of polymer film materials.

[0035] like Figure 2 As shown, the upper cover plate 1 is provided with a sample liquid inlet A11 and four positioning holes A12.

[0036] like Figure 3 As shown, the upper guide layer 2 is provided with a sample liquid inlet B21, four straight channels 22, four curved channels 23, a sample liquid outlet 24 and four positioning holes B25. One end of the straight channel 22 is connected to the sample liquid inlet B21, and the other end is connected to the middle of the curved channel 23; both ends of the curved channel 23 are connected to the sample liquid outlet 24. The four straight channels 22 are connected to the four curved channels 23 to form eight channels of equal length, so that the sample liquid flows from the sample liquid inlet B21 to the sample liquid outlet 24.

[0037] like Figure 4 As shown, at least 8 identical inertial spiral units are provided on the primary sorting module 3, each of which includes a sample liquid inlet C31, a spiral flow channel 32, a spiral unit inner outlet 33, a spiral unit outer outlet 34, and 4 positioning holes C35. One end of the spiral flow channel 32 is connected to the sample liquid inlet C31, and the other end is connected to the spiral unit inner outlet 33 and the spiral unit outer outlet 34, respectively.

[0038] As a specific embodiment, the spiral flow channel 32, the spiral unit inner outlet 33 and the spiral unit outer outlet 34 are in a Y-shaped structure. The spiral unit inner outlet 33 of each spiral flow channel 32 is connected to the spiral unit inner outlet 33 of the spiral flow channel 32 adjacent to the left, and the spiral unit outer outlet 34 of each spiral flow channel 32 is connected to the spiral unit outer outlet 34 of the spiral flow channel 32 adjacent to the right. The vertical cross-section of the spiral flow channel 32 is a rectangular structure or a trapezoidal structure with a width greater than a height.

[0039] like Figure 5 As shown, four primary sorting collection liquid inlets A41, four primary sorting waste liquid inlets A42, a primary sorting collection liquid outlet A43, a primary sorting collection liquid flow channel 44 and four positioning holes D45 are provided on the inner outlet guide layer 4. The primary sorting collection liquid inlet A41 and the primary sorting collection liquid outlet A43 are connected through the primary sorting collection liquid flow channel 44.

[0040] As a specific embodiment, the primary sorting collection liquid flow channel 44 is in an "H"-shaped structure, and the middle of the transverse flow channel represented by "-" in the "H"-shaped structure is connected to the primary sorting collection liquid outlet A43, and its four end points are respectively connected to the primary sorting collection liquid inlet A41.

[0041] like Figure 6 As shown, four primary sorting waste liquid inlets B51, a primary sorting waste liquid outlet 52, a primary sorting collection liquid outlet B53, a primary sorting waste liquid flow channel 54 and four positioning holes E55 are provided on the outer outlet guide layer 5. The primary sorting waste liquid inlet B51 and the primary sorting waste liquid outlet 52 are connected through the primary sorting waste liquid flow channel 54.

[0042] As a specific embodiment, the primary sorting waste liquid flow channel 54 is in an "H"-shaped structure, and the transverse flow channel represented by "-" in the "H"-shaped structure is in a "︿" shape, with its four end points respectively connected to the primary sorting waste liquid inlet B51, and the middle of the "︿" shape connected to the primary sorting waste liquid outlet 52.

[0043] like Figure 7 As shown, the concentration module 6 is provided with a primary sorting collection liquid inlet B61, at least 4 concentration units, a primary sorting waste liquid inlet C66 and 4 positioning holes F67. Each concentration unit includes a concentration flow channel 62 and a concentration unit waste liquid flow channel 64. One end of the concentration flow channel 62 is connected to the primary sorting collection liquid inlet B61, and the other end is connected to the concentration unit collection liquid outlet 63; both ends of the concentration unit waste liquid flow channel 64 are connected to the concentration unit waste liquid outlet 65.

[0044] As a specific embodiment, the concentration channel 62 is an asymmetric sinusoidal structure.

[0045] like Figure 8 As shown, eight concentration unit waste liquid inlets 71, a concentration unit waste liquid collection outlet 72, four concentration unit collection liquid inlets A73, a concentration unit waste liquid collection channel 74 and four positioning holes G75 are provided on the side outlet guide layer 7. The concentration unit waste liquid inlet 71 and the concentration unit waste liquid collection outlet 72 are connected through the concentration unit waste liquid collection channel 74.

[0046] As a specific embodiment, the symmetry center of the concentrating unit waste liquid collecting channel 74 is connected to the concentrating unit waste liquid collecting outlet 72; and the eight ends of the concentrating unit waste liquid collecting channel 74 are respectively connected to the concentrating unit waste liquid inlet 71.

[0047] like Fig. 9 As shown, four concentration unit collection liquid inlets B81, a concentration unit collection liquid collection channel 82, a concentration unit collection liquid collection outlet 83, a separation and concentration waste liquid outlet 84 and four positioning holes H85 are provided on the middle outlet guide layer 8. The concentration unit collection liquid inlet B81 and the concentration unit collection liquid collection outlet 83 are connected through the concentration unit collection liquid collection channel 82.

[0048] As a specific embodiment, the concentration unit collected liquid collection channel 82 is in a "cross" shape, the symmetry center is connected to the concentration unit collected liquid collection outlet 83, and the four end points are respectively connected to the concentration unit collected liquid inlet B81.

[0049] like Fig.10 As shown, the acoustic sorting module 9 is provided with an acoustic sorting sample liquid inlet 91, a symmetrical sinusoidal flow channel 92, a straight flow channel 93, a secondary sorting collection liquid outlet A94, a first waste liquid outlet A95, a second waste liquid inlet 96, a second waste liquid flow channel 97, a second waste liquid outlet A98 and four positioning holes I99.

[0050] An empty slot structure 910 is provided below the direct flow channel 93. One end of the symmetrical sinusoidal flow channel 92 is connected to the acoustic sorting sample liquid inlet 91, and the other end is connected to the direct flow channel 93; one end of the direct flow channel 93 is respectively connected to the secondary sorting collection liquid outlet A94 and the first waste liquid outlet A95; one end of the second waste liquid flow channel 97 is connected to the second waste liquid inlet 96, and the other end is connected to the second waste liquid outlet A98.

[0051] As a specific embodiment, the direct flow channel 93, the secondary sorting collection liquid outlet A94 and the first waste liquid outlet A95 are in a "Y" shape.

[0052] like Fig.11 As shown, the piezoelectric substrate 10 is provided with an interdigital transducer 101, a secondary sorting collection liquid outlet B102, a first waste liquid outlet B103, a second waste liquid outlet B104 and four positioning holes J105.

[0053] The vertical distance between the direct current channel (93) and the interdigital transducer 101 is 1 mm. The interdigital transducer 101 includes a plurality of pairs of interdigits. The piezoelectric substrate 10 is made of 128°-cut lithium niobate material, and the interdigital transducer 101 is manufactured on the surface of the lithium niobate by sputtering and stripping technology.

[0054] like Figure 1As shown, a circulating tumor cell sorting chip integrating inertial and acoustic wave technologies of an embodiment of the present application is assembled sequentially from bottom to top through four positioning holes set on each layer, wherein the sample liquid inlet A11 of the upper cover plate 1 is connected to the sample liquid inlet B21 of the upper guide layer 2, the sample liquid outlet 24 of the upper guide layer 2 is connected to the sample liquid inlet C31 of the primary sorting module 3, the spiral unit inner outlet 33 and the spiral unit outer outlet 34 of the primary sorting module 3 are respectively connected to the primary sorting collection liquid inlet A41 and the primary sorting waste liquid inlet A42 of the inner outlet guide layer 4, the primary sorting collection liquid outlet A43 and the primary sorting waste liquid inlet A42 of the inner outlet guide layer 4 are respectively connected to the primary sorting collection liquid outlet B53 and the primary sorting waste liquid inlet B51 of the outer outlet guide layer 5, the primary sorting collection liquid outlet B53 and the primary sorting waste liquid outlet 52 of the outer outlet guide layer 5 are connected to the primary sorting collection liquid inlet A41 of the concentration module 6, and the primary sorting collection liquid outlet B53 and the primary sorting waste liquid outlet 52 of the concentration module 6 are connected to the primary sorting collection liquid inlet A41 of the concentration module 6. The concentrating unit collecting liquid outlet 63 and the concentrating unit waste liquid outlet 65 of the concentrating module 6 are respectively connected with the concentrating unit collecting liquid inlet A73 and the concentrating unit waste liquid inlet 71 of the side outlet guide layer 7; the concentrating unit collecting liquid inlet A73 and the concentrating unit waste liquid collection outlet 72 of the side outlet guide layer 7 are respectively connected with the concentrating unit collecting liquid inlet B81 and the sorting concentrated waste liquid outlet 84 of the middle outlet guide layer 8; the sorting concentrated waste liquid outlet 84 and the concentrating unit collecting liquid collection outlet 83 of the middle outlet guide layer 8 are respectively connected with the second waste liquid inlet 96 and the acoustic sorting sample liquid inlet 91 of the acoustic sorting module 9; the secondary sorting collecting liquid outlet A94, the first waste liquid outlet A95 and the second waste liquid outlet A98 of the acoustic sorting module 9 are respectively connected with the secondary sorting collecting liquid outlet B102, the first waste liquid outlet B103 and the second waste liquid outlet B104 of the piezoelectric substrate 10.

[0055] like Figure 4The diagram shows the structure of the primary sorting module. The sample liquid enters the spiral flow channel 32 from the sample liquid inlet C31 of each spiral sorting unit. The fluid velocity near the center line of the flow channel is higher than the velocity near the wall of the flow channel. Therefore, it flows outward under the combined action of centrifugal force and radial pressure gradient imbalance. Due to the conservation of mass in the closed flow channel, the fluid near the outer wall will be squeezed to generate backflow along the upper and lower walls of the spiral flow channel 32, and finally form two vortices with opposite rotation directions in the vertical direction, namely Dean flow. At the same time, since the flow velocity of the fluid from the center of the flow channel to the wall of the flow channel is a parabolic distribution, the velocity gradient generated thereby forms a shear induced lift pointing to the wall of the flow channel. In addition, due to the interaction between the cell and the wall of the flow channel, the wall induced lift is formed. The shear induced lift drives the cell to move toward the wall, and the wall induced lift makes the cell move away from the wall when it approaches the wall. The combined force of these two lifts is called inertial lift FL. After the sample liquid enters the spiral flow channel 32, under the combined effect of the inertial lift FL and the Dean drag FD generated by the Dean flow, cells of different sizes in the flow channel will stabilize at different equilibrium positions. In this embodiment, the circulating tumor cells with larger sizes migrate to the equilibrium position close to the inner wall surface, and the blood cells with smaller sizes migrate to the equilibrium position close to the outer wall surface. Therefore, the collection liquid containing circulating tumor cells flows from the inner outlet 33 of the spiral unit to the next level, and the waste liquid containing a large amount of background blood cells flows out from the outer outlet 34 of the spiral unit.

[0056] like Figure 7 The diagram shows the structure of the concentration module. The collected liquid obtained by the primary sorting module enters the concentration module 6 from the primary sorting collected liquid inlet B61. The primary sorting collected liquid first enters the concentration channel 62. The concentration channel 62 adopts an asymmetric sinusoidal structure. The cells in the channel are simultaneously affected by the inertial lift FL and the Dean drag FD generated by the Dean flow. By designing the channel size and controlling the appropriate flow rate, the larger circulating tumor cells can be focused at the center of the channel and flow from the concentration unit collection liquid outlet 63 to the next level, and the small blood cells can flow out from the concentration unit waste liquid outlet 65. A concentration unit waste liquid channel 64 of a certain length is provided between the terminal outlet of the concentration channel 62 and the concentration unit waste liquid outlet 65 to match the flow resistance so that the fluid in the channel meets the flow rate requirements. In addition, the concentration module 6 is provided with 4 concentration units. After the collected liquid obtained by the primary sorting module is concentrated by the 4 concentration units, most of the collected liquid containing background blood cells flows out from the waste liquid outlet 65 of the concentration unit, and the remaining small volume of the collected liquid containing circulating tumor cells enters the acoustic sorting module 9. The purpose of setting multiple concentration units is to increase the concentration multiple. Since the acoustic sorting has a smaller flux than the inertial sorting, the concentration module 6 is used to make the sample liquid flux entering the acoustic sorting meet the requirements.

[0057] like Fig.10 The structure diagram of the acoustic sorting module is shown in FIG. The acoustic sorting module 9 is provided with a symmetrical sinusoidal flow channel 92 and a direct flow channel 93. The sample liquid obtained by the concentration module 6 first enters the symmetrical sinusoidal flow channel 92 from the acoustic sorting sample liquid inlet 91. The cells are simultaneously acted upon by the inertial lift FL and the Dean drag FD generated by the Dean flow in the flow channel, so that the cells are focused to the center of the flow channel and enter the direct flow channel 93. A group of interdigital transducers 101 is provided 1 mm below the direct flow channel 93. When an alternating current signal is applied to the interdigital transducers 101, surface acoustic waves are generated on the piezoelectric substrate according to the inverse piezoelectric effect. When the cells in the sample liquid pass through the direct flow channel 93, they are acted upon by the acoustic radiation force, which is positively correlated with the cell volume. Therefore, the acoustic radiation force on the circulating tumor cells with a larger volume is much greater than that on the blood cells, causing the circulating tumor cells to migrate upward and flow out from the secondary sorting collection liquid outlet A94, while the effect of the acoustic radiation force on the blood cells is not obvious, and they can maintain their original motion trajectory and flow out from the first waste liquid outlet A95.

[0058] like Figure 1 As shown, the working process of a circulating tumor cell sorting chip integrating inertial and acoustic wave technology according to an embodiment of the present invention is as follows:

[0059] 1. The syringe injects the sample liquid into the chip from the sample liquid inlet A11 at a certain flow rate. After passing through the straight channel 22 and the curved channel 23 of the upper guide layer 2, the sample liquid is divided into 8 parts and injected into the sample liquid inlet C31 of the 8 inertial spiral units on the primary sorting module 3. After the sample liquid passes through the spiral flow channel 32, the cells are simultaneously affected by the inertial lift FL and the Dean drag FD generated by the Dean flow. The sample liquid containing circulating tumor cells flows out from the spiral unit inner outlet 33, and the waste liquid containing a large amount of background blood cells flows out from the spiral unit outer outlet 34, completing the primary sorting of the sample liquid.

[0060] 2. The sample liquid obtained after primary sorting is collected together through the inner outlet guide layer 4 and flows into the primary sorting collection liquid inlet B61 of the concentration module 6, and the outer outlet guide layer 5 collects the waste liquid at the outer outlet and flows to the primary sorting waste liquid inlet C66 of the concentration module 6. The collected liquid obtained by primary sorting is concentrated through four concentration units on the concentration module 6. The collected liquid passes through the asymmetric sinusoidal structure concentration flow channel 62. Under the combined action of the inertial lift FL and the Dean drag FD generated by the Dean flow, the collected liquid containing circulating tumor cells flows out from the concentration unit collection liquid outlet 63, and the waste liquid containing blood cells flows out from the concentration unit waste liquid outlet 65 through the concentration unit waste liquid flow channel 64. After passing through the concentration module 6, the injected high-throughput sample liquid is greatly concentrated, and the waste liquid containing background blood cells does not enter the next level of sorting.

[0061] 3. The waste liquid obtained by the concentration module 6 is collected together through the waste liquid inlet 71 of the concentration unit of the side outlet guide layer 7 and flows to the second waste liquid inlet 96 of the acoustic sorting module 9. The middle outlet guide layer 8 collects the concentrated collected liquid through the collection liquid inlet B81 of the concentration unit and flows to the acoustic sorting sample liquid inlet 91 of the acoustic sorting module 9. The concentrated collected liquid first passes through the symmetrical sinusoidal flow channel 92. The cells can be focused to the center of the flow channel under the combined action of the inertial lift FL and the Dean drag FD generated by the Dean flow, and enter the direct flow channel 93. In the direct flow channel 93, an AC signal is applied to the interdigital transducer 101, and the circulating tumor cells migrate upward under the action of the acoustic radiation force and flow out from the secondary sorting collection liquid outlet A94, while the blood cells are not significantly affected by the acoustic radiation force and flow out from the first waste liquid outlet A95 along the original path. The waste liquid obtained after primary sorting and concentration flows out from the second waste liquid outlet A98 through the second waste liquid flow channel 97. This enables high-throughput and high-precision sorting of circulating tumor cells.

[0062] The above shows and describes the basic principles, main features and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above specific embodiments. The above specific embodiments and the description in the specification are only for further illustrating the principles of the present invention. Without departing from the spirit and scope of the present invention, the present invention may have various changes and improvements, which fall within the scope of the present invention to be protected. The scope of the present invention to be protected is defined by the claims and their equivalents.

Claims

1. A circulating tumor cell sorting chip integrating inertial and acoustic wave technology, characterized in that: It comprises an upper cover plate (1), an upper guide layer (2), a primary sorting module (3), an inner outlet guide layer (4), an outer outlet guide layer (5), a concentration module (6), a side outlet guide layer (7), an intermediate outlet guide layer (8), an acoustic sorting module (9) and a piezoelectric substrate (10) which are arranged in sequence from top to bottom; The upper cover plate (1) is provided with a sample liquid inlet A (11) and four positioning holes A (12); The upper guide layer (2) is provided with a sample liquid inlet B (21), four straight flow channels (22), four curved flow channels (23), a sample liquid outlet (24) and four positioning holes B (25); one end of the straight flow channel (22) is connected to the sample liquid inlet B (21), and the other end is connected to the middle of the curved flow channel (23); both ends of the curved flow channel (23) are connected to the sample liquid outlet (24); The primary sorting module (3) is provided with at least 8 identical inertial spiral units, each of which comprises a sample liquid inlet C (31), a spiral flow channel (32), a spiral unit inner outlet (33), a spiral unit outer outlet (34) and 4 positioning holes C (35); one end of the spiral flow channel (32) is connected to the sample liquid inlet C (31), and the other end is respectively connected to the spiral unit inner outlet (33) and the spiral unit outer outlet (34); The inner outlet guide layer (4) is provided with four primary sorting collection liquid inlets A (41), four primary sorting waste liquid inlets A (42), a primary sorting collection liquid outlet A (43), a primary sorting collection liquid flow channel (44) and four positioning holes D (45); the primary sorting collection liquid inlet A (41) and the primary sorting collection liquid outlet A (43) are connected through the primary sorting collection liquid flow channel (44); The outer outlet guide layer (5) is provided with four primary sorting waste liquid inlets B (51), a primary sorting waste liquid outlet (52), a primary sorting collection liquid outlet B (53), a primary sorting waste liquid flow channel (54) and four positioning holes E (55); the primary sorting waste liquid inlet B (51) and the primary sorting waste liquid outlet (52) are connected via the primary sorting waste liquid flow channel (54); The concentration module (6) is provided with a primary sorting collection liquid inlet B (61), at least four concentration units, a primary sorting waste liquid inlet C (66) and four positioning holes F (67); each concentration unit comprises a concentration flow channel (62) and a concentration unit waste liquid flow channel (64); one end of the concentration flow channel (62) is connected to the primary sorting collection liquid inlet B (61), and the other end is connected to the concentration unit collection liquid outlet (63); both ends of the concentration unit waste liquid flow channel (64) are connected to the concentration unit waste liquid outlet (65); The side outlet guide layer (7) is provided with eight concentration unit waste liquid inlets (71), a concentration unit waste liquid collection outlet (72), four concentration unit collection liquid inlets A (73), a concentration unit waste liquid collection channel (74) and four positioning holes G (75); the concentration unit waste liquid inlets (71) and the concentration unit waste liquid collection outlet (72) are connected via the concentration unit waste liquid collection channel (74); The intermediate outlet guide layer (8) is provided with four concentration unit collection liquid inlets B (81), a concentration unit collection liquid collection channel (82), a concentration unit collection liquid collection outlet (83), a separation and concentration waste liquid outlet (84) and four positioning holes H (85); the concentration unit collection liquid inlet B (81) and the concentration unit collection liquid collection outlet (83) are connected via the concentration unit collection liquid collection channel (82); The acoustic sorting module (9) is provided with an acoustic sorting sample liquid inlet (91), a symmetrical sinusoidal flow channel (92), a direct flow channel (93), a secondary sorting collection liquid outlet A (94), a first waste liquid outlet A (95), a second waste liquid inlet (96), a second waste liquid flow channel (97), a second waste liquid outlet A (98) and four positioning holes I (99); a hollow groove structure (910) is provided below the direct flow channel (93); one end of the symmetrical sinusoidal flow channel (92) is connected to the acoustic sorting sample liquid inlet (91), and the other end is connected to the direct flow channel (93); one end of the direct flow channel (93) is respectively connected to the secondary sorting collection liquid outlet A (94) and the first waste liquid outlet A (95); one end of the second waste liquid flow channel (97) is connected to the second waste liquid inlet (96), and the other end is connected to the second waste liquid outlet A (98); The piezoelectric substrate (10) is provided with an interdigital transducer (101), a secondary sorting collection liquid outlet B (102), a first waste liquid outlet B (103), a second waste liquid outlet B (104) and four positioning holes J (105); The sample liquid inlet A (11) is connected to the sample liquid inlet B (21); the sample liquid outlet (24) is connected to the sample liquid inlet C (31); the spiral unit inner outlet (33) and the spiral unit outer outlet (34) are respectively connected to the primary sorting collection liquid inlet A (41) and the primary sorting waste liquid inlet A (42); the primary sorting collection liquid outlet A (43) and the primary sorting waste liquid inlet A (42) are respectively connected to the primary sorting collection liquid outlet B (53) and the primary sorting waste liquid inlet B (51); the primary sorting collection liquid outlet B (53) and the primary sorting waste liquid outlet (52) are respectively connected to the primary sorting collection liquid inlet B (61) and the primary sorting waste liquid inlet C (66); the concentration unit collection liquid outlet (63) and the concentration unit waste liquid outlet (65) are respectively connected to the primary sorting collection liquid outlet B (53) and the primary sorting waste liquid inlet B (51). The concentration unit collection liquid inlet A (73) and the concentration unit waste liquid inlet (71) are connected; the concentration unit collection liquid inlet A (73) and the concentration unit waste liquid collection outlet (72) are connected to the concentration unit collection liquid inlet B (81) and the separation concentration waste liquid outlet (84); the separation concentration waste liquid outlet (84) and the concentration unit collection liquid collection outlet (83) are respectively connected to the second waste liquid inlet (96) and the acoustic separation sample liquid inlet (91); the secondary separation collection liquid outlet A (94), the first waste liquid outlet A (95) and the second waste liquid outlet A (98) are respectively connected to the secondary separation collection liquid outlet B (102), the first waste liquid outlet B (103) and the second waste liquid outlet B (104); and the interdigital transducer (101) is arranged in the empty slot structure (910).

2. The sorting chip according to claim 1, characterized in that: The lengths of the straight channel (22) and the curved channel (23) are equal.

3. The sorting chip according to claim 1, characterized in that: The spiral flow channel (32), the spiral unit inner outlet (33) and the spiral unit outer outlet (34) are in a Y-shaped structure; the spiral unit inner outlet (33) of each spiral flow channel (32) is connected to the spiral unit inner outlet (33) of the spiral flow channel (32) adjacent to the left, and the spiral unit outer outlet (34) of each spiral flow channel (32) is connected to the spiral unit outer outlet (34) of the spiral flow channel (32) adjacent to the right; the vertical cross section of the spiral flow channel (32) is a rectangular structure or a trapezoidal structure with a width greater than a height.

4. The sorting chip according to claim 1, characterized in that: The primary sorting collection liquid flow channel (44) is in an "H"-shaped structure. The middle of the transverse flow channel represented by "-" in the "H"-shaped structure is connected to the primary sorting collection liquid outlet A (43), and its four end points are respectively connected to the primary sorting collection liquid inlet A (41).

5. The sorting chip according to claim 1, characterized in that: The primary sorting waste liquid flow channel (54) is in an "H"-shaped structure. The transverse flow channel indicated by "-" in the "H"-shaped structure is in a "︿" shape. Its four end points are respectively connected to the primary sorting waste liquid inlet B (51), and the middle of the "︿" shape is connected to the primary sorting waste liquid outlet (52).

6. The sorting chip according to claim 1, characterized in that: The concentrating flow channel (62) is an asymmetric sinusoidal structure.

7. The sorting chip according to claim 1, characterized in that: The symmetry center of the concentrating unit waste liquid collection channel (74) is connected to the concentrating unit waste liquid collection outlet (72); and the eight ends of the concentrating unit waste liquid collection channel (74) are respectively connected to the concentrating unit waste liquid inlet (71).

8. The sorting chip according to claim 1, characterized in that: The concentrated unit collected liquid collection channel (82) is in a "cross"-shaped structure, with its symmetry center connected to the concentrated unit collected liquid collection outlet (83), and its four end points respectively connected to the concentrated unit collected liquid inlet B (81).

9. The sorting chip according to claim 1, characterized in that: The direct flow channel (93) forms a "Y"-shaped structure with the secondary sorting collection liquid outlet A (94) and the first waste liquid outlet A (95).

10. The sorting chip according to claim 1, characterized in that: The vertical distance between the direct current channel (93) and the interdigital transducer (101) is 1 mm, and the interdigital transducer (101) comprises a plurality of pairs of interdigital fingers; the piezoelectric substrate (10) is made of 128°-cut lithium niobate material, and the interdigital transducer (101) is manufactured on the surface of the lithium niobate by sputtering and stripping technology.

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