A micro-particle capture chip, system and method

Abstract

This application discloses a microparticle capture chip, system, and method. The microparticle capture chip includes a micro-pit layer and a top cap layer. A micro-pit array is formed at multiple locations on the first surface of the micro-pit layer. The top cap layer covers the first surface of the micro-pit layer. On the side of the top cap layer facing the micro-pit layer, multiple flow channels are formed, each corresponding to a different micro-pit array. The flow channels extend linearly and are parallel to each other. At each end of the flow channel are a first opening and a second opening, respectively, extending to the side of the top cap layer facing away from the micro-pit layer. The flow channels and the corresponding micro-pit arrays form independent sample loading cavities. The microparticle capture chip of this application can provide helpful assistance in constructing a platform that simultaneously possesses high throughput, high single-particle resolution, and low operator skill requirements.

Description

Technical Field

[0001] This application relates to the field of microparticle capture technology, and in particular to a microparticle capture chip, system and method. Background Technology

[0002] To accurately and precisely reconstruct the composition and regulatory mechanisms of functional biological units, and to clarify the expression networks and dynamic interactions between cells, single-cell level transcriptional analysis has been developed as an ideal technique to accomplish this task with the highest resolution and sensitivity. This powerful, high-resolution tool is widely used in various biological fields, including cell development, immune regulation, neuroscience, cancer heterogeneity and microenvironment, and epigenetics.

[0003] Traditional single-cell isolation methods primarily rely on manual picking or fluorescence-activated cell sorting; however, limitations in efficiency, throughput, and long operating times often hinder their widespread application. In recent years, highly integrated microfluidic chips, due to their size matching cells, have proven capable of isolating single cells in a highly efficient, low-consumption, low-external-contamination, highly compatible, and economical manner.

[0004] To date, microfluidic chips based on different separation principles have been developed. For example, some chips use physical structures such as tiny obstacles or microdams to capture single cells in microfluidics; some use precisely controlled microdroplets to encapsulate single cells; some use integrated photoelectron tweezers or dielectrophoresis to separate cells; and some use micropores or chambers the size of cells to immobilize cells that have settled due to gravity.

[0005] However, the practical application of these devices in clinical or general biological laboratories is still limited by four main drawbacks. First, most devices require pumps or pressure control systems to introduce and guide the liquid, and some also require specialized electronic or optical instruments. These auxiliary devices are not common in routine medical or biological laboratories, increasing the difficulty of operation. Second, many microfluidic chips are complex in structure due to their multi-layered structures, channels, and valves, making them difficult to manufacture and operate. Third, auxiliary components occupy a lot of space, significantly reducing the area density of single-cell arrays. Last but not least, many devices have less than ideal performance. For example, some devices with dense micro-barriers or dams have high throughput but low sample utilization; some devices that rely on gravity sedimentation have simple structures but low or medium single-cell resolution and long separation times; and some devices have low throughput and a limited number of cell capture sites. Therefore, building a platform that simultaneously offers high throughput, high single-cell resolution, and low operator skill requirements remains a challenge. Summary of the Invention

[0006] In view of this, this application provides a microparticle capture chip, system and method, which can help build a platform that has high throughput, high single microparticle resolution and low technical requirements for operators.

[0007] To achieve the above objectives, this application provides the following technical solution:

[0008] A microparticle capture chip, comprising:

[0009] A micro-pit layer, wherein a micro-pit array is provided at multiple locations on the first surface of the micro-pit layer;

[0010] A top cover layer covers the first surface. The top cover layer has multiple flow channels on the side facing the micro-pit layer, each corresponding to a micro-pit array. The flow channels extend in a straight line and are parallel to each other. The two ends of each flow channel have a first opening and a second opening that extend to the side of the top cover layer facing away from the micro-pit layer. The flow channels and the corresponding micro-pit arrays form independent sample loading cavities.

[0011] Optionally, the above-mentioned microparticle capture chip includes:

[0012] A support layer is stacked on the second surface of the micro-pit layer, the second surface facing opposite to the first surface, and the stiffness of the support layer is greater than that of the micro-pit layer.

[0013] Optionally, the above-mentioned microparticle capture chip includes:

[0014] The bottom plate layer is provided with a sink trough for accommodating the support layer and the micro-pit layer, and the bottom of the sink trough is provided with an observation port.

[0015] Optionally, in the above-mentioned microparticle capture chip, at least one of the micro-pit arrays is composed of a plurality of first micro-pit units, wherein the first micro-pit unit is configured as a single-layer hole.

[0016] Optionally, in the above-mentioned microparticle capture chip, at least one of the micro-pit arrays is composed of a plurality of second micro-pit units, the second micro-pit units being configured as double-layer holes, the double-layer holes including a first hole connected to the first surface and a second hole connected to the bottom of the first hole, the area of ​​the second hole being smaller than the area of ​​the first hole.

[0017] Optionally, in the above-mentioned microparticle capture chip, the first hole is a regular hexagon, and the second hole is a circle or an ellipse.

[0018] A microparticle capture system, comprising:

[0019] Microparticle capture chips as disclosed in any of the above;

[0020] A centrifugal turntable, wherein the upper surface of the centrifugal turntable is provided with a plurality of card holders distributed around the rotation center of the centrifugal turntable, the card holders being used to load the microparticle capture chip;

[0021] A drive unit is used to drive the centrifugal turntable to rotate.

[0022] A method for capturing microparticles, using the above-described microparticle capture system, includes the following steps:

[0023] After the sample is added, the first port and the second port are sealed with a film, and the microparticle capture chip is loaded into the holder of the centrifugal turntable. The side of the top cover layer that is away from the micro-pit layer faces the rotation center of the centrifugal turntable, and the flow channel is parallel to the rotation axis of the centrifugal turntable.

[0024] The centrifugal turntable rotates to sequentially complete the alignment, arrangement, and extrusion stages. In the alignment stage, the centrifugal turntable performs multiple forward and reverse rotation cycles at a first rotation speed. In the arrangement stage, the centrifugal turntable rotates forward at a second rotation speed greater than the first rotation speed. In the extrusion stage, the centrifugal turntable rotates forward at a third rotation speed greater than the second rotation speed.

[0025] Optionally, in the above microparticle capture method, the alignment step, the arrangement step, and the extrusion step take equal time. In the alignment step, each forward and reverse cycle takes 1 second, and the number of forward and reverse cycles is not less than 3.

[0026] As can be seen from the above technical solution, in the microparticle capture chip provided in this application, the flow channels of the micro-pit array corresponding to the micro-pit layer on the top cover layer are all set to extend in a straight line. Both ends of the flow channels are connected to the side of the top cover layer facing away from the micro-pit layer through the first port and the second port, respectively. Therefore, only a pipette is needed for sample addition and cleaning, eliminating the need for external valves or pipelines, making it convenient and user-friendly with low technical requirements for operators. Furthermore, since each flow channel and its corresponding micro-pit array form an independent sample addition cavity, it effectively solves the problem of simultaneously analyzing multiple samples, improving the flexibility and adaptability of sample loading throughput. In summary, the microparticle capture chip of this application can provide valuable assistance in constructing a platform that simultaneously possesses high throughput, high single-particle resolution, and low operator technical requirements. Attached Figure Description

[0027] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of this application. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0028] Figure 1 This is a schematic diagram of a microparticle capture chip according to an embodiment of this application;

[0029] Figure 2 This is a schematic diagram of a microparticle capture chip according to an embodiment of this application from another perspective;

[0030] Figure 3 This is a disassembly diagram of a microparticle capture chip according to an embodiment of this application;

[0031] Figure 4 This is a cross-sectional view of a microparticle capture chip according to an embodiment of this application;

[0032] Figure 5 This is an enlarged schematic diagram of the first micro-pit unit according to an embodiment of this application;

[0033] Figure 6 This is an enlarged schematic diagram of the second micro-pit unit according to an embodiment of this application;

[0034] Figure 7 This is a schematic diagram of a microparticle capture chip and a centrifugal turntable according to an embodiment of this application;

[0035] Figure 8 This is a schematic diagram of a microparticle capture system according to an embodiment of this application;

[0036] Figure 9 This is a top-down microscopic photograph of a portion of the cells after centrifugation and capture.

[0037] Figure 10 These are top-down microscopic images of the microspheres after centrifugation and capture.

[0038] Figure 11 This is a top-down microscopic photograph of the cells and microspheres after centrifugation and pairing.

[0039] The annotations in the attached figures are explained as follows:

[0040] 100. Bottom plate layer; 110. Settling tank; 120. Observation port; 130. Threaded hole;

[0041] 200. Support layer;

[0042] 300, Micro-pit layer; 310, Micro-pit array region; 311, First micro-pit unit; 312, Second micro-pit unit; 312a, First hole; 312b, Second hole;

[0043] 400, Top cover layer; 410, First opening; 420, Second opening; 430, Mounting hole; 440, Flow channel;

[0044] 500. Fasteners;

[0045] 600. Centrifugal turntable; 610. Card holder; 620. Weight reduction groove; 630. Notch;

[0046] 700. Drive unit;

[0047] 810. Base casing; 820. Protective casing; 821. Operating window; 830. Partition. Detailed Implementation

[0048] The technical solutions of the embodiments of this application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of this application, and not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this application.

[0049] In the description of this application, the references to terms such as "one embodiment," "some embodiments," "example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of this application. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of those different embodiments or examples.

[0050] In the description of this specification, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0051] See Figures 1-8This application provides a microparticle capture chip, which mainly includes a micro-pit layer 300 and a top cover layer 400. The first surface of the micro-pit layer 300 is provided with a micro-pit array at multiple locations. The top cover layer 400 covers the first surface of the micro-pit layer 300. The side of the top cover layer 400 facing the micro-pit layer 300 is provided with a plurality of flow channels 440 corresponding to the micro-pit array at each location. The flow channels 440 extend in a straight line and are parallel to each other. The two ends of the flow channels 440 are respectively provided with a first port 410 and a second port 420 that extend to the side of the top cover layer 400 facing away from the micro-pit layer 300. The flow channels 440 and the corresponding micro-pit array form an independent sample loading cavity.

[0052] The top cover layer 400 and the micro-pit layer 300 are stacked, with the top cover layer 400 contacting the first surface of the micro-pit layer 300. On the first surface of the micro-pit layer 300, a plurality of micro-pit arrays are arranged within the micro-pit array region 310, each micro-pit array consisting of multiple micro-pit units arranged in an array. Each micro-pit array corresponds to a flow channel 440 located on the side of the top cover layer 400 facing the micro-pit layer 300. These flow channels 440 of the top cover layer 400 are parallel straight lines. At one end of the flow channel 440, it connects to the side of the top cover layer 400 facing away from the micro-pit layer 300 through a first port 410; at the other end, it connects to the side of the top cover layer 400 facing away from the micro-pit layer 300 through a second port 420. One of the first port 410 and the second port 420 can serve as a sample loading port, and the other as a sample discharging port. The flow channel 440 and the corresponding micro-pit array form a sample loading cavity. The sample loading cavities where different flow channels 440 are located are independent of each other, that is, they are not connected to each other.

[0053] In traditional related technologies, rinsing multiple cells and changing the environmental fluid within the micropits requires extremely complex professional skills, making it impossible to guarantee batch-to-batch stability and the removal of free RNA from the environment, resulting in technical noise that is even difficult to assess. However, when using the microparticle capture chip of this application, since the flow channels 440 are all designed to extend in a straight line, only a pipette is needed for sample addition and washing, eliminating the need for external valves or pipelines, making it convenient and user-friendly with low operator skill requirements. Furthermore, because each flow channel 440 forms an independent sample addition cavity with the corresponding micropit array, it effectively solves the problem of analyzing multiple samples in parallel at once, improving the flexibility and adaptability of sample loading throughput. In summary, the microparticle capture chip of this application can provide valuable assistance in building a platform that simultaneously possesses high throughput, high single-cell resolution, and low operator skill requirements.

[0054] In some embodiments, the microparticle capture chip may include a support layer 200, which is stacked on the second surface of the micropit layer 300. The second surface faces opposite to the first surface, and the stiffness of the support layer 200 is greater than that of the micropit layer 300. The first and second surfaces are two surfaces of the micropit layer 300 opposite each other in the thickness direction. The first surface is the surface on which the micropit array is formed. The support layer 200 contacts the second surface, providing support for the micropit layer 300. The support layer 200 and the top cover layer 400 clamp the micropit layer 300, ensuring a sealed contact between the top cover layer 400 and the micropit layer 300. The support layer 200 also enhances the overall structural stiffness, reducing the possibility of local deformation of the micropit layer 300 leading to local separation from the top cover layer 400. This more effectively ensures that different sample loading cavities remain completely isolated from each other.

[0055] The materials of the micro-pit layer 300, the top cover layer 400, and the support layer 200 can be flexibly set as needed. For example, the material of the micro-pit layer 300 can be PDMS (polydimethylsiloxane), the material of the top cover layer 400 can be high-transparency materials such as PMMA (polymethyl methacrylate) or PC (polycarbonate), and the material of the support layer 200 can be high-transparency materials such as glass.

[0056] When the material of the micro-pit layer 300 is PDMS, the micro-pit layer 300 can be configured to have a sealed surface obtained by immersion in a sealing reagent (i.e., a surface after surface sealing treatment). PDMS is a porous material with a certain degree of viscosity. If the micro-pit layer 300 is not surface sealed, it is easy to cause non-specific adsorption of microparticles, affecting the capture effect. When performing surface sealing treatment, the sealing reagent can be the commonly used 1% BSA (1% bovine serum albumin solution by mass / volume). That is, the micro-pit layer 300 can be immersed in 1% BSA to achieve surface sealing treatment.

[0057] In some embodiments, the microparticle capture chip may include a substrate layer 100, which may have a recess 110 for accommodating a support layer 200 and a micro-pit layer 300. An observation port 120 is provided at the bottom of the recess 110. In such an embodiment, the support layer 200 and the micro-pit layer 300 are mounted within the recess 110 of the substrate layer 100, and the substrate layer 100 can form a protective frame to protect the support layer 200 and the micro-pit layer 300. The observation port 120 of the substrate layer 100 allows the support layer 200 to be observed, enabling the capture results of the micro-pit layer 300 to be observed under a microscope through the support layer 200. That is, the observation port 120 corresponds at least to the micro-pit array region 310 of the micro-pit layer 300, meaning the area of ​​the observation port 120 is not less than the area of ​​the micro-pit array region 310. The material of the substrate layer 100 can be flexibly chosen as needed; for example, the material of the substrate layer 100 may be a hard metallic material such as aluminum alloy.

[0058] In some embodiments, the top cover layer 400 may be positioned outside the recess 110 of the bottom plate layer 100 and in contact with the surface of the bottom plate layer 100. In such embodiments, the depth of the recess 110 may be set to be slightly smaller than the sum of the thicknesses of the support layer 200 and the micro-pit layer 300 when the top cover layer 400 is not covered (for example, a difference of 1mm to 2mm). This way, after the micro-pit layer 300 and the support layer 200 are placed in the recess 110 of the bottom plate layer 100, they will be slightly higher than the outer surface of the bottom plate layer 100. After the top cover layer 400 is placed on top, the thickness of the micro-pit layer 300 will slightly decrease under the pressure of the top cover layer 400 (i.e., the micro-pit layer 300 will be slightly compressed and deformed in the thickness direction), resulting in a more reliable sealing contact between the top cover layer 400 and the first surface of the micro-pit layer 300. Of course, in other embodiments, the top cover layer 400 may also be positioned within the recess 110, as long as it ensures that the top cover layer 400 presses firmly against the micro-pit layer 300.

[0059] In some embodiments, the top cover layer 400 may be provided with mounting holes 430 for mounting fasteners 500, and the bottom plate layer 100 may be provided with threaded holes 130 corresponding to the mounting holes 430. The top cover layer 400 and the bottom plate layer 100 can be connected by fasteners 500 (e.g., screws). This configuration allows the microparticle capture chip to achieve a simple structure that is easy to manufacture and assemble. After disassembling the fasteners 500, the various structural layers of the microparticle capture chip can be disassembled, facilitating the separate cleaning of the microparticle capture chip components. After cleaning, each structural layer can be reused and reassembled into a microparticle capture chip, which has good economic value. Of course, in other embodiments, the top cover layer 400 and the bottom plate layer 100 can be fixedly connected by other means, such as by adhesive bonding or welding. However, such embodiments are not convenient or allow for the disassembly and cleaning of the microparticle capture chip components.

[0060] The capture of microparticles includes, but is not limited to, suspensions of regular particles such as cells and microspheres. The size of the capture unit (i.e., the size of the micropit unit) of the micropit array of the micropit layer 300 can be designed according to the microparticles to be captured. For example, for cell capture, it can be designed according to the average size of the cell line; for microsphere capture, it can be designed according to the average size of the microspheres; for cell and microsphere pairing, the micropit unit of the micropit array can be designed as a two-layer structure.

[0061] Multiple micro-pit arrays are provided within the micro-pit array region 310 of the micro-pit layer 300. In some embodiments, at least one micro-pit array can be configured to consist of multiple first micro-pit units 311, where each first micro-pit unit 311 is a single-layer hole. Figure 5As shown, the first micro-pit unit 311 is a single-layer structure, that is, the first micro-pit unit 311 is a blind hole with a uniform cross-section. The shape of the first micro-pit unit 311 can be set in various forms, such as a regular hexagon, a circle, etc.

[0062] In some embodiments, at least one micropit array can be configured to consist of a plurality of second micropit units 312, wherein the second micropit unit 312 is configured as a double-layer hole, the double-layer hole including a first hole 312a connected to the first surface and a second hole 312b connected to the bottom of the first hole 312a, the area of ​​the second hole 312b being smaller than the area of ​​the first hole 312a. Figure 6 As shown, the second micro-pit unit 312 has a double-layer structure, that is, the second micro-pit unit 312 is a segmented blind hole with different cross-sectional sizes for different segments, among which the segment near the hole opening (i.e., the first hole 312a) has a relatively larger cross-section. In some embodiments, the first hole 312a can be set as a regular hexagon, and the second hole 312b can be set as a circle or an ellipse.

[0063] In some embodiments, the micropit arrays at different locations can be configured to consist of the same micropit units. For example, the micropit arrays at each location can consist of multiple first micropit units 311, or the micropit arrays at each location can consist of multiple second micropit units 312. Of course, in other embodiments, at least two micropit arrays can be configured to consist of different micropit units. For example, the micropit layer 300 can include both a micropit array composed of multiple first micropit units 311 and a micropit array composed of multiple second micropit units 312.

[0064] See Figures 1-8 This application also provides a microparticle capture system, which includes a centrifugal disc 600, a driving device 700, and the microparticle capture chip disclosed in any of the above embodiments. The driving device 700 drives the centrifugal disc 600 to rotate. A plurality of mounting slots 610 are disposed on the upper surface of the centrifugal disc 600 around its rotation center. The mounting slots 610 are used to hold the microparticle capture chip. The structure and working principle of the microparticle capture chip can be referred to the preceding description of the microparticle capture chip, and will not be repeated here. Since the microparticle capture chip disclosed in the above embodiments has the above-mentioned technical effects, the microparticle capture system having this microparticle capture chip also has the above-mentioned technical effects, and will not be repeated here.

[0065] See Figure 7 and Figure 8In some embodiments, the holder 610 on the centrifugal turntable 600 can be configured as a slot structure, allowing the microparticle capture chip to be vertically inserted into the holder 610 for easy and quick loading. The cross-section of the holder 610 is adapted to the size of the microparticle capture chip, ensuring that the chip is fixedly connected to the centrifugal turntable 600 after insertion. To facilitate removal, a notch 630 can be provided at the upper port of the holder 610, allowing the operator to grasp a larger area of ​​the microparticle capture chip with their fingers. The notch 630 can be provided only on the side of the upper port of the holder 610 closest to the rotation center of the centrifugal turntable 600, thus allowing the inner wall of the holder 610 on the other side to have a relatively large area to support the microparticle capture chip, providing more reliable support for the chip during the rotation of the centrifugal turntable 600. In other embodiments, the holder 610 on the centrifugal turntable 600 can be configured in other forms, such as a hole structure with the opening facing the rotation center of the centrifugal turntable 600. In such an embodiment, the microparticle capture chip moves radially along the centrifugal turntable 600 in an upright state and is inserted into the holder 610.

[0066] In some embodiments, the centrifugal turntable 600 may be provided with a plurality of weight-reducing grooves 620 distributed around the rotation center of the centrifugal turntable 600, and the weight-reducing grooves 620 and the holder 610 are arranged alternately along the circumference of the centrifugal turntable 600. By providing the weight-reducing grooves 620, the weight of the centrifugal turntable 600 is reduced, which can reduce the wind resistance of the centrifugal turntable 600 during the centrifugation process and ensure stability. It is easy to understand that in order to improve the rotational stability, the weight-reducing grooves 620 and the holder 610 should be evenly distributed around the rotation center of the centrifugal turntable 600.

[0067] See Figure 7 In embodiments where the card holder 610 is configured as a slot structure, the depth and size of the card holder 610 should ensure that the entire or most of the volume of the microparticle capture chip can be loaded inside, so as to prevent it from flying out during centrifugation. In some embodiments, the circumferential diameter of the centrifugal turntable 600 can be set to 16 cm, under which eight card holders 610 can be arranged. After the microparticle capture chip is loaded into the card holder 610, its plane (i.e., the surface of the top cover layer 400 facing away from the micro-pit layer 300) is perpendicular to the rotation plane of the centrifugal turntable 600. Of course, in other embodiments, the circumferential diameter of the centrifugal turntable 600 can be set to other values, such as 18 cm, 20 cm, etc.

[0068] For single-cell analysis, microwell array devices offer several key advantages over droplet-based devices, including lower sample and reagent consumption, no peripheral equipment, ease of parallelization, enhanced compatibility and openness, and suitability for short-term cell culture and cell-cell interaction experiments. However, in traditional related techniques, most microwells employ gravity sedimentation for cell capture, and the long sedimentation time poses a significant challenge to preserving the viability of cells and tissue samples. Moreover, to minimize crosstalk risks, these methods make significant compromises in well occupancy, resulting in a substantial increase in reagent consumption and cost during library construction.

[0069] The microparticle capture system of this application includes a centrifuge disc 600, which utilizes centrifugal force to actively transport individual microparticles into micropit units, achieving rapid and efficient loading and capture of single microparticles. Therefore, it provides a fast, efficient, and economical platform for capturing single microparticles in a scalable throughput manner, and allows for easy washing away of multiple-particle captures and environmental transcript contamination. Using the microparticle capture system of this application, the capture time is simple and rapid, with a single centrifugation capture time not exceeding 2 minutes, making it more practical for fields such as live cell analysis.

[0070] The drive unit 700 can take many forms; see [link / reference]. Figure 8 In some embodiments, the drive device 700 can be configured as a servo motor directly connected to the centrifugal turntable 600, that is, the drive device 700 and the centrifugal turntable 600 can be in a direct drive configuration. Of course, in other embodiments, the drive device 700 can also be configured to be connected to the centrifugal turntable 600 through a transmission mechanism, as long as the drive device 700 can drive the centrifugal turntable 600 to rotate.

[0071] In some embodiments, the microparticle capture system may include a base housing 810, and the drive device 700 may be disposed within the base housing 810. Furthermore, the microparticle capture system may include a protective housing 820, which covers the centrifuge disc 600 and is fixedly connected to the base housing 810, ensuring safety during the centrifugation process. The top of the protective housing 820 may have an operation window 821, allowing an operator's hand or an industrial robot to reach the centrifuge disc 600 to perform experimental operations (e.g., installing or removing a microparticle capture chip from the centrifuge disc 600, etc.).

[0072] In some embodiments, the microparticle capture system may include a partition 830, which is connected to the top of the base housing 810, separating the drive unit 700 and the centrifugal disc 600 into different spaces, and is assembled and connected to the drive unit 700 to provide stable support for the drive unit 700.

[0073] This application also provides a microparticle capture method using the microparticle capture system disclosed in the above embodiments, comprising the following steps:

[0074] Step A: After the sample is added, seal the first port 410 and the second port 420 with a film, load the microparticle capture chip into the card holder 610 of the centrifuge turntable 600, with the side of the top cover layer 400 facing away from the micro-pit layer 300 facing the rotation center of the centrifuge turntable 600, and the flow channel 440 parallel to the rotation axis of the centrifuge turntable 600.

[0075] The sample loading operation involves adding a microparticle suspension into the sample loading cavity. During loading, one of the first port 410 and the second port 420 serves as the sample loading port, and the other as the sample outlet. After loading, the first port 410 and the second port 420 are sealed with a film to close the sample loading cavity. The film can be adhesive tape, which needs to be removed when cleaning the sample loading cavity. The microparticle capture chip is mounted vertically on the centrifuge disc 600, with the extension direction of the flow channel 440 parallel to the rotation axis of the centrifuge disc 600.

[0076] In step B, the centrifugal turntable 600 rotates, sequentially completing the alignment, arrangement, and extrusion stages. In the alignment stage, the centrifugal turntable 600 performs multiple forward and reverse rotation cycles at a first rotation speed; in the arrangement stage, the centrifugal turntable 600 rotates forward at a second rotation speed greater than the first rotation speed; and in the extrusion stage, the centrifugal turntable 600 rotates forward at a third rotation speed greater than the second rotation speed.

[0077] The alignment stage aligns the microparticle suspension in the flow channel 440 to the plane where the micro-pit layer 300 is located. The arrangement stage aligns the microparticles above each micro-pit unit to reduce the amount of microparticles remaining between micro-pit units. The extrusion stage extrudes the microparticles aligned above the micro-pit unit to the bottom of the micro-pit unit to achieve capture.

[0078] One forward and reverse cycle of the centrifugal turntable 600 refers to one forward rotation and one reverse rotation. It should be noted that the direction of forward rotation can be either counterclockwise or clockwise. The forward and reverse rotations of the alignment stage, as well as the above three stages, are carried out continuously and without interruption. Therefore, no pause time is set between forward and reverse rotations.

[0079] In some embodiments, the microparticle capture method can be configured such that the alignment, arrangement, and compression stages take equal time. In the alignment stage, each forward and reverse cycle takes 1 second, and the number of forward and reverse cycles is not less than 3. For example, the alignment stage can be set to have 5 forward and reverse cycles, then the alignment, arrangement, and compression stages each take 5 seconds.

[0080] The first, second, and third rotational speeds can be flexibly set as needed, with the speeds increasing sequentially. This means the centrifugation process begins with low-speed centrifugation followed by high-speed centrifugation. In some embodiments, the first rotational speed can be set to 500 rpm, the second to 600 rpm, and the third to 4000 rpm.

[0081] After step B, the microparticle capture chip can be removed from the centrifuge disc 600 for cleaning. Cleaning can be performed by pipetting and aspirating the sample loading chamber 10 times to flush out any excess microparticles in the flow channel 440 that have not entered the micropit unit. To illustrate the pipetting and aspiration process, an example is given below: Using a 200 μL pipette, aspirate 200 μL of cleaning solution (usually consistent with the suspension containing the microparticles) and quickly inject it into the flow channel 440 through the inlet; this process is called blowing. Keeping the pipette in place, quickly aspirate the liquid from the flow channel 440 into the pipette tip; this process is called aspiration.

[0082] The following describes three exemplary experiments conducted using the microparticle capture method of this application.

[0083] Experiment 1: Cell centrifugation and capture

[0084] According to the attached Figure 3 The capture chips are assembled sequentially according to the layer order shown in the attached diagram. Figure 5 The image shows a single-layer micro-pit array. Cell suspension (HeLa-GFP cells with green fluorescence were used for easy visualization) was added to the sample loading cavity of the capture chip. After sealing the loading and unloading ports, the chip was placed on an attached... Figure 8 The centrifugation capture process, as shown in Table 1, was performed in the centrifuge discs. The samples were then observed using a fluorescence microscope. Figure 9 The image shows a top-down microscopic photograph of the cells after centrifugation and capture, revealing that the cells occupy more than 95% of the microchip.

[0085] Table 1 Centrifugal Capture Process Parameters

[0086]

[0087] Experiment 2: Centrifugal capture of microspheres

[0088] According to the attached Figure 3 The capture chips are assembled sequentially according to the layer order shown in the attached diagram. Figure 5 The image shows a single-layer micro-pit array. A microsphere suspension (green fluorescent polystyrene microspheres were used for easy visualization) was added to the sample loading cavity of the capture chip. After sealing the loading and unloading ports, the chip was placed on an attached... Figure 8The centrifugation capture process, as shown in Table 1, was performed in the centrifuge discs. The samples were then observed using a fluorescence microscope. Figure 10 The image shows a top-down microscopic photograph of the microspheres after centrifugation and capture, revealing that the microspheres occupy more than 98% of the micro-pit chip.

[0089] Experiment 3: Centrifugal pairing of cells and microspheres

[0090] According to the attached Figure 3 The capture chips are assembled sequentially according to the layer order shown in the attached diagram. Figure 6 The diagram shows the micro-pit layers of the double-layer micro-pit array. First, cell suspension was added to the sample loading chamber of the capture chip. After centrifugation, the cells entered the lower pit of the double-layer micro-pit array unit. Then, microsphere suspension was added to the capture chip, and after centrifugation, the microspheres entered the upper pit of the double-layer micro-pit array unit. The cells were then observed using a fluorescence microscope. Figure 11 The image shows a top-down microscopic photograph of the cells and microspheres after centrifugation and pairing, revealing a pairing rate of over 95% between the cells and microspheres on the micro-pit chip.

[0091] The three experiments described above demonstrate that the microparticle capture system of this application can achieve efficient capture and pairing of microparticles, and is simple, fast and user-friendly.

[0092] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on the differences from other embodiments. The same or similar parts between the various embodiments can be referred to each other.

[0093] The above description of the disclosed embodiments enables those skilled in the art to make or use this application. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of this application. Therefore, this application is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A micro-particle capture chip, characterized by, include: A micro-pit layer (300) has a micro-pit array disposed at multiple locations on its first surface; A top cover layer (400) covers the first surface. The top cover layer (400) facing the micro-pit layer (300) has a plurality of flow channels (440) corresponding to the micro-pit arrays at each location. The flow channels (440) extend in a straight line and are parallel to each other. The two ends of the flow channels (440) are respectively provided with a first opening (410) and a second opening (420) that penetrate to the side of the top cover layer (400) facing away from the micro-pit layer (300). The flow channels (440) and the corresponding micro-pit arrays form independent sample loading cavities.

2. The microparticle capture chip of claim 1, wherein, include: A support layer (200) is stacked on the second surface of the micro-pit layer (300), the second surface being opposite to the first surface, and the stiffness of the support layer (200) being greater than the stiffness of the micro-pit layer (300).

3. The microparticle capture chip of claim 2, wherein, include: The bottom plate layer (100) is provided with a sink (110) for accommodating the support layer (200) and the micro-pit layer (300), and the bottom of the sink (110) is provided with an observation port (120).

4. The micro-particle capture chip according to any one of claims 1 to 3, wherein, At least one of the micro-pit arrays is composed of a plurality of first micro-pit units (311), wherein the first micro-pit unit (311) is configured as a single-layer hole.

5. The microparticle capture chip of claim 4, wherein, At least one of the micro-pit arrays is composed of a plurality of second micro-pit units (312), the second micro-pit unit (312) is configured as a double-layer hole, the double-layer hole includes a first hole (312a) connected to the first surface and a second hole (312b) connected to the bottom of the first hole (312a), the area of ​​the second hole (312b) is smaller than the area of ​​the first hole (312a).

6. The micro-particle capture chip according to any one of claims 1 to 3, wherein, At least one of the micro-pit arrays is composed of a plurality of second micro-pit units (312), the second micro-pit unit (312) is configured as a double-layer hole, the double-layer hole includes a first hole (312a) connected to the first surface and a second hole (312b) connected to the bottom of the first hole (312a), the area of ​​the second hole (312b) is smaller than the area of ​​the first hole (312a).

7. The microparticle capture chip according to claim 6, characterized in that, The first hole (312a) is a regular hexagon, and the second hole (312b) is a circle or an ellipse.

8. A microparticle capture system, characterized in that, include: The microparticle capture chip as described in any one of claims 1 to 7; A centrifugal turntable (600) has a plurality of card holders (610) arranged around the rotation center of the centrifugal turntable (600) on its upper surface. The card holders (610) are used to load the microparticle capture chip. A drive unit (700) is used to drive the centrifugal turntable (600) to rotate.

9. A method for capturing microparticles, characterized in that, Using the microparticle capture system as described in claim 8 includes the following steps: After the sample is added, the first port (410) and the second port (420) are sealed with a film. The microparticle capture chip is loaded into the holder (610) of the centrifugal turntable (600). The side of the top cover layer (400) facing away from the micro-pit layer (300) faces the rotation center of the centrifugal turntable (600). The flow channel (440) is parallel to the rotation axis of the centrifugal turntable (600). The centrifugal turntable (600) rotates to sequentially complete the alignment, arrangement, and extrusion stages. In the alignment stage, the centrifugal turntable (600) performs multiple forward and reverse rotation cycles at a first rotation speed. In the arrangement stage, the centrifugal turntable (600) rotates forward at a second rotation speed greater than the first rotation speed. In the extrusion stage, the centrifugal turntable (600) rotates forward at a third rotation speed greater than the second rotation speed.

10. The microparticle capture method according to claim 9, characterized in that, The alignment, arrangement, and compression steps take equal time. In the alignment step, each forward and reverse cycle takes 1 second, and the number of forward and reverse cycles is not less than 3.

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