Extracellular vesicle extraction device

Abstract

The invention discloses an extracellular vesicle extraction device, which is characterized by comprising: a housing, the housing comprises an upper housing and a lower housing, and the upper housing is communicated with the lower housing; the filtering assembly is arranged on the upper shell and is communicated with the upper shell; the centrifugal assembly is arranged in the lower shell, and the upper part of the centrifugal assembly is communicated with the lower part of the filter assembly through a filter screen. The filtering assembly is used for filtering blood, alveolar lavage fluid and cell culture fluid, the blood, the alveolar lavage fluid and the cell culture fluid directly enter the centrifugal assembly after being filtered, extracellular vesicles are separated out through ultracentrifugation, and finally precipitation and collection are carried out in combination with chromatography, so that the extraction efficiency of the extracellular vesicle suspension is greatly improved; and the whole process consumes short time.

Description

Technical Field

[0001] The present invention belongs to the technical field related to extracellular vesicle extraction, and particularly relates to an extracellular vesicle extraction device. Background Art

[0002] Extracellular vesicles have important value in many aspects, mainly in the fields of medical diagnosis and disease treatment. Medical diagnosis includes: Disease markers: Biomolecules such as proteins and nucleic acids carried by extracellular vesicles can be used as markers for disease diagnosis. For example, in the blood of cancer patients, exosomes released by tumor cells may contain specific tumor-associated antigens or gene mutation information. By detecting these markers, it can help with early diagnosis, disease monitoring and prognosis assessment of tumors. Liquid biopsy: Compared with traditional tissue biopsy, liquid biopsy based on extracellular vesicles has the advantages of being non-invasive and highly reproducible. By analyzing extracellular vesicles in body fluids such as blood and urine, disease-related information can be obtained, providing a basis for diagnosis and treatment of the disease.

[0003] Disease treatment includes: Drug carriers: Extracellular vesicles can serve as natural drug carriers to deliver therapeutic drugs, nucleic acids, etc. to target cells. They have good biocompatibility and low immunogenicity, can protect the drugs they carry from the influence of the internal environment, improve the stability and targeting of the drugs, and reduce the side effects of drugs on normal tissues. Cell therapy: Extracellular vesicles derived from mesenchymal stem cells and other sources have functions such as immune regulation and tissue repair. In the treatment of some diseases, such as cardiovascular diseases and neurodegenerative diseases, extracellular vesicles can play a therapeutic role by regulating cell signaling pathways, promoting cell proliferation and differentiation, etc.

[0004] Extracellular vesicles (EVs) are nanoscale membrane-bound vesicles formed by active cell secretion or shedding. Their density and size differ from impurities such as cells, cell debris, and protein aggregates. Ultracentrifugation utilizes the differences in sedimentation rates of different particles under a strong centrifugal field to separate EVs from complex biological samples by gradually increasing the centrifugal force and extending the centrifugation time.

[0005] Patent publication number CN118516219A discloses an extracellular vesicle extraction device, comprising a 30-nanometer pore membrane, a central-pore organic glass gasket, an eccentric-pore organic glass gasket, and a 200-nanometer pore membrane 15, sequentially installed within an outlet housing. Before extraction, a centrifuge is used to remove impurities such as cell debris, yielding a plasma supernatant. This plasma supernatant is then placed within the housing and filtered to yield an extracellular vesicle suspension. However, because the centrifuge and extracellular vesicle extraction device are designed separately, the overall process is time-consuming and inefficient. Summary of the Invention

[0006] The present invention aims to provide an extracellular vesicle extraction device to solve the technical problems of time-consuming overall process and low efficiency of existing extracellular vesicle extraction devices.

[0007] The extracellular vesicle extraction device of the present invention comprises: A filter assembly is provided above the centrifugal housing to filter the plasma supernatant; The centrifugal assembly is disposed within the centrifugal housing and below the filter assembly to centrifuge the plasma supernatant after filtration by the filter assembly. The filter assembly and the centrifugal assembly are combined to extract extracellular vesicles, thereby improving work efficiency.

[0008] Preferably, The filter assembly comprises: The filter housing is disposed above the centrifugal housing, and a first filter hole is provided at the bottom of the filter housing. The plasma supernatant is filtered through the filter hole and then flows into the centrifugal assembly, thereby improving the filtration effect of the plasma supernatant.

[0009] The filter group also includes: The negative pressure pump is arranged on the filter housing and sucks the plasma supernatant through the suction tube and transports it to the filter housing for filtration, thereby realizing automatic extraction of the plasma supernatant.

[0010] The centrifugal assembly comprises: A rotating component, the bottom of which is connected to the bottom of the centrifugal housing; The first centrifugal cavity is arranged on the top of the rotating component. The top of the first centrifugal cavity is provided with a second filter hole. The plasma supernatant in the filter shell flows into the first centrifugal cavity through the first filter hole and the second filter hole.

[0011] The centrifugal assembly further comprises: a second centrifugal cavity, disposed on the periphery of the first centrifugal cavity and connected to the first centrifugal cavity through a pipeline, wherein a through hole is provided at the lower portion of the second centrifugal cavity; a filter membrane disposed below the second centrifugal chamber, wherein the rotating component passes through the filter membrane and is fixedly connected thereto, and a distance is left between the filter membrane and the bottom of the housing; A chromatographic column is arranged on the filter membrane, and the chromatographic column is aligned with the through hole at the lower part of the second centrifugal cavity.

[0012] The filter membrane is provided with waste liquid filtering holes in an array, and the plasma supernatant on the filter membrane flows into the gap between the filter membrane and the bottom of the shell through the waste liquid filtering holes.

[0013] Preferably, There are multiple chromatographic columns, which are circumferentially arranged on the filter membrane; There are multiple second centrifugal cavities, which are circumferentially arranged on the outer wall of the first centrifugal cavity. The multiple second centrifugal cavities correspond one-to-one to the multiple chromatographic columns.

[0014] The centrifugal assembly further comprises: a first transmission part, disposed on the rotating component and located below the first centrifugal cavity; The second transmission part is arranged below the second centrifugal cavity and is rotationally connected. The first transmission part is rotationally connected to the second transmission part.

[0015] The centrifugal assembly further comprises: The filter screen is arranged on the top of the first centrifugal cavity and is fixedly connected. The filter screen is located between the first filter hole and the second filter hole. The plasma supernatant filtered by the first filter hole flows into the filter screen for secondary filtration, and the plasma supernatant filtered by the filter screen flows into the second filter hole for tertiary filtration.

[0016] In the above, the filter, the first centrifugal cavity, and the second centrifugal cavity are designed in proportion, that is, the filter area is smaller than the first centrifugal cavity, and the diameter of the first centrifugal cavity is smaller than the second centrifugal cavity. In this way, the filter, the first centrifugal cavity, and the second centrifugal cavity divide the interior of the centrifugal shell into three centrifugal zones, so that the apoptotic bodies with the largest diameter, the microvesicles, and the exosomes with the smallest diameter are centrifuged out in the three centrifugal zones respectively.

[0017] Preferably, The upper centrifugal housing is provided with a through hole, and the filter housing is provided on the centrifugal housing, wherein the first filter hole is communicated with the through hole; The lower centrifugal shell is arranged below the upper centrifugal shell and is rotatably connected to the upper centrifugal shell. The centrifugal assembly is arranged in the upper centrifugal shell and the lower centrifugal shell.

[0018] The positive progress effect of the present invention is: 1) The present invention designs a filtration component and a centrifugal component. The filtration component filters blood, alveolar lavage fluid, and cell culture fluid. After filtration, the filtered fluid directly enters the centrifugal component for centrifugation. Extracellular vesicles are separated by ultracentrifugation, and finally precipitated and collected by chromatography. This greatly improves the extraction efficiency of the extracellular vesicle suspension and shortens the overall process time.

[0019] 2) The filter screen, the first centrifugal chamber, and the second centrifugal chamber of the present invention are designed in proportion, that is, the filter screen area is smaller than the first centrifugal chamber, and the diameter of the first centrifugal chamber is smaller than the second centrifugal chamber. In this way, the filter screen, the first centrifugal chamber, and the second centrifugal chamber divide the interior of the centrifugal shell into three centrifugal zones, so that the apoptotic bodies with the largest diameter, the microvesicles, and the exosomes with the smallest diameter are centrifuged respectively in the three centrifugal zones.

[0020] 3) The present invention is designed to have a rotating component, an upper centrifugal housing, and a lower centrifugal housing. The lower centrifugal housing is rotatably connected to the upper centrifugal housing. The rotating component drives the filter screen, the first centrifugal chamber, the second centrifugal chamber, and the upper housing to rotate. Among them, the upper housing rotates in the opposite direction to the filter screen, the first centrifugal chamber, and the second centrifugal chamber. In this way, the extracellular vesicles rotate several times along the inner walls of the filter screen, the first centrifugal chamber, and the second centrifugal chamber to buffer the collision caused by excessive centrifugal force and reduce damage to the extracellular vesicles. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 It is a schematic diagram of the overall structure of the present invention; Figure 2 This is a schematic diagram of the filter housing structure of the present invention; Figure 3 for Figure 2 Cross-sectional view; Figure 4 This is a schematic diagram of the connection between the upper centrifugal housing and the lower centrifugal housing of the present invention; Figure 5 This is a schematic diagram of the connection between the rotating component and the lower centrifugal housing of the present invention; Figure 6 This is a schematic diagram of the filter housing structure of the present invention; Figure 7 Schematic diagram of the centrifugal assembly structure of the present invention Figure 1 ; Figure 8 Schematic diagram of the centrifugal assembly structure of the present invention Figure 2 ; Figure 9 Schematic diagram of the centrifugal assembly structure of the present invention Figure 3 ; Figure 10 Schematic diagram of the centrifugal assembly structure of the present invention Figure 4 ; Figure 11 Schematic diagram of the centrifugal assembly structure of the present invention Figure 5 ; Figure 12 This is a schematic diagram of the connection between the filter housing, the upper centrifugal housing, and the lower centrifugal housing of the present invention; Figure 13 This is a schematic diagram of the connection between the upper centrifugal housing and the lower centrifugal housing of the present invention; Figure 14 This is a schematic diagram of the connection between the upper centrifugal housing, the lower centrifugal housing, and the centrifugal assembly of the present invention; Figure 15 Schematic diagram of the connection between the upper centrifugal housing and the centrifugal assembly of the present invention Figure 1 ; Figure 16 Schematic diagram of the connection between the upper centrifugal housing and the centrifugal assembly of the present invention Figure 2 ; Figure 17 This is a schematic diagram of the connection between the second centrifugal cavity, the second transmission part, and the first centrifugal cavity of the present invention.

[0022] In the figure: 1 upper centrifugal housing; 101 inner annular gear; 2 lower centrifugal housing; 201 rotating component; 202 chromatographic column; 203 filter membrane; 204 second transmission component; 205 first centrifugal cavity; 206 second centrifugal cavity; 207 pipeline; 208 first transmission component; 209 filter screen; 210 fixing rod; 3 filter housing; 301 suction tube; 302 negative pressure pump; 303 first filter hole. DETAILED DESCRIPTION

[0023] The following describes the embodiments of the present invention through specific examples. Those skilled in the art will readily understand the other advantages and benefits of the present invention from the disclosure herein. The present invention may also be implemented or applied through specific embodiments, and the details in this specification may be modified or altered based on different viewpoints and applications without departing from the spirit of the present invention.

[0024] There are many methods for extracting extracellular vesicles, including ultracentrifugation, Principle: Extracellular vesicles (EVs) are separated from other cellular components by varying the centrifugal force based on their density and size. By gradually increasing the centrifugation speed, cell debris and larger organelles are removed before the EVs are collected.

[0025] Operation steps: Usually, biological samples (such as blood, alveolar lavage fluid, cell culture fluid, etc.) are first centrifuged at a low speed to remove cells and large impurities; then the supernatant is transferred to a new centrifuge tube and centrifuged multiple times at a higher speed to finally obtain extracellular vesicle precipitation.

[0026] Advantages: Currently the most commonly used method, it can obtain extracellular vesicles of high purity and is applicable to a variety of biological samples. Disadvantages: The operation is complex, requires expensive ultracentrifuges, is time-consuming, and may cause aggregation and damage of extracellular vesicles.

[0027] Density gradient centrifugation, principle: a continuous or discontinuous density gradient medium (such as sucrose, iodixanol, etc.) is formed in a centrifuge tube. Under the action of centrifugal force, extracellular vesicles are distributed in different density layers according to their density, thereby achieving separation.

[0028] Procedure: After mixing the sample with a density gradient medium, centrifuge. Extracellular vesicles (EVs) are distributed in a layer of medium with the same density as the EVs and can be collected by puncture or siphoning.

[0029] Advantages: It can achieve more precise separation based on the density differences of extracellular vesicles, thereby improving purity. Disadvantages: The operation is relatively cumbersome, requiring special density gradient media and centrifugation equipment, and the separation process is relatively long.

[0030] Ultrafiltration: Principle: Using an ultrafiltration membrane with a specific pore size, components in a solution are separated based on molecular size. Extracellular vesicles (EVs) typically range in size from tens to hundreds of nanometers. Choosing an ultrafiltration membrane with an appropriately sized pore size can retain EVs while allowing small molecules and solvents to pass through.

[0031] Operation steps: The biological sample is passed through the ultrafiltration device. Under the action of pressure or centrifugal force, the liquid passes through the ultrafiltration membrane, and the extracellular vesicles are retained on the membrane. Finally, the extracellular vesicles are eluted with an appropriate buffer.

[0032] Advantages: The procedure is relatively simple and rapid, can be performed at room temperature, and can better preserve the biological activity of extracellular vesicles. Disadvantages: Extracellular vesicles may be lost due to membrane adsorption, and the separation effect of extracellular vesicles of different sizes is limited.

[0033] Immunoaffinity capture method, Principle: Utilize the specific binding between specific antigens on the surface of extracellular vesicles and corresponding antibodies to capture extracellular vesicles onto solid phase carriers (such as magnetic beads, microplates, etc.), thereby achieving separation.

[0034] Operation steps: Mix a solid phase carrier coated with specific antibodies with the biological sample to allow the antibodies to bind to the antigens on the surface of extracellular vesicles; then separate the solid phase carrier bound to the extracellular vesicles by magnetic force or centrifugation, and after washing to remove impurities, the extracellular vesicles can be obtained.

[0035] Advantages: Highly specific, capable of enriching specific types of extracellular vesicles and improving purity. Disadvantages: Requires selection of appropriate antibodies for different extracellular vesicles, which is costly and may affect the integrity and function of extracellular vesicles.

[0036] Polymer precipitation: Principle: Using a precipitant such as a polymer (e.g., polyethylene glycol, PEG) to precipitate extracellular vesicles by altering the physicochemical properties of the solution. PEG binds to water molecules, reducing their activity and promoting the aggregation and precipitation of extracellular vesicles.

[0037] Operation steps: Add an appropriate amount of polymer precipitant to the biological sample, mix evenly, incubate at a certain temperature for a period of time, and then collect the precipitate by centrifugation or other methods to obtain extracellular vesicles.

[0038] Advantages: Simple and rapid operation, no need for special equipment, and suitable for large-scale sample processing. Disadvantages: The product obtained by precipitation may contain a large number of impurities and require further purification, and the polymer may affect the structure and function of extracellular vesicles.

[0039] like Figure 1 As shown, the present invention mainly provides an extracellular vesicle extraction device for ultracentrifugation, which has a long process and low efficiency. The device comprises an upper centrifugal shell 1, a lower centrifugal shell 2, and a filter shell 3; the upper centrifugal shell 1 is a circular shell with a flange, a through hole is provided on the circular surface, and is made of stainless steel, the lower centrifugal shell 2 is a circular shell with a flange, a stainless steel material, and the filter shell 3 is a circular chamber, and an array of first filter holes 303 are provided on the circular surface of the circular chamber; the upper centrifugal shell 1 and the lower centrifugal shell 2 are connected through an internal centrifugal assembly, and the connected upper centrifugal shell 1 is on the lower centrifugal shell and does not contact each other; the first filter hole 303 on the filter shell 3 is aligned with the through hole on the upper centrifugal shell 1, and the plasma supernatant in the filter shell 3 is filtered through the first filter hole 303 and flows into the centrifugal assembly, and the centrifugal assembly is separated under different centrifugal forces, and extracellular vesicles are obtained after separation, which greatly improves the extraction efficiency of the extracellular vesicle suspension.

[0040] Plasma supernatant extraction, blood collection and anticoagulation: Blood collection is generally done at room temperature but needs to be processed as quickly as possible. Anticoagulants such as heparin and EDTA are usually added to prevent blood clotting.

[0041] Centrifugation: The collected anticoagulated blood is centrifuged to separate the plasma and blood cells. The centrifugation temperature is generally controlled between 4°C and 25°C. In experiments requiring high stability of bioactive substances in plasma, centrifugation at 4°C is used to reduce enzyme activity and minimize degradation of bioactive substances. In some routine tests, centrifugation at room temperature (approximately 20°C to 25°C) can also be performed to improve efficiency. After centrifugation, the pale yellow, transparent liquid on the top is the plasma supernatant. Carefully aspirate the supernatant and transfer it to a clean container for storage.

[0042] Storage Temperature: If the extracted plasma supernatant cannot be immediately tested or used for subsequent experiments, an appropriate storage temperature should be selected based on the specific test being performed. Generally, short-term storage (hours to days) can be refrigerated at 4°C. Long-term storage (months to years) requires freezing at -20°C or -80°C to maintain the stability of the various components in the plasma.

[0043] Cell culture fluid extraction, cell culture: Cell culture is typically performed in an incubator at 37°C and 5% CO2 to simulate the in vivo environment and provide optimal cell growth conditions. Different cell types may require slightly different culture conditions, but 37°C is a common temperature for most mammalian cell cultures.

[0044] Collecting cell culture fluid: To extract cell culture fluid, first remove the culture flask or dish from the incubator and perform the procedure under sterile conditions. For adherent cells, remove the culture fluid and gently wash the cells with a buffer such as PBS. Then, add an appropriate amount of a digestive solution such as trypsin to detach the cells from the culture vessel surface. Add a serum-containing culture medium to terminate digestion. Centrifuge the cell suspension and collect the supernatant to obtain the cell culture fluid. For suspended cells, centrifuge the cell suspension directly and collect the supernatant. During these procedures, maintain the temperature close to the cell culture temperature (around 37°C) to minimize any effects on the cells and components of the culture fluid. However, in practice, due to the short duration of the procedure, performing the procedure at room temperature (20°C-25°C) generally does not significantly affect the results.

[0045] Storage Temperature: If the extracted cell culture medium needs to be stored, the appropriate temperature should be selected based on the experimental purpose and requirements. For short-term storage for subsequent cell experiments, such as cytokine detection, it can be stored at 4°C. For long-term storage or for specialized experiments, such as the preservation of extracellular vesicles, it may be necessary to freeze at -80°C or lower.

[0046] The volume of extracellular vesicle suspension extracted from blood, alveolar lavage fluid, and cell culture medium is usually 10-100 ml, for example, 50 ml. Centrifugation speed and time: The first step is low-speed centrifugation: generally at 300-500×g, centrifugation for 10-15 minutes, such as 400×g centrifugation for 10 minutes, the purpose is to remove cell debris and large cell aggregates. The second step is medium-speed centrifugation: the speed can be set at 10,000-20,000×g, centrifugation for 20-30 minutes, such as 15,000×g centrifugation for 25 minutes, to further remove smaller impurities. The third step is high-speed centrifugation: the speed can reach 100,000-200,000×g, the centrifugation time is 60-120 minutes, and 110,000×g centrifugation for 90 minutes is often used to precipitate extracellular vesicles. The final volume of extracellular vesicle suspension obtained is generally around 0.5-5 mL, depending on the starting culture medium volume and extraction efficiency. For example, approximately 1 mL of extracellular vesicle suspension can be obtained from 50 mL of blood, bronchoalveolar lavage fluid, or cell culture medium. These values ​​are examples only and may require optimization and adjustment based on cell type, culture conditions, and specific experimental requirements.

[0047] like Figure 2 、 Figure 3As shown, in order to improve the extraction efficiency of the extracellular vesicle suspension, a negative pressure pump 302 is provided on the filter housing 3, and the negative pressure pump 302 model is DQB410-SB). The outlet of the negative pressure pump 302 is connected to the interior of the filter housing 3, and a suction tube 301 is provided on the water inlet of the negative pressure pump 302. In this way, the negative pressure pump 302 directly draws the plasma supernatant into the filter housing 3 for filtration; the existing technology is manual filtration followed by centrifugation, which saves some steps.

[0048] like Figure 7 、 8 , 9, 10, 11, and 12, the centrifugal assembly includes a rotating component 201, which is arranged on the bottom inner portion of the lower centrifugal shell 2 and fixedly connected. The rotating component 201 includes: a motor model of YET-YQDJ586 and a transmission shaft model of DZ9114315156. The motor is fixed to the bottom inner portion of the lower centrifugal shell 2 by bolts; the transmission shaft is connected to the motor output shaft by a coupling, and a first centrifugal cavity 205 is fixedly provided on the top of the transmission shaft. The first centrifugal cavity 205 is a circular stainless steel chamber, and a second filter hole is provided on the upper end surface of the first centrifugal cavity 205, so that the plasma supernatant inside the filter shell 3 is filtered through the first filter hole 303 and then flows into the first centrifugal cavity 205 for centrifugation.

[0049] A second centrifugal chamber 206 is circumferentially provided on the periphery of the first centrifugal chamber 205 (a circular ring with one end closed, a pipe 207 provided on the closed end, the pipe 207 communicating with the interiors of the first centrifugal chamber 205 and the second centrifugal chamber 206, and the pipe 207 is welded to the second centrifugal chamber 206 and the first centrifugal chamber 205). During the centrifugation of the first centrifugal chamber 205, the second centrifugal chamber 206 is driven to centrifuge. The diameter of the first centrifugal chamber 205 is smaller than that of the second centrifugal chamber 206. Thus, the first centrifugal chamber 205 and the second centrifugal chamber 206 divide the interior of the upper centrifugal housing 1 into two centrifugal zones, so that microvesicles and exosomes with the smallest diameter are respectively centrifuged out of the two centrifugal zones.

[0050] The overall planning dimensions of the product are as follows: the overall height of the equipment is controlled at 600-800mm, the width is 400-600mm, and the length is 500-700mm, which is convenient for placement and operation on the laboratory table. The dimensions of each component are set, the centrifugal shell part, the upper centrifugal shell 1: the height is about 150-200mm, the inner diameter is designed according to the inner ring gear 101, and it is assumed that the inner diameter of the circle where the inner ring gear is located is 200-250mm, in order to accommodate the rotating parts and other matching structures of the lower centrifugal shell. The lower centrifugal shell 2: the height is about 250-350mm, of which the diameter of the rotating part 201 can be designed to be 150-200mm, which is compatible with the inner diameter of the upper centrifugal shell to ensure the matching rotation. The chromatographic column 202 is 100-150mm in length and 10-20mm in diameter. According to the analysis requirements, the appropriate specifications are selected and placed in the second centrifugal cavity 206. The first centrifugal cavity 205 and the second centrifugal cavity 206 can be designed to be distributed up and down, with heights of 100-150mm and 150-200mm respectively, and the diameter is adapted to the lower centrifugal shell. The diameter of the filter screen 209 is slightly smaller than the diameter of the centrifugal cavity, such as 180-230mm. The length of the fixing rod 210 is adjusted according to the height of the centrifugal cavity, generally 20-30mm shorter than the cavity height, for stabilizing the components. The inner diameter of the pipe (207) is 5-10mm, and the length is determined according to the connection requirements between the centrifugal cavities and the filter shell, generally 100-300mm. The size of the first transmission part 208 and the second transmission part 204 is designed according to the power transmission requirements and the size of the rotating parts, such as a diameter of 30-50mm and a thickness of 10-20mm. The filter shell part, filter shell 3: height 150-200mm, width and length are adapted to the centrifugal shell, such as width 200-250mm, length 250-300mm. The suction tube 301 has an inner diameter of 8-12 mm and a length determined by the distance from the centrifuge housing, typically 200-400 mm. The dimensions of the negative pressure pump 302 are selected based on the required suction force, assuming the occupied space is 100 mm x 100 mm x 100 mm. The first filter holes 303 have a diameter of 0.1-1 mm and are set based on the required filtration accuracy and are reasonably distributed on the filter housing.

[0051] Specific experimental process, used for separation and analysis of biological samples (such as blood, alveolar lavage fluid, cell culture fluid) as an example: Preparation: Component Installation and Inspection: Install the chromatographic column 202, filter membrane 203, filter screen 209, and other components into the lower centrifuge housing 2 as required, ensuring that the fixing rod 210 is securely fastened. Check that the transmission components, including the inner ring gear 101, first transmission component 208, and second transmission component 204, are correctly installed and functioning smoothly. Connect the pipe 207 and suction tube 301 to ensure proper airtightness.

[0052] Equipment commissioning: Turn on the negative pressure pump 302 and check its suction for normal operation. Adjust the negative pressure to an appropriate value (e.g., 20kPa to 50kPa, depending on sample characteristics and experimental requirements). Perform a low-speed pre-run on the centrifuge housing and check that the rotating component 201 rotates smoothly and generates no abnormal noise.

[0053] Sample preparation: Collect biological samples (such as blood, bronchoalveolar lavage fluid, cell culture fluid), add appropriate anticoagulants (such as heparin) to prevent coagulation, and gently invert to mix.

[0054] Centrifugation process Sample loading: slowly add the processed sample into the first centrifugal chamber 205 through a suitable interface.

[0055] Centrifugal separation: The centrifuge is activated, causing the rotating component 201 of the lower centrifuge housing 2 to drive the internal components into high-speed rotation (the speed should be set appropriately based on the sample type; for example, when separating blood components, the speed can be set to 3000-5000 rpm). Under the action of centrifugal force, the different components in the sample are initially separated within the first centrifugal chamber 205 and the second centrifugal chamber 206 due to their density differences. Heavier components (such as blood cells) are located near the outside of the centrifuge chamber, while lighter components (such as plasma) are located near the inside.

[0056] Chromatographic analysis process Component transfer: After the initial centrifugal separation, a portion of the sample (such as plasma) flows through the pipeline 207 into the second centrifugal chamber 206 equipped with the chromatographic column 202.

[0057] Chromatographic separation: The sample components are further separated using a chromatographic column 202. Different substances migrate at different speeds within the column, depending on the characteristics of the column packing and the sample components, resulting in separation of the components. For example, when using a gel filtration column to separate proteins from plasma, small molecules of protein reside longer in the column, while larger molecules of protein elute first.

[0058] Subsequent analysis and closing Sample collection and analysis: Collect filtered target samples, use corresponding detection equipment (such as spectrum analyzer, mass spectrometer, etc.) to perform qualitative and quantitative analysis on the sample components to obtain experimental data.

[0059] Equipment cleaning and arrangement: After the experiment, turn off the power of the equipment, disassemble the cleanable parts (such as the chromatographic column, which needs to be cleaned as required if it is reusable, filter membrane, filter screen, etc.), and clean the equipment with a suitable cleaning agent (such as deionized water, dilute acid or alkali solution, etc.).

[0060] Filter 209, first centrifuge chamber 205, and second centrifuge chamber 206 divide the interior of the centrifuge housing into three centrifuge zones. Each zone contains containers of varying diameters, such as 15ml and 50ml centrifuge tubes, as well as larger ones like 250ml centrifuge bottles and smaller ones like 2ml microtubes. Containers of varying diameters hold varying capacities, so the starting culture volume will vary. Centrifugation speed and time may need to be adjusted, primarily based on centrifugal force (g) rather than speed (rpm), as g is related to radius. However, users may need rpm data, so the centrifuge rotor radius should be considered. Assuming a typical rotor radius of 10cm, the relationship between rpm and g can be calculated using the formula g = 1.118 × 10^-5 × rpm² × r, where r is the rotor radius in centimeters. Here, assuming r = 10cm, rpm = √(g / (1.118 × 10^-5 × 10)).

[0061] Next, divide the sample into three groups: small diameter (for example, 2 ml, approximately 1 cm diameter), medium diameter (15 ml, approximately 2.5 cm diameter), and large diameter (50 ml, approximately 3.5 cm diameter). Each group should include the vessel diameter, capacity, starting culture volume, speed and time for each centrifugation step, and the final suspension volume. Ensure that the centrifugation conditions for each group are appropriate, with the g-ranges for low, medium, and high speed centrifugation consistent with those previously described: 300-500 g for low speed, 10,000-20,000 g for medium speed, and 100,000-200,000 g for high speed. Also, adjust the starting volume based on the vessel capacity; small vessels might process 5-10 ml, medium vessels 20-50 ml, and large vessels 100-200 ml. Estimate the final volume proportionally, for example, to achieve a recovery rate of approximately 10%. Maintaining consistent centrifugal force is crucial; regardless of vessel diameter, centrifugal force parameters should remain within a standard range. Therefore, the speed may need to be adjusted based on the radius. However, assuming the rotor radius is the same, the vessel diameter alone affects the capacity, not the centrifugal force calculation. In addition, users may need specific rpm values, so conversion is required. For example, a low speed of 400g corresponds to approximately 1900rpm (r=10cm), a medium speed of 15000g corresponds to approximately 11600rpm, and a high speed of 110000g corresponds to approximately 31600rpm. These values ​​must be calculated accurately.

[0062] Finally, ensure that each set of data is clearly laid out, step by step, and indicate that parameters can be adjusted to optimize the actual equipment and samples. At the same time, remind users to pay attention to the maximum capacity of the centrifuge and the specifications of the rotor to avoid overloading or parameter errors.

[0063] The following is the centrifugation parameter data for extracellular vesicle extraction based on three containers with different diameters (using common centrifuge tubes / bottles as an example). The grouping design is based on the container specifications, processing volume, and centrifugation conditions: Group 1: Small-diameter containers (e.g., 2 mL centrifuge tubes, approximately 1.0 cm in diameter). Container specifications: 1.0 cm diameter, maximum capacity: 2 mL. Applicable sample volumes: 5–10 mL of blood, bronchoalveolar lavage fluid, and cell culture medium. Centrifugation parameters: Low-speed clarification, speed / centrifugal force: 300 × g (approximately 1500 rpm, assuming a 10 cm rotor radius), time: 10 minutes. Purpose: Remove cell debris and large particles. Medium-speed clarification, speed / centrifugal force: 10,000 × g (approximately 6900 rpm), time: 20 minutes. Purpose: Remove medium-sized impurities such as dead cells and cell debris. High-speed extracellular vesicle sedimentation, speed / centrifugal force: 100,000 × g (approximately 31,600 rpm), time: 90 minutes. Purpose: Enrich extracellular vesicles (precipitate at the bottom of the tube). The final suspension volume is 0.1–0.3 mL (based on 10 mL of starting culture medium).

[0064] Group 2: Medium-diameter containers (e.g., 15 mL centrifuge tubes, approximately 2.5 cm in diameter). Container specifications: 2.5 cm diameter, maximum capacity: 15 mL. Applicable sample volumes: 20–50 mL of blood, bronchoalveolar lavage fluid, or cell culture medium. Centrifugation parameters: Low-speed impurity removal, speed / centrifugal force: 400 × g (approximately 2100 rpm, rotor radius 10 cm), time: 15 minutes. Purpose: Remove cell debris and large particles. Medium-speed clarification, speed / centrifugal force: 15,000 × g (approximately 8700 pm), time: 25 minutes. Purpose: Remove small particles and apoptotic bodies. High-speed extracellular vesicle precipitation, speed / centrifugal force: 110,000 × g (approximately 3,000 rpm), time: 100 minutes. Purpose: Efficient precipitation of extracellular vesicles. The final suspension volume is 0.5–1.5 mL (based on a 50 mL starting culture medium).

[0065] Group 3: Large-diameter containers (e.g., 50 mL centrifuge tubes / bottles, approximately 3.5 cm in diameter). Container specifications: 3.5 cm diameter, maximum capacity: 50 mL. Applicable sample volumes: 100–200 mL of blood, bronchoalveolar lavage fluid, or cell culture medium. Centrifugation parameters: Low-speed impurity removal, speed / centrifugal force: 500 × g (approximately 2500 rpm, rotor radius 10 cm), time: 15 minutes. Purpose: Rapidly remove cellular debris from large-volume samples. Medium-speed clarification, speed / centrifugal force: 20,000 × g (approximately 10,000 rpm), time: 30 minutes. Purpose: Further remove cellular debris and impurities. High-speed extracellular vesicle precipitation, speed / centrifugal force: 120,000 × g (approximately 34,600 rpm), time: 120 minutes. Purpose: Enrich extracellular vesicles from large-volume samples (note the maximum centrifuge capacity and rotor load). Results: Final suspension volume: 1–5 mL (based on 200 mL starting culture medium). Note: Centrifugal force conversion: The above speeds are calculated based on a rotor radius of 10 cm (formula: g = 1.118 × 10−5 × rpm² × rotor radius). The actual speed should be adjusted based on the laboratory centrifuge rotor parameters. Container matching: Ensure that the centrifuge tubes / bottles are compatible with the centrifuge rotor specifications (e.g., compatible with a 50 mL swing-out rotor or angle rotor). Optimization suggestions: If the sample contains significant impurities, a 0.22 μm filter can be added after low-speed centrifugation. After high-speed centrifugation, avoid vigorous pipetting when resuspending the pellet in PBS or exosome-free medium to minimize damage to the exosomes. Depending on the actual sample characteristics (e.g., cell type, culture medium composition) and experimental objectives (e.g., subsequent detection sensitivity), further adjustment of centrifugation parameters or incorporation of ultrafiltration, density gradient centrifugation, or other methods can be used to improve exosome purity.

[0066] Furthermore, the device is used to shorten the sample centrifugation time experimental process; 1. Experimental Preparation (1) Equipment inspection and debugging Check the sealing performance of the connection between the upper centrifugal housing 1 and the lower centrifugal housing 2 to ensure that the inner ring gear 101 is well engaged with the first transmission part 208 and the second transmission part 204, and that the rotating component 201 rotates smoothly without any jamming. The negative pressure pump 302 is turned on, the suction force of the suction tube 301 is detected, and the suction force of the suction tube is stabilized at 30 kPa-50 kPa by adjusting the negative pressure pump parameters to ensure that the first filter hole 303 of the filter housing 3 can filter effectively.

[0067] Perform a no-load test on the equipment, set the centrifugal speed to 80% of the maximum speed, run for 5 minutes, and observe the operating status of the equipment to ensure that there is no abnormal noise and vibration. (2) Sample pretreatment Blood Sample: After collecting blood, quickly transfer it to a container containing an anticoagulant (such as sodium heparin) and gently invert 5-8 times to mix thoroughly. When transferring the blood to a centrifuge tube, perform a preliminary filtration using filter 209 to remove any impurities such as fibrin and minimize interference with separation during centrifugation. Bronchoalveolar lavage fluid sample: After collecting the bronchial lavage fluid, if the fluid is viscous, add an appropriate amount of mucolytic agent (such as acetylcysteine) and incubate in a 37°C water bath for 10 minutes, gently shaking the container to promote dissolution. The treated lavage fluid is initially filtered through filter membrane 203 to remove larger particulate matter. Cell culture medium sample: Observe the cell growth status in the cell culture medium. If the cells are in the logarithmic growth phase, transfer the culture medium to a centrifuge tube. If there are suspended cells in the culture medium, perform low-speed centrifugation (800 rpm for 3 minutes) to initially pellet the cells and reduce mechanical damage to the cells during subsequent high-speed centrifugation. 2. Centrifugal separation process (1) Sample loading The pretreated blood, alveolar lavage fluid, and cell culture fluid samples are slowly added to the first centrifugal cavity 205 of the lower centrifugal shell 2. The amount of sample added does not exceed 70% of the volume of the first centrifugal cavity to ensure that the sample does not overflow during centrifugation. Correctly install and lock the lower centrifuge housing 2 containing the sample and the upper centrifuge housing 1 to ensure the stability of the equipment during operation. (II) Centrifugation parameter setting and operation Based on the optimized parameters determined in the preliminary experiments, the centrifuge speed was set to 4000-5000 rpm, the centrifuge temperature was set to 4°C, and the centrifuge time was set to 60 minutes. The rapid acceleration and deceleration function of the equipment was used to shorten the acceleration and deceleration time during the centrifugation process and improve the overall centrifugal efficiency. Start the centrifugal equipment, closely observe the equipment's operating status, and monitor the speed, temperature and other parameters during the centrifugation process in real time to ensure that the equipment operates stably according to the set parameters. 3. Filtration and Collection Process (1) Sample transfer and filtration After centrifugation, the upper layer of liquid (such as plasma, alveolar lavage fluid supernatant, cell culture fluid supernatant) separated in the first centrifugal chamber 205 is drained to the second centrifugal chamber 206 through the pipeline 207, and the sample is further separated and purified using the chromatographic column 206. The negative pressure pump 302 is started to suck the sample processed by the chromatographic column in the second centrifugal chamber 206 into the filter housing 3 through the suction tube 301. The sample is finely filtered through the first filter hole 303 to remove tiny particle impurities and residual chromatographic column fillers. (2) Target component collection A collection container is placed below the filter housing 3 to collect the filtered target component liquid, such as separated plasma, purified active components of the lavage fluid, specific metabolites in the cell culture fluid, etc.

[0068] After the collection is completed, the negative pressure pump 302 is turned off, the filter housing 3 is disassembled, and the collected samples are marked with information such as the sample type, centrifugation time, and collection time. 4. Experimental results detection and analysis (1) Sample testing Blood samples: Use a hematology analyzer to detect the residual amount of blood cells in the plasma after separation to evaluate the centrifugal separation effect; use a biochemical analyzer to detect various biochemical indicators in the plasma (such as blood sugar, blood lipids, liver function indicators, etc.), and compare them with the test results of the original sample to determine the impact of shortening the centrifugation time on the sample composition. For BALF samples, use a protein quantification kit to detect the protein content and purity in the lavage fluid. Observe the morphology and activity of cells in the lavage fluid under a microscope to assess the effects of the centrifugation process on the cells. Cell culture fluid samples: Use a cell counter to detect the viability and number of cells in the culture fluid. Use high-performance liquid chromatography (HPLC) and other methods to detect the content of specific metabolites in the culture fluid, and analyze whether shortening the centrifugation time affects the separation effect of cell metabolites. (2) Data Analysis The test results of this experiment (centrifugation time 60 minutes) were compared with the test results of the original centrifugation time (120 minutes), and the significance of the data difference was analyzed using statistical methods (such as t-test).

[0069] Based on the data analysis results, the feasibility and effectiveness of using this equipment to shorten the centrifugation time from 120 minutes to 60 minutes were evaluated, and the problems and improvement directions in the experimental process were summarized. 5. Cleaning and sorting of experimental equipment Disassemble the upper centrifugal housing 1, lower centrifugal housing 2, filter housing 3 and other components, rinse them with deionized water to remove residual samples and impurities. The chromatographic column 206, the filter membrane 203, the filter screen 209, etc. are specially cleaned and cleaned and regenerated using a suitable cleaning solution according to the requirements of their instructions to ensure the accuracy of the next experiment.

[0070] After cleaning, dry the equipment parts and store them properly. Close and clean the laboratory table to prepare for the next experiment. like Figure 16As shown, a filter screen 209 is provided on the top of the first centrifugal chamber 205 and is fixedly connected. The filter screen 209 is located between the first filter hole 303 and the second filter hole. The plasma supernatant after being filtered through the first filter hole 303 flows into the filter screen 209 for secondary filtration, and the plasma supernatant after being filtered through the filter screen 209 flows into the second filter hole for tertiary filtration. This improves the filtration efficiency of the plasma supernatant. At the same time, the filter screen 209, the first centrifugal chamber 205, and the second centrifugal chamber 206 are designed in a proportional manner. That is, the area of ​​the filter screen 209 is smaller than that of the first centrifugal chamber 205, and the diameter of the first centrifugal chamber 205 is smaller than that of the second centrifugal chamber 206. In this way, the filter screen 209, the first centrifugal chamber 205, and the second centrifugal chamber 206 divide the interior of the centrifugal housing into three centrifugal zones, so that the three centrifugal zones can respectively centrifuge apoptotic bodies with the largest diameter, separate microvesicles, and separate exosomes with the smallest diameter.

[0071] like Figure 6 、 8 , 12, 13, 14, 15, the transmission shaft model is DZ9114315156, the transmission shaft is fixed with a filter membrane 203, the filter membrane 203 is a medical waterproof breathable membrane bacteria filter 50mm circular hydrophobic filter membrane, the filter membrane 203 and the bottom of the lower centrifugal shell 2 are spaced apart, the spacing is used to receive the filtered waste liquid; the filter membrane 203 is provided with a plurality of chromatographic columns 202 on the circumference, the chromatographic column 202 is LAEK-A300, the lower end of the chromatographic column 202 is bonded to the upper end surface of the filter membrane 203, and the filter membrane 2 03 has an upper end lower than the second centrifugal chamber 206, and an upper end of each chromatographic column 202 is arranged corresponding to each second centrifugal chamber 206; the plasma supernatant flows toward the chromatographic column 202 under the centrifugal effect of the second centrifugal chamber 206, so that the extracellular vesicles are bound to the chromatographic column 12; chromatographic column balancing: a size exclusion chromatographic column is taken and the column is balanced; 150 μL of the diluted sample is added to the chromatographic column, and the initial 1 mL of filtrate that flows out is discarded; extracellular vesicles are collected: multiple fractions containing extracellular vesicles are collected, and a total of 600 μL of extracellular vesicles are collected.

[0072] Continue as Figure 6 、 8 As shown in Figures 12, 13, 14, and 15, the filter membrane 203 is arrayed with waste liquid filter holes, and a gap is left between the filter membrane 203 and the bottom of the lower centrifugal shell 2, which is used to receive the filtered waste liquid; the second centrifugal chamber 206 flows toward the chromatographic column 202 due to centrifugal action, and part of the waste liquid flows into the waste liquid filter holes arrayed on the filter membrane 203 and is collected in the gap between the filter membrane 203 and the bottom of the lower centrifugal shell 2.

[0073] like Figure 4 、 5As shown in , 6, 11, and 17, a first transmission part 208 is fixedly provided on the transmission shaft, and the first transmission part 208 is a disc gear. The first transmission part 208 is arranged below the first centrifugal cavity 205, and a second transmission part 204 is arranged below the second centrifugal cavity 206. The second transmission part 204 is an outer ring gear, and the hollow part on the second transmission part 204 is aligned with the hollow part of the second centrifugal cavity 206. The second transmission part 204 is fixedly connected to the lower part of the first centrifugal cavity 205 through a fixing rod, and the first transmission part 208 is rotationally connected to the second transmission part 204.

[0074] like Figure 14 、 15 As shown, an inner ring gear 101 is provided on the lower inner edge of the upper centrifugal housing 1, and the inner ring gear 101 is engaged with the second transmission part 204. In this way, the second transmission part 204 is engaged and rotates around the inner ring gear 101, that is, the upper centrifugal housing 1 does not rotate, and the filter screen 209, the first centrifugal cavity 205, and the inner wall of the second centrifugal cavity 206 rotate several times, which can buffer the collision caused by excessive centrifugal force and reduce damage to extracellular vesicles.

[0075] The development of an extracellular vesicle extraction device is crucial for controlling different centrifugal forces for the same raw material and protecting the integrity of extracellular vesicles under excessive centrifugal forces. By varying the radius of the centrifugal zones, the device can differentiate centrifugal forces to achieve the separation of apoptotic bodies, microvesicles, and small extracellular vesicles. A planetary gear design allows the zones receiving the greatest centrifugal force to orbit around the center while simultaneously generating self-rotation, providing a buffering effect and protecting the integrity of the isolated exosomes. Chromatographic columns are positioned at corresponding positions in the three centrifugal zones to assist in extracellular vesicle purification. This approach, which allows for the zoned extraction of three types of extracellular vesicles, the protection of the exosome cell wall, and the integration of ultracentrifugation and chromatography, is crucial for improving ease of use and enhancing medical quality. This not only reflects advancements in medical technology but also safeguards the rights and safety of patients.

[0076] Centrifugal extraction: From sample pretreatment to the final extraction of extracellular vesicles, multiple centrifugation steps are required, and the entire process can take several hours or even longer. For example, for blood sample processing, the ultracentrifugation step alone can take 70-120 minutes. Combined with the initial plasma separation steps, the total time is long, which is inefficient for situations where rapid experimental results are required. This also impacts sample throughput: Due to the long single centrifugation time, when processing a large number of samples, the samples must wait in line for centrifugation, which prolongs the overall experimental cycle and cannot meet the needs of rapid large-scale sample extraction.

[0077] The usage process is as follows: start the motor, the motor drives the transmission shaft to rotate, the transmission shaft drives the filter membrane 203, the chromatographic column 202, the first centrifugal chamber 205, the second centrifugal chamber 206, the filter screen 209 to rotate, and drives the first transmission part 208 and the second transmission part 204 to rotate. Among them, the second transmission part 204 rotates around the inner ring gear 101, that is, the inner ring gear 101 is fixed. After rotating several circles, the inner ring gear 101 can buffer the collision caused by excessive centrifugal force and reduce damage to extracellular vesicles. Negative pressure pump 302 is driven to draw blood, BALF, and cell culture fluid through suction tube 301 through first filter hole 303 into filter screen 209. Filter screen 209 filters out impurities in the blood, BALF, and cell culture fluid, and the fluid then flows directly into first centrifugal chamber 205. Under centrifugal force, the blood, BALF, and cell culture fluid enter pipe 207, pass through pipe 207 into second centrifugal chamber 206, and continue to flow through centrifugation into chromatography column 202 and filter membrane 203. Waste fluid is filtered out through the waste fluid holes in filter membrane 203 and stored at the bottom of lower centrifugal housing 2. Finally, the fluid is combined with chromatography column 202. Column balancing: A size exclusion chromatography column is taken and equilibrated. 150 μL of the diluted sample is added to the column, and the initial 1 mL of filtrate is discarded. Extracellular vesicle collection: Multiple fractions containing extracellular vesicles are collected, totaling 600 μL of extracellular vesicles.

[0078] The present invention has been described in detail above with reference to the embodiments of the accompanying drawings. A person skilled in the art can make various modifications to the present invention based on the above description. Therefore, certain details in the embodiments should not be construed as limiting the present invention. The scope of protection of the present invention shall be determined by the scope defined by the appended claims.

Claims

1. An extracellular vesicle extraction device, characterized in that: include: A filter assembly is provided above the centrifugal housing to filter the plasma supernatant; The centrifugal component is arranged in the centrifugal shell and is located below the filter component, and is used to centrifuge the plasma supernatant filtered by the filter component.

2. The extracellular vesicle extraction device according to claim 1, characterized in that The filter assembly comprises: The filter housing is arranged above the centrifugal housing. The bottom of the filter housing is provided with a first filter hole. The plasma supernatant flows into the centrifugal assembly after being filtered through the filter hole.

3. The extracellular vesicle extraction device according to claim 2, characterized in that The filter assembly further comprises: The negative pressure pump is arranged on the filter housing, and the negative pressure pump sucks the plasma supernatant through the suction tube and transports it to the filter housing for filtration.

4. The extracellular vesicle extraction device according to claim 2, wherein The centrifugal assembly comprises: A rotating component, the bottom of which is connected to the bottom of the centrifugal housing; The first centrifugal cavity is arranged on the top of the rotating component. The top of the first centrifugal cavity is provided with a second filter hole. The plasma supernatant in the filter shell flows into the first centrifugal cavity through the first filter hole and the second filter hole.

5. The extracellular vesicle extraction device according to claim 4, characterized in that The centrifugal assembly further comprises: a second centrifugal cavity, disposed on the periphery of the first centrifugal cavity and connected to the first centrifugal cavity through a pipeline, wherein a through hole is provided at the lower portion of the second centrifugal cavity; a filter membrane disposed below the second centrifugal chamber, wherein the rotating component passes through the filter membrane and is fixedly connected thereto, and a distance is left between the filter membrane and the bottom of the housing; A chromatographic column is arranged on the filter membrane, and the chromatographic column is aligned with the through hole at the lower part of the second centrifugal cavity.

6. The extracellular vesicle extraction device according to claim 5, characterized in that The filter membrane is provided with waste liquid filtering holes in an array, and the plasma supernatant on the filter membrane flows into the gap between the filter membrane and the bottom of the shell through the waste liquid filtering holes.

7. The extracellular vesicle extraction device according to claim 5, characterized in that There are multiple chromatographic columns, which are circumferentially arranged on the filter membrane; There are multiple second centrifugal cavities, which are circumferentially arranged on the outer wall of the first centrifugal cavity. The multiple second centrifugal cavities correspond one-to-one to the multiple chromatographic columns.

8. The extracellular vesicle extraction device according to claim 4, wherein The centrifugal assembly further comprises: a first transmission part, disposed on the rotating component and located below the first centrifugal cavity; The second transmission part is arranged below the second centrifugal cavity and is rotationally connected to the first transmission part.

9. The extracellular vesicle extraction device according to claim 4, characterized in that The centrifugal assembly further comprises: The filter screen is arranged on the top of the first centrifugal cavity and is fixedly connected. The filter screen is located between the first filter hole and the second filter hole. The plasma supernatant filtered by the first filter hole flows into the filter screen for secondary filtration, and the plasma supernatant filtered by the filter screen flows into the second filter hole for tertiary filtration.

10. The extracellular vesicle extraction device according to claim 2, wherein The centrifugal housing comprises: an upper centrifugal housing, wherein the upper centrifugal housing is provided with a through hole, the filter housing is provided on the centrifugal housing, wherein the first filter hole is communicated with the through hole; The lower centrifugal shell is arranged below the upper centrifugal shell and is rotatably connected to the upper centrifugal shell. The centrifugal assembly is arranged in the upper centrifugal shell and the lower centrifugal shell.

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