Exosome extraction device and exosome extraction method

Abstract

The present application discloses an exosome extraction device and an exosome extraction method, and relates to the field of exosome extraction. The exosome extraction device includes a loading platform, a filtering component, a liquid injection device and a vibration device. A placement tank is provided on the loading platform, and a liquid outlet connected to the placement tank is provided. The filtering component is detachably installed in the placement tank, and the filtering component includes a filter membrane and a liquid inlet and a waste liquid outlet located on two opposite sides of the filter membrane, and the waste liquid outlet is connected to the liquid outlet. The vibration device is arranged near the filtering component, and is used to vibrate the particles in the filtering component. The exosome extraction device provided in the present application vibrates the liquid, particles and filter membrane in the filter component through the vibration device, so as to avoid the particles from stopping on the filter membrane, prevent the filter membrane from being blocked, and improve the extraction efficiency of exosomes in the liquid.

Description

Technical Field

[0001] The present application relates to the technical field of exosome extraction, and in particular to an exosome extraction device and an exosome extraction method. Background Art

[0002] Exosomes are a type of extracellular vesicles with a size of 30-150nm. They are released outside the cell in the form of exocytosis after the fusion of multivesicular bodies in the cell with the cell membrane. They are widely present in cell culture supernatant and various body fluids, including blood, lymph, saliva, urine, semen, milk, etc.

[0003] For the study of exosomes, the first step of separation and purification is difficult. The lack of technically standardized tools for high-throughput and high-purity separation poses a great obstacle to the discovery of downstream biomarkers. Existing separation methods include ultracentrifugation, ultrafiltration, precipitation, immunoaffinity and microfluidics. Although ultracentrifugation is still the most commonly used "gold standard" separation method in the research community, it requires instruments, manpower and operation time, so it is difficult to truly apply it in clinical practice. The precipitation method using the hydrophilicity of polyethylene glycol has low purity for downstream analysis due to polymer contamination. The capture method based on immunomagnetic beads is expensive and heavily dependent on specific antibodies, so the collected products cannot reflect all exosomes.

[0004] Ultrafiltration is actually a very good purification method. It does not require high-end equipment and is simple to operate. It does not affect the biological activity of exosomes. The equipment used in ultrafiltration requires a syringe with a fixed injection speed and a device with an ultrafine filter membrane. Connect the syringe containing the sample to the device with the ultrafine filter membrane, start the fixed speed injection, and the exosomes obtained can be enriched on the filter membrane. The excess liquid is discharged during the injection process. Finally, the sample on the filter membrane is extracted and the next step can be carried out.

[0005] However, under the actual injection pressure of the syringe, exosomes are often intercepted on the filter membrane. The accumulation of particles on the filter membrane eventually leads to liquid blockage. Therefore, during the experiment, the device with the filter membrane needs to be rotated manually from time to time to reduce the impact of blockage. At the same time, the product finally obtained by the ultrafiltration method is usually of average purity, and the filtration speed is slow due to the blockage of the filter membrane, and a high pressure is generated inside. At the least, the syringe will rebound, resulting in incomplete injection of the sample. This is a factor that affects the filtration efficiency and purity. In severe cases, the device with the filter membrane or the syringe and other injection pipelines will be damaged during the operation. Summary of the invention

[0006] In view of this, the purpose of the present application is to overcome the deficiencies in the prior art. The present application provides an exosome extraction device to at least solve the problem of filter membrane clogging in the prior art.

[0007] This application provides:

[0008] An exosome extraction device, comprising:

[0009] Base;

[0010] A loading platform, wherein the loading platform is provided with a placement groove and a liquid outlet communicated with the placement groove;

[0011] A filter assembly, the filter assembly is detachably mounted in the placement tank, and the filter assembly comprises a filter membrane and a liquid inlet and a waste liquid outlet located on two opposite sides of the filter membrane, the waste liquid outlet is communicated with the liquid outlet;

[0012] A liquid injection device, the liquid injection device is used to output liquid into the filter assembly or absorb liquid;

[0013] A vibration device is arranged close to the filter assembly and is used to generate vibrations for the liquid, particles and the filter membrane in the filter assembly.

[0014] In addition, the exosome extraction device according to the present application may also have the following additional technical features:

[0015] In some embodiments of the present application, the exosome extraction device further comprises a rotating member, the rotating member is rotatably disposed on the base, and the vibration device is disposed on the rotating member.

[0016] In some embodiments of the present application, the loading station comprises:

[0017] A loading base, one end of which is connected to the rotating member, and the other end of which is provided with a mounting hole;

[0018] A loading turntable, the loading turntable is rotatably arranged at the mounting hole, the placement slot is arranged on the loading turntable, and a receiving slot communicating with the placement slot is arranged on a side away from the placement slot;

[0019] An end cover is connected to a side of the loading turntable where the receiving groove is provided, and covers the receiving groove, and the end cover is provided with the liquid outlet.

[0020] In some embodiments of the present application, the loading platform also includes a first pipe joint with openings at both ends, the first pipe joint is passed through the connection between the placement groove and the receiving groove, and abuts against the edge of the waste liquid port to connect the first pipe joint with the waste liquid port.

[0021] In some embodiments of the present application, the mounting hole is a stepped hole;

[0022] The loading turntable is sleeved with a first bearing, which is located in the stepped hole on a side away from the end cover; and / or, the loading turntable is also sleeved with a second bearing, which is located in the stepped hole on a side close to the end cover.

[0023] In some embodiments of the present application, the liquid injection device comprises:

[0024] An infusion pump having an infusion port for sucking in or delivering liquid;

[0025] The telescopic mechanism comprises a driving member and a moving member, wherein the driving member is fixedly connected to the injection pump, and an output end of the driving member is connected to the moving member, and is used to drive the moving member to move relative to the loading platform;

[0026] A turntable assembly connected to the moving part and moving with the moving part;

[0027] Wherein, the turntable assembly includes a rotating mechanism, a turntable base, an injection turntable, a mounting seat and a second pipe joint, the mounting seat is connected to the moving part, and the rotating mechanism and the turntable base are respectively arranged on the mounting seat; the injection turntable is rotatably connected to the turntable base and is connected to the rotating end of the rotating mechanism; the second pipe joint has a first end and a second end that are relatively arranged, the second pipe joint is passed through the injection turntable, and the first end is connected to the infusion port through a hose, and the second end can abut against the edge of the liquid inlet to connect the second pipe joint with the liquid inlet.

[0028] In some embodiments of the present application, the rotating mechanism includes a driving motor and a transmission member, the driving motor drives the transmission member to rotate, and the transmission member is drivingly connected to the injection turntable;

[0029] The driving motor and the turntable base are respectively fixedly connected to the mounting base.

[0030] In some embodiments of the present application, the transmission member is a first gear, the first gear is connected to the output end of the drive motor, and the injection turntable is provided with a second gear meshing with the first gear; or

[0031] The transmission member includes a first pulley and a belt, the first pulley is connected to the output end of the driving motor, a second pulley is arranged on the injection turntable, and the belt transmission is connected to the first pulley and the second pulley.

[0032] In some embodiments of the present application, the turntable base is provided with a step hole, the injection turntable is provided with a third bearing, and the third bearing is located on the side of the step hole close to the moving part; and / or, the injection turntable is also provided with a fourth bearing, and the fourth bearing is located on the side of the step hole away from the moving part.

[0033] In some embodiments of the present application, an annular groove is formed on the end surface of the second end, and the annular groove is used for clamping with the probe gun head.

[0034] In some embodiments of the present application, a vent hole communicating with the annular groove is provided on the second pipe joint.

[0035] In some embodiments of the present application, the exosome extraction device further includes a transverse moving mechanism, which is disposed on the rotating member, and the injection pump and the driving member are disposed on the transverse moving mechanism through a slider, and the transverse moving mechanism is used to drive the slider to move along the length direction of the base.

[0036] In some embodiments of the present application, the lateral movement mechanism includes a bracket, a driving part and a transmission part, the bracket is installed on the rotating member, the driving part and the transmission part are respectively installed on the bracket, and the driving part is used to drive the transmission part to rotate, and the slider is arranged on the transmission part.

[0037] In some embodiments of the present application, the filter assembly includes an upper cover, the filter membrane, filter paper and a lower cover, the upper cover is provided with the liquid inlet, the lower cover is provided with a groove, the filter membrane and the filter paper are respectively located in the groove, and the filter paper is located between the filter membrane and the bottom of the groove, the lower cover is provided with the waste liquid port, the waste liquid port is connected to the groove, and the upper cover is connected to the lower cover.

[0038] In some embodiments of the present application, a filtration cavity is formed between the upper cover and the filtration membrane.

[0039] In some embodiments of the present application, the upper cover and the lower cover are both circular, the liquid inlet is located at a non-center position of the upper cover, and the waste liquid port is located at a non-center position of the lower cover.

[0040] In some embodiments of the present application, the exosome extraction device also includes a sample consumable rack, and the sample consumable rack is provided with at least one first mounting position and at least one second mounting position, the first mounting position is used to place a test tube, and the second mounting position is used to place a probe gun tip.

[0041] In some embodiments of the present application, a waste box with an opening is further provided on the sample consumables rack, a gun tip scraper is provided above the waste box, and a notch matching the probe gun tip is provided on the gun tip scraper.

[0042] In some embodiments of the present application, the vibration device is any one of an ultrasonic instrument, an ultrasonic transducer, a vortex vibrator, a high-frequency vibration motor, a pneumatic vibrator or an exciter.

[0043] The embodiments of the present application also provide a method for extracting exosomes, using the exosome extraction device described in the above embodiments, comprising the following steps:

[0044] The injection device extracts a probe gun tip on the sample consumables rack through the second pipe joint, moves to the wetting reagent tube through the lateral moving mechanism to take the wetting agent and temporarily stores it in the injection pump, and after sucking the wetting agent, moves to the gun tip scraper to scrape off the probe gun tip;

[0045] The injection turntable is moved onto the filter assembly through the telescopic mechanism and pressed and sealed, and the absorbed wetting agent is injected into the filter assembly and the filter paper is wetted;

[0046] The injection device extracts another probe gun tip on the sample consumable rack through the second pipe joint, and moves to the blood sample test tube through the lateral moving mechanism to take the blood sample and temporarily store it in the injection pump, and after sucking the blood sample, moves to the gun tip scraper to scrape off the probe gun tip;

[0047] The injection turntable is moved to the filter assembly through the telescopic mechanism and pressed and sealed. The injection device and the loading platform are rotated 90° counterclockwise through the rotating member, so that the second pipe joint on the injection turntable is in the lower position and the first pipe joint on the loading platform is in the upper position. The injection device injects the blood sample into the filter assembly at a constant speed for filtration. During the filtration process, the vibration device starts to work;

[0048] The injection device extracts another probe gun tip on the sample consumables rack through the second pipe joint, and moves to the cleaning agent tube through the lateral moving mechanism to take the cleaning agent and temporarily store it in the injection pump, and after sucking the cleaning agent, moves to the gun tip scraper to scrape off the probe gun tip;

[0049] The injection turntable is moved to the filter assembly through the telescopic mechanism and pressed and sealed. The injection device and the loading platform are rotated 90° counterclockwise through the rotating parts. At the same time, the rotating mechanism of the injection device drives the filter assembly on the loading platform to rotate together, so that the second pipe joint on the injection turntable is in the lower position and the first pipe joint on the loading platform is in the upper position. The injection device injects the cleaning agent into the filter assembly at a constant speed for filtration. During the filtration process, the vibration device starts to work;

[0050] The liquid injection device extracts another probe gun head on the sample consumables rack through the second pipe joint, and moves the probe gun head to the liquid inlet on the filter assembly to absorb the filtered sample;

[0051] The obtained sample was injected into the exosome collection reagent tube to complete the filtration experiment.

[0052] In some embodiments of the present application, the method for extracting exosomes further comprises the following steps:

[0053] Pre-treating the blood sample in the blood sample tube to be placed on the sample consumables rack;

[0054] The pretreatment includes: centrifuging, diluting and filtering the blood sample.

[0055] Furthermore, the pretreatment also includes enzymatic hydrolysis of the blood sample.

[0056] Furthermore, the centrifugal force of the centrifugation is 9000g to 12000g; and the centrifugal time is 15min to 20min.

[0057] Furthermore, the pore size of the filter for filtering is 0.15 μm to 0.22 μm.

[0058] Furthermore, the temperature of the enzymolysis is 36°C to 40°C; the time of the enzymolysis is 10min to 30min.

[0059] In some embodiments of the present application, the vibration frequency of the vibration device is 20 Hz to 100 kHz.

[0060] Compared with the prior art, the beneficial effect of the present application is as follows: the present application proposes an exosome extraction device, through the setting of a loading platform and an ultrasonic transducer, when a sample reagent is subjected to a filtering experiment, the filtering component is assembled into a placement slot, and the sample reagent is injected into the filtering component through a liquid injection device, thereby performing a filtering operation. In this process, the vibration device is turned on to vibrate the liquid, particles and filter membrane in the filtering component, and the particles and the filter membrane move relative to each other, that is, the particles are constantly vibrated on the filter membrane, so that the particles are prevented from stopping on the filter membrane and the filter membrane is prevented from being blocked, thereby improving the extraction efficiency of exosomes in the liquid. BRIEF DESCRIPTION OF THE DRAWINGS

[0061] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings required for use in the embodiments will be briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present application and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without paying creative work.

[0062] Figure 1 A three-dimensional schematic diagram of an exosome extraction device in some embodiments of the present application is shown;

[0063] Figure 2 A schematic diagram showing the structure of an exosome extraction device without a liquid injection device in some embodiments of the present application is shown;

[0064] Figure 3 A schematic structural diagram of a liquid injection device from one perspective in some embodiments of the present application is shown;

[0065] Figure 4 Shows Figure 3 AA-direction cross-sectional structural diagram;

[0066] Figure 5 A schematic structural diagram of a second pipe joint from one perspective in some embodiments of the present application is shown;

[0067] Figure 6 Shows Figure 5 Schematic diagram of the cross-sectional structure in the middle BB direction;

[0068] Figure 7 A schematic diagram showing the clamping state of the annular clamping groove of the second pipe joint and the probe gun head in some embodiments of the present application is shown;

[0069] Figure 8 A schematic diagram of the three-dimensional structure of a loading platform in some embodiments of the present application is shown;

[0070] Fig. 9 A schematic diagram of the structure of a loading platform from one perspective in some embodiments of the present application is shown;

[0071] Fig.10 Shows Fig. 9 Schematic diagram of the cross-sectional structure in CC direction;

[0072] Fig.11 A schematic diagram of the structure of a filter assembly from one perspective in one embodiment of the present application is shown;

[0073] Fig.12 Shows Fig.11 Schematic diagram of the cross-sectional structure in the middle DD direction;

[0074] Fig.13 A schematic structural diagram of a filter assembly from one perspective in another embodiment of the present application is shown;

[0075] Fig.14 Shows Fig.13 Schematic diagram of the cross-sectional structure in the EE direction;

[0076] Fig.15 A schematic diagram of the three-dimensional structure of a sample consumables rack in some embodiments of the present application is shown;

[0077] Fig.16 A schematic diagram showing an implementation of a vibration device in some embodiments of the present application is shown;

[0078] Fig.17 The schematic diagram of the operation process of the exosome extraction device in some embodiments of the present application is shown Figure 1 ;

[0079] Fig.18 The schematic diagram of the operation process of the exosome extraction device in some embodiments of the present application is shown Figure 2 ;

[0080] Fig.19 The schematic diagram of the operation process of the exosome extraction device in some embodiments of the present application is shown Figure 3 ;

[0081] Fig. 20 The schematic diagram of the operation process of the exosome extraction device in some embodiments of the present application is shown Figure 4 ;

[0082] Fig.21 The schematic diagram of the operation process of the exosome extraction device in some embodiments of the present application is shown Figure 5 ;

[0083] Fig. 22 Schematic diagrams of the method for extracting exosomes in other embodiments of the present application are shown.

[0084] Description of main component symbols:

[0085] 100-exosome extraction device; 10-base; 20-rotating member; 30-loading platform; 31-loading base; 311-mounting hole; 32-loading turntable; 321-placing slot; 322-accommodating slot; 33-end cover; 331-liquid outlet; 34-first pipe joint; 35-first bearing; 36-second bearing; 40-filter assembly; 41-upper cover; 411-liquid inlet; 42-filter membrane; 43-filter paper; 44-lower cover; 441-waste liquid outlet; 442-groove; 45-filter cavity; 50-liquid injection device; 51-liquid injection pump; 511-infusion port; 52-telescopic mechanism; 521-driving member; 522-moving member; 53-rotating mechanism; 531-driving motor; 5 32-transmission member; 54-turntable assembly; 541-turntable base; 5411-step hole; 542-injection turntable; 5421-second gear; 543-second pipe joint; 5431-first end; 5432-second end; 5433-annular groove; 5434-vent; 544-mounting seat; 55-third bearing; 56-fourth bearing; 57-L-shaped bracket; 60-vibration device; 70-lateral movement mechanism; 71-slider; 72-bracket; 73-driving unit; 74-transmission unit; 80-sample consumables rack; 801-first mounting position; 802-second mounting position; 81-waste box; 82-gun tip scraper; 821-notch; 200-probe gun tip; 300-test tube. DETAILED DESCRIPTION

[0086] The embodiments of the present application are described in detail below, and examples of the embodiments are shown in the accompanying drawings, wherein the same or similar reference numerals throughout represent the same or similar elements or elements having the same or similar functions. The embodiments described below with reference to the accompanying drawings are exemplary and are only used to explain the present application, and cannot be understood as limiting the present application.

[0087] In the description of the present application, it should be understood that the terms "center", "longitudinal", "lateral", "length", "width", "thickness", "up", "down", "front", "back", "left", "right", "vertical", "horizontal", "top", "bottom", "inside", "outside", "clockwise", "counterclockwise", "axial", "radial", "circumferential" and the like indicate orientations or positional relationships based on the orientations or positional relationships shown in the accompanying drawings, and are only for the convenience of describing the present application and simplifying the description, and do not indicate or imply that the referred device or element must have a specific orientation, be constructed and operated in a specific orientation, and therefore should not be understood as a limitation on the present application.

[0088] In addition, the terms "first" and "second" are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Thus, a feature defined as "first" or "second" may explicitly or implicitly include one or more of the features. In the description of this application, the meaning of "plurality" is two or more, unless otherwise clearly and specifically defined.

[0089] In this application, unless otherwise clearly specified and limited, the terms "installed", "connected", "connected", "fixed" and the like should be understood in a broad sense, for example, it can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection or an electrical connection; it can be a direct connection or an indirect connection through an intermediate medium, it can be the internal connection of two elements or the interaction relationship between two elements. For ordinary technicians in this field, the specific meanings of the above terms in this application can be understood according to specific circumstances.

[0090] In the present application, unless otherwise clearly specified and limited, a first feature being “above” or “below” a second feature may mean that the first and second features are in direct contact, or the first and second features are in indirect contact through an intermediate medium. Moreover, a first feature being “above”, “above”, and “above” a second feature may mean that the first feature is directly above or obliquely above the second feature, or simply means that the first feature is higher in level than the second feature. A first feature being “below”, “below”, and “below” a second feature may mean that the first feature is directly below or obliquely below the second feature, or simply means that the first feature is lower in level than the second feature.

[0091] like Figure 1 and Figure 2 As shown, an embodiment of the present application provides an exosome extraction device 100, which is mainly used for filtering and extracting exosomes. The exosome extraction device 100 includes a base 10, a loading platform 30, a filtering assembly 40, a liquid injection device 50 and a vibration device 60.

[0092] The rotating member 20 is rotatably disposed on the base 10, and can be driven by a motor to rotate on the base 10. The loading platform 30 is provided with a placement groove 321, and a liquid outlet 331 communicated with the placement groove 321 is provided.

[0093] The filter assembly 40 is detachably installed in the placement groove 321, and the filter assembly 40 includes a filter membrane 42 and a liquid inlet 411 and a waste liquid port 441 located on two opposite sides of the filter membrane 42. The waste liquid port 441 is connected to the liquid outlet 331, so that the filtered waste liquid flows into the waste liquid pool or waste liquid tank through the liquid outlet 331.

[0094] The liquid injection device 50 is used to output liquid or absorb liquid into the filter assembly 40. The vibration device 60 is arranged near the filter assembly 40 to achieve vibration of the liquid, particles and filter membrane 42 in the filter assembly 40.

[0095] In the exosome extraction device 100 provided in the embodiment of the present application, when a filtering experiment is performed on a sample reagent, the filter assembly 40 is assembled into the placement groove 321, and the placement groove 321 is used to place and position the filter assembly 40, and the sample reagent is injected into the filter assembly 40 through the liquid injection device 50 to perform a filtering operation. During this process, the vibration device 60 is turned on to vibrate the liquid, particles and filter membrane 42 in the filter assembly 40, so as to prevent the particles from resting on the filter membrane 42 and prevent the filter membrane 42 from being blocked, thereby improving the extraction efficiency of exosomes in the liquid.

[0096] Optionally, the exosome extraction device 100 further includes a rotating member 20, the rotating member 20 is rotatably disposed on the base 10, and the vibration device 60 is disposed on the rotating member 20. In addition, when the rotating member 20 is driven to rotate a preset angle, the loading platform 30 also rotates a preset angle, such as 90 degrees counterclockwise. Of course, in other embodiments, such as Fig.16 As shown, the vibration device 60 can also be arranged on the base 10 or at the bottom of the loading platform 30 near the base 10, that is, it can vibrate the liquid, particles and filter membrane 42 in the filter assembly 40.

[0097] It should be noted that the filter assembly 40 is a consumable, and the filter assembly 40 is replaced after each experiment. In addition, taking the base 10 as an L-shaped plate as an example, the rotating member 20 can be rotatably arranged on the vertical plate of the L-shaped plate through a deep groove ball bearing, and at the same time, the motor is fixed on the vertical plate, and the motor is meshed with the rotating member 20 through an output gear. Of course, the motor can also be connected to the rotating member 20 through a belt transmission mechanism. In other embodiments, the motor can also be connected to the rotating member 20 through a chain sprocket, but it is not limited to this. It should be understood that the motor can be provided with a reducer, which is transmitted to the rotating member 20 through the reducer. The vibration device 60 is any one of an ultrasonic instrument, an ultrasonic transducer, a vortex vibrator, a high-frequency vibration motor, a pneumatic vibrator or an exciter. The vibration frequency of the vibration device is 20Hz~100kHz (for example 20Hz, 25Hz, 30Hz, 40Hz, 80Hz, 100Hz, 120Hz, 200Hz, 400Hz, 700Hz, 900Hz, 1kHz, 5kHz, 10kHz, 20kHz, 30kHz, 33kHz, 35kHz, 38kHz, 40kHz, 43kHz, 45kHz, 48kHz, 50kHz, 70kHz, 80kHz, 100kHz). In the present embodiment, the vibration device is selected to use an ultrasonic transducer as an example. Of course, in other embodiments, the vibration device is not limited to an ultrasonic instrument, an ultrasonic transducer, a vortex vibrator, a high-frequency vibration motor, a pneumatic vibrator or an exciter, and can also be other equipment with a vibration function.

[0098] like Figures 8 to 10 As shown, in some embodiments of the present application, optionally, the loading platform 30 includes a loading base 31, a loading turntable 32 and an end cover 33. One end of the loading base 31 is connected to the rotating member 20, and the other end is provided with a mounting hole 311, the loading turntable 32 is rotatably penetrated at the mounting hole 311, and the placement slot 321 is opened on the loading turntable 32.

[0099] The loading turntable 32 has a receiving groove 322 on one side away from the placement groove 321, which is connected to the placement groove 321. The end cover 33 is connected to the side of the loading turntable 32 on which the receiving groove 322 is provided, and covers the receiving groove 322, and the end cover 33 is provided with the liquid outlet 331. It should be noted that a soft rubber hose can be installed at the liquid outlet 331, and the waste liquid is discharged into the waste liquid pool or waste liquid tank through the soft rubber hose.

[0100] In this embodiment, the loading turntable 32 is rotatably disposed on the loading base 31 so that the loading turntable 32 rotates relative to the loading base 31 . Meanwhile, the placement slot 321 is opened on the loading turntable 32 so that the filter assembly 40 located in the placement slot 321 can rotate.

[0101] Continue reading Fig.10 In the above-mentioned embodiment of the present application, further, the loading platform 30 also includes a first pipe joint 34 with openings at both ends, and the first pipe joint 34 is passed through the connection between the placement groove 321 and the receiving groove 322, and abuts against the edge of the waste liquid port 441, so that one end of the first pipe joint 34 is connected to the waste liquid port 441, and the other end is connected to the waste liquid pool or waste liquid tank through a soft rubber tube through the liquid outlet 331.

[0102] In this embodiment, the shape of the first pipe joint 34 matches that of the waste liquid port 441, so that when the filter assembly 40 is compressed, a sealing effect is formed at the first pipe joint 34 and the waste liquid port 441, allowing the waste liquid to enter the first pipe joint 34 from the waste liquid port 441, and then be discharged into the waste liquid pool or waste liquid tank through the soft rubber tube connected to the first pipe joint 34.

[0103] Furthermore, the mounting hole 311 is a stepped hole. The loading turntable 32 is sleeved with a first bearing 35, and the first bearing 35 is located in the stepped hole on a side away from the end cover 33. In this embodiment, since the first pipe joint 34 and the filter assembly 40 are in axial contact, in order to ensure the sealing effect, a certain axial force must exist. In this way, the axial force is borne by the setting of the first bearing 35. Specifically, the first bearing 35 uses a thrust cylindrical roller bearing, and the thrust cylindrical roller bearing bears the axial force, thereby reducing the friction generated by the rotation of the loading turntable 32 and the loading base 31.

[0104] It should be understood that the loading turntable 32 may also be provided with a second bearing 36, which is located in the stepped hole on one side close to the end cover 33. That is, bearings are provided at the rotation points of the upper and lower ends of the loading turntable 32 and the loading base 31, and the friction between the loading turntable 32 and the loading base 31 is reduced by providing the second bearing 36. In this embodiment, the second bearing 36 adopts a deep groove ball bearing, thereby reducing the friction generated by the rotation between the loading turntable 32 and the loading base 31. At the same time, the end cover 33 is connected to the loading turntable 32 to limit the position of the second bearing 36, and can rotate with the loading turntable 32.

[0105] It should be noted that, in this embodiment, the first bearing 35 uses a thrust cylindrical roller bearing, and the second bearing 36 uses a deep groove ball bearing. Of course, in other embodiments, the first bearing 35 and the second bearing 36 may also use other bearings.

[0106] like Figure 3 and Figure 4 As shown, in some embodiments of the present application, optionally, the injection device 50 includes an injection pump 51 , a telescopic mechanism 52 and a turntable assembly 54 .

[0107] Specifically, the infusion pump 51 has an infusion port 511 for sucking or discharging liquid. The telescopic mechanism 52 includes a driving member 521 and a moving member 522. The driving member 521 is fixedly connected to the infusion pump 51. Optionally, the driving member 521 and the infusion pump 51 can be fixed together on the slider 71. The output end of the driving member 521 is connected to the moving member 522 for driving the moving member 522 to move relative to the loading platform 30.

[0108] More specifically, the turntable assembly 54 is connected to the moving member 522, and the moving member 522 moves in a direction away from or close to the loading platform 30, thereby driving the turntable assembly 54 to move. The turntable assembly 54 includes a rotating mechanism 53, a turntable base 541, an injection turntable 542, a second pipe joint 543 and a mounting seat 544. The mounting seat 544 is connected to the moving member 522, and the rotating mechanism 53 and the turntable base 541 are respectively arranged on the mounting seat 544. The injection turntable 542 is rotatably connected to the turntable base 541 and is connected to the rotating end of the rotating mechanism 53. The second pipe connector 543 has a first end 5431 and a second end 5432 that are relatively arranged. The second pipe connector 543 is passed through the injection turntable 542, and the first end 5431 is connected to the infusion port 511 through a hose. The second end 5432 can abut against the edge of the liquid inlet 411 to connect the second pipe connector 543 with the liquid inlet 411.

[0109] In the present embodiment, the moving member 522 is driven by the driving member 521 to move in a direction away from or close to the loading platform 30, so as to drive the turntable assembly 54 to move in a direction away from or close to the loading platform 30, so that the turntable assembly 54 can be pressed against the loading platform 30 when the turntable assembly 54 moves downward a preset distance. At this time, the injection turntable 542 abuts against the filter assembly 40, pressing the filter assembly against the loading turntable 32, and also restricting the filter assembly 40. At this time, the injection turntable 542 is driven to rotate by the rotating mechanism 53, thereby causing the filter assembly 40 to rotate, and the filter assembly 40 drives the loading turntable 32 to rotate.

[0110] It should be noted that the driving member 521 of the telescopic mechanism 52 can be a hydraulic cylinder or a pneumatic cylinder, and the moving member 522 can be a push rod, which is easy to operate. Of course, in other embodiments, the telescopic mechanism 52 can also use a linear motor.

[0111] like Figure 3As shown, in the above embodiment of the present application, optionally, the rotating mechanism 53 includes a driving motor 531 and a transmission member 532 , the driving motor 531 drives the transmission member 532 to rotate, and the transmission member 532 is drivingly connected to the injection turntable 542 .

[0112] Specifically, the driving motor 531 is fixed on the mounting seat 544 , and the turntable base 541 is also fixed on the mounting seat 544 .

[0113] Optionally, the liquid injection device further comprises an L-shaped bracket 57, a mounting seat 544 is mounted on a horizontal plate of the L-shaped bracket 57, the moving member 522 is also mounted on the horizontal plate of the L-shaped bracket 57, and the vertical plate of the L-shaped bracket 57 is parallel to the telescopic direction of the moving member 522. At the same time, the driving motor 531 and the turntable base 541 can be connected to the mounting seat 544 by bolts.

[0114] Optionally, the transmission member 532 is a first gear connected to the output end of the drive motor 531, and a second gear 5421 meshing with the first gear is provided on the injection turntable 542. The drive motor 531 drives the first gear to rotate, thereby driving the second gear 5421 to rotate, thereby driving the injection turntable 542 to rotate.

[0115] Of course, the transmission member 532 can also use a belt transmission mechanism, such as the transmission member 532 includes a first pulley and a belt, the first pulley is connected to the output end of the driving motor 531, the injection turntable 542 is provided with a second pulley, and the belt transmission is connected to the first pulley and the second pulley. The driving motor 531 drives the first pulley to rotate, and drives the second pulley to rotate through the belt, thereby driving the injection turntable 542 to rotate.

[0116] It should be noted that the transmission member 532 is not limited to the above structure, and the transmission member 532 can also use a sprocket chain structure, that is, the transmission member 532 includes a chain and a first sprocket, the output end of the drive motor 531 is connected to the first sprocket, and a second sprocket is provided on the injection turntable 542, and the chain is connected to the first sprocket and the second sprocket. When in use, the drive motor 531 drives the first sprocket to rotate, and the second sprocket is driven to rotate through the chain, thereby driving the injection turntable 542 to rotate.

[0117] It should be understood that in other embodiments, the transmission member 532 may also be driven by a planetary gear or a worm gear mechanism.

[0118] like Figure 4As shown, in the above-mentioned embodiment of the present application, the turntable base 541 is provided with a step hole 5411, and the injection turntable 542 is provided with a third bearing 55, and the third bearing 55 is located in the step hole 5411 on the side close to the moving part 522; and / or, the injection turntable is also provided with a fourth bearing 56, and the fourth bearing 56 is located in the step hole 5411 on the side away from the moving part 522.

[0119] In this embodiment, since the injection turntable 542 and the turntable base 541 rotate relative to each other, in order to make the injection turntable 542 and the turntable base 541 rotate more smoothly and reduce radial clearance shaking, the third bearing 55 is provided to achieve a rotational fixed relationship between the injection turntable 542 and the turntable base 541, and the third bearing 55 is used to achieve a rotation function of reducing friction. In this embodiment, the third bearing 55 uses a deep groove ball bearing, thereby reducing the friction generated by the rotation of the injection turntable 542 and the turntable base 541.

[0120] It should be understood that the loading turntable may also be provided with a fourth bearing 56. Since the second pipe joint 543 and the filter assembly 40 are in axial contact, a certain axial force must exist in order to ensure the sealing effect. In this way, the axial force is borne by the fourth bearing 56. Specifically, the fourth bearing 56 uses a thrust cylindrical roller bearing, and the axial force is borne by the thrust cylindrical roller bearing, thereby reducing the axial force between the injection turntable 542 and the turntable base 541, thereby reducing the friction generated by the rotation between the two.

[0121] It should be noted that the third bearing 55 and the fourth bearing 56 in this embodiment, wherein the third bearing 55 located at the upper end of the step hole 5411 is selected to use a deep groove ball bearing, and the fourth bearing 56 located at the lower end of the step hole 5411 is selected to use a thrust cylindrical roller bearing.

[0122] Of course, in other embodiments, the injection turntable 542 may be directly rotatably connected to the turntable base 541 via a third bearing 55 .

[0123] like Figures 5 to 7 As shown, in some embodiments of the present application, the end surface of the second end 5432 is provided with an annular groove 5433, and the annular groove 5433 is used to engage with the probe gun head 200. In this way, when the telescopic mechanism 52 drives the turntable assembly 54 to move downward, the probe gun head 200 can be engaged through the annular groove 5433, thereby driving the probe gun head 200 to move to the test tube 300 or to the liquid inlet 411 of the filter assembly 40 to output liquid or absorb liquid.

[0124] See also Figure 6Furthermore, the second pipe joint 543 is provided with an air vent 5434 in communication with the annular groove 5433. Thus, when the probe gun tip 200 at the annular groove 5433 is scraped off by the gun tip scraper 82, the air entering from the air vent 5434 can generate a certain atmospheric pressure on the probe gun tip 200 at the annular groove 5433, so that the probe gun tip 200 is easily scraped off.

[0125] like Figure 1 and Figure 2 As shown, in some embodiments of the present application, optionally, the exosome extraction device 100 further includes a lateral movement mechanism 70, the lateral movement mechanism 70 is disposed on the rotating member 20, the injection pump 51 and the driving member 521 are disposed on the lateral movement mechanism 70 through a slider 71, and the lateral movement mechanism 70 is used to drive the slider 71 to move along the length direction of the base 10. In this way, the injection device 50 can move along the length direction of the base 10, that is, Figure 1 The left and right directions of movement make it easier for the injection device 50 to reach different process positions for operation.

[0126] Specifically, the lateral moving mechanism 70 includes a bracket 72 , a driving part 73 and a transmission part 74 .

[0127] The bracket 72 is installed on the rotating member 20, the driving part 73 and the transmission part 74 are respectively installed on the bracket 72, and the driving part 73 is used to drive the transmission part 74 to rotate, the slider 71 is arranged on the transmission part 74, and the driving part 73 drives the transmission part 74 to rotate, so that the slider 71 moves relative to the bracket 72, thereby driving the injection device 50 to move.

[0128] It should be noted that, in this embodiment, the driving part 73 is a motor, the transmission part 74 is a screw, the slider 71 is sleeved on the screw, and the motor drives the screw to rotate to drive the slider 71. Of course, in other embodiments, the transmission part 74 can also be a belt transmission mechanism or a chain transmission mechanism to move the slider 71.

[0129] like Fig.11 and Fig.12 As shown, in some embodiments of the present application, the filter assembly 40 includes an upper cover 41 , the filter membrane 42 , filter paper 43 and a lower cover 44 .

[0130] Specifically, the upper cover 41 is provided with the liquid inlet 411, the lower cover 44 is provided with a groove 442, the filter membrane 42 and the filter paper 43 are respectively located in the groove 442, and the filter paper 43 is located between the filter membrane 42 and the bottom of the groove 442, the lower cover 44 is provided with the waste liquid port 441, the waste liquid port 441 is communicated with the groove 442, and the upper cover 41 is connected to the lower cover 44.

[0131] In this embodiment, specifically, the filter paper 43 is placed in the groove 442, and the filter membrane 42 is plastic welded or bonded to the groove 442 of the lower cover 44, the filter membrane 42 is pressed by the upper cover 41, and then the upper cover 41 and the lower cover 44 are sealed into a whole by ultrasonic welding or glue bonding. The function of the filter paper 43 is to support the filter membrane 42 and to accelerate the absorption speed of the filtered liquid by using its good water absorption.

[0132] It should be noted that, preferably, the pore size of the filter membrane 42 is 0.03 μm to 0.20 μm (eg, 0.03 μm, 0.05 μm, 0.08 μm, 0.10 μm, 0.13 μm, 0.16 μm or 0.20 μm).

[0133] In addition, since the purpose of designing this filter assembly 40 is to make it a disposable consumable, it can be mass-produced to reduce manual installation operations, and all parts need to be assembled before leaving the factory. In order to ensure airtightness and reliability, all parts need to be firmly fixed, and ultrasonic welding or other glue bonding methods can be used. In this embodiment, ultrasonic welding technology is used, so the materials of the upper cover 41 and the lower cover 44 are both polycarbonate (PC) or acrylic, which is a plastic transparent material that is easy to fuse with the filter membrane 42 (PC material), so as to facilitate observation of the experimental process.

[0134] like Fig.12 As shown, further, a filtering cavity 45 is formed between the upper cover 41 and the filtering membrane 42. In this way, the filtering cavity 45 can be used to temporarily store the liquid sample before / after filtering, so as to facilitate the aspiration of the probe gun head 200.

[0135] Furthermore, the upper cover 41 and the lower cover 44 are both circular, the waste liquid port 441 is located at a non-center position on the lower cover 44, and the liquid inlet 411 is located at a non-center position on the upper cover 41. In this way, the filter assembly 40 and the loading platform 30 are relatively fixed by utilizing the eccentric circle feature, which is convenient for driving the loading turntable 32 to rotate, and the shape of the waste liquid port 441 matches the shape of the first pipe joint 34, and a sealing surface can be formed during operation.

[0136] In addition, the liquid inlet 411 and the waste liquid port 441 are staggered, with the liquid inlet 411 being arranged close to the edge of the upper cover 41, and the waste liquid port 441 being arranged on the lower cover 44 at a position away from the liquid inlet 411, so that the distance between the liquid inlet 411 and the waste liquid port 441 becomes larger. In this way, the distance over which the liquid flows is extended as much as possible to avoid blockage due to the close distance between the two.

[0137] It should be noted that, in other embodiments, Fig.13 and Fig.14 As shown, the liquid inlet 411 and the waste liquid outlet 441 can also be coaxially arranged. Of course, the two can be arranged at the center position of the circular upper cover 41 and the lower cover 44, or at the non-center position of the circular upper cover 41 and the lower cover 44.

[0138] like Figure 1 and Fig.15 As shown, in some embodiments of the present application, the exosome extraction device 100 further includes a sample consumable rack 80, and the sample consumable rack 80 is provided with at least one first mounting position 801 and at least one second mounting position 802, wherein the first mounting position 801 is used to place the test tube 300, and the second mounting position 802 is used to place the probe gun tip 200. In this way, more first mounting positions 801 can be provided to place test tubes 300 with different reagents, and more second mounting positions 802 can be provided to place more probe gun tips 200, so as to facilitate taking and placing.

[0139] In this embodiment, the number of first mounting positions 801 is four and the number of second mounting positions 802 is four as an example. Of course, in other embodiments, one, two, five, etc. can also be used.

[0140] Furthermore, a waste box 81 with an opening is also provided on the sample consumables rack 80 , a gun tip scraper 82 is provided above the waste box 81 , and a notch 821 matching the probe gun tip 200 is provided on the gun tip scraper 82 .

[0141] In this embodiment, the tip scraper 82 is provided to facilitate the injection device 50 to move to the position and use the tip scraper 82 to scrape off the used probe gun tip 200. At the same time, the scraped probe gun tip 200 falls into the waste box 81, which is used to temporarily store the discarded probe gun tip 200.

[0142] like Fig. 22 As shown, the embodiment of the present application also provides a method for extracting exosomes, using the exosome extraction device 100 in the above embodiment, and the specific operation steps are:

[0143] Before operating, Fig.17As shown, in the initial state of the exosome extraction device 100, the sample to be tested, reagents and consumables (probe gun tip 200, filter assembly 40) are placed;

[0144] Step S1, such as Fig.18 As shown, the injection device 50 extracts a probe gun tip 200 on the sample consumables rack through the second pipe joint 543, and then moves to the wetting reagent tube through the lateral moving mechanism 70 to take the wetting agent and temporarily store it in the injection pump 51, and after the wetting agent is absorbed, it moves to the gun tip scraper 82 to scrape off the probe gun tip 200;

[0145] Step S2, then move the injection turntable 542 to the filter assembly 40 through the telescopic mechanism 52 and press and seal it, inject the absorbed wetting agent into the filter assembly 40 to wet the filter paper 43 in the filter assembly 40;

[0146] Step S3, the injection device 50 extracts another probe gun tip 200 on the sample consumable rack again through the second pipe joint 543, then moves to the blood sample test tube to take the blood sample and temporarily stores it in the injection pump 51, and after the absorption, moves to the gun tip scraper 82 to scrape off the probe gun tip 200;

[0147] Step S4, such as Fig.19 As shown, the injection turntable 542 is then moved to the filter assembly 40 through the telescopic mechanism 52 and pressed and sealed. The injection device 50 and the loading platform 30 are rotated 90° counterclockwise through the rotating member 20. At this time, the second pipe joint 543 on the injection turntable 542 is in the lower position, and the first pipe joint 34 on the loading platform 30 is in the upper position. The blood sample is injected into the filter assembly 40 at a constant speed for filtration. During the filtration process, the vibration device 60 is turned on, such as using an ultrasonic transducer to emit ultrasonic waves toward the filter assembly 40. During this process, the rotating mechanism 53 rotates to drive the injection turntable 542 to rotate, thereby driving the filter assembly 40 to rotate. It should be noted that the filter assembly 40 can be controlled to rotate at any time or at a preset time. When the filtration is completed, the injection device 50 and the loading platform 30 are rotated 90° clockwise through the rotating member 20 to reset them to a vertical state;

[0148] Step S5, the injection device 50 extracts another probe gun tip 200 on the sample consumable rack through the second pipe joint 543, then moves to the cleaning agent tube to take the cleaning agent and temporarily stores it in the injection pump 51, and after the absorption, moves to the gun tip scraper 82 to scrape off the probe gun tip 200;

[0149] Step S6, as Fig. 20As shown, the injection turntable 542 then moves to the filter assembly 40 and is pressed and sealed. The injection device 50 and the loading platform 30 rotate counterclockwise by 90° through the rotating member 20. At the same time, the rotating mechanism 53 of the injection device 50 drives the filter assembly 40 on the loading platform 30 to rotate together, so that the second pipe joint 543 on the injection turntable 542 is in the upper position and the first pipe joint 34 on the loading platform 30 is in the lower position. The cleaning agent is injected into the filter assembly 40 at a constant speed for filtration. During the filtration process, the vibration device 60 starts to work, such as using an ultrasonic transducer to emit ultrasonic waves toward the filter assembly 40. In this process, the rotating mechanism 53 rotates, driving the injection turntable 542 to rotate, thereby driving the filter assembly 40 to rotate. It should be noted that the filter assembly 40 can be controlled to rotate at any time or at a preset time. When the filtration is completed, the injection device 50 and the loading platform 30 are rotated clockwise by 90° through the rotating member 20 to reset them to a vertical state;

[0150] Step S7, Fig.21 As shown, the injection device 50 extracts another probe gun head 200 on the sample consumable rack through the second pipe joint 543, and then moves the probe gun head 200 to the liquid inlet 411 on the filter assembly 40 to absorb the filtered sample. Since the amount of sample obtained after filtration is very small, the absorbed sample is stored in the probe gun head 200 and will not flow into the injection pump 51 and its pipeline.

[0151] Step S8, then inject the obtained sample into the exosome collection reagent tube to complete the filtration experiment.

[0152] It should be noted that during the sampling process, the vibration device 60 also continues to work to ensure that the target is suspended in the liquid and is easier to be absorbed.

[0153] Before the operation of step S3 in the above embodiment, the step of pre-treating the blood sample in the blood sample tube to be placed on the sample consumable rack is also included. The pre-treatment includes centrifuging, diluting and filtering the blood sample in sequence.

[0154] Preferably, the pore size of the filter used for filtering is 0.15 μm to 0.22 μm (eg, 0.15 μm, 0.16 μm, 0.18 μm, 0.20 μm, 0.21 μm or 0.22 μm).

[0155] The diluted extract was preliminarily filtered using a syringe filter to further remove large-diameter microvesicles and possible apoptotic bodies, thereby obtaining a sample to be filtered with relatively simple components.

[0156] The pretreatment step can reduce the risk of various impurities such as cell debris and proteins being accumulated on the filter membrane during the ultrafiltration process, and reduce the possibility of clogging of the filter membrane 42.

[0157] Preferably, the pretreatment further comprises enzymatic hydrolysis of the blood sample.

[0158] Preferably, the centrifugal force of the centrifugation is 9000g to 12000g (for example, 9000g, 9500g, 10000g, 10500g, 11000g, 11500g or 12000g).

[0159] Preferably, the centrifugation time is 15 min to 20 min (eg, 15 min, 16 min, 17 min, 18 min, 19 min or 20 min).

[0160] The blood samples were centrifuged to remove cell debris and large cell microvesicles.

[0161] Preferably, the temperature of the enzymatic hydrolysis is 36-40°C (eg, 36°C, 37°C, 38°C or 40°C).

[0162] Preferably, the enzymatic hydrolysis time is 10 min to 30 min (e.g., 10 min, 12 min, 15 min, 20 min, 21 min, 25 min, 28 min or 30 min).

[0163] Preferably, the enzymatic enzyme comprises at least one of proteinase K, pronase, trypsin, pepsin, elastase, thrombin or chymotrypsin.

[0164] The extract is subjected to enzymatic hydrolysis to remove impure proteins therein.

[0165] Preferably, the vibration frequency of the vibration device is 20 Hz to 100 kHz (for example, 20 Hz, 25 Hz, 30 Hz, 40 Hz, 80 Hz, 100 Hz, 120 Hz, 200 Hz, 400 Hz, 700 Hz, 900 Hz, 1 kHz, 5 kHz, 10 kHz, 20 kHz, 30 kHz, 33 kHz, 35 kHz, 38 kHz, 40 kHz, 43 kHz, 45 kHz, 48 kHz, 50 kHz, 70 kHz, 80 kHz, 100 kHz).

[0166] In summary, by using the exosome extraction device 100 provided in this embodiment, the exosomes can be automatically extracted, effectively avoiding the uncertainty caused by manual operation. In addition, the ultrasonic wave emitted by the vibration device 60 vibrates the liquid, particles and filter membrane 42 in the filter assembly 40, preventing the particles from resting on the filter membrane 42, preventing the filter membrane 42 from being blocked, and improving the extraction efficiency of exosomes in the liquid.

[0167] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" etc. means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present application. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art may combine and combine the different embodiments or examples described in this specification and the features of the different embodiments or examples, without contradiction.

[0168] Although the embodiments of the present application have been shown and described above, it can be understood that the above embodiments are exemplary and cannot be understood as limitations on the present application. Ordinary technicians in this field can change, modify, replace and modify the above embodiments within the scope of the present application.

Claims

1. An exosome extraction device, characterized in that: include: Base; A loading platform, wherein the loading platform is provided with a placement groove and a liquid outlet communicated with the placement groove; The filter assembly is detachably mounted in the placement groove, and the filter assembly comprises an upper cover, a filter membrane, filter paper, a lower cover, and a liquid inlet and a waste liquid outlet located on two opposite sides of the filter membrane, the waste liquid outlet being communicated with the liquid outlet; the upper cover is provided with the liquid inlet, the lower cover is provided with a groove, the filter membrane and the filter paper are respectively located in the groove, and the filter paper is located between the filter membrane and the bottom of the groove, the lower cover is provided with the waste liquid outlet, the waste liquid outlet is communicated with the groove, and the upper cover is connected with the lower cover; the upper cover and the lower cover are both circular, the waste liquid outlet is located at a non-center position on the lower cover, the liquid inlet is located at a non-center position on the upper cover, the liquid inlet and the waste liquid outlet are staggered, the liquid inlet is arranged close to the edge of the upper cover, and the waste liquid outlet is arranged on the lower cover at a position away from the liquid inlet; A liquid injection device, the liquid injection device is used to output liquid into the filter assembly or absorb liquid; a vibration device, disposed near the filter assembly, for vibrating the liquid, particles and the filter membrane in the filter assembly; A rotating member is rotatably disposed on the base, the vibration device is disposed on the rotating member, and the rotating member drives the vibration device and the loading platform to rotate a preset angle in a vertical plane.

2. The exosome extraction device according to claim 1, characterized in that: The loading platform comprises: A loading base, one end of which is connected to the rotating member, and the other end of which is provided with a mounting hole; A loading turntable, the loading turntable is rotatably arranged at the mounting hole, the placement slot is arranged on the loading turntable, and a receiving slot communicating with the placement slot is arranged on a side away from the placement slot; An end cover is connected to a side of the loading turntable where the receiving groove is provided, and covers the receiving groove, and the end cover is provided with the liquid outlet.

3. The exosome extraction device according to claim 2, characterized in that: The loading platform also includes a first pipe joint with openings at both ends. The first pipe joint is penetrated at the connection between the placement groove and the containing groove, and abuts against the edge of the waste liquid port to connect the first pipe joint with the waste liquid port.

4. The exosome extraction device according to claim 2, characterized in that: The mounting hole is a stepped hole; The loading turntable is sleeved with a first bearing, which is located in the stepped hole on a side away from the end cover; and / or, the loading turntable is also sleeved with a second bearing, which is located in the stepped hole on a side close to the end cover.

5. The exosome extraction device according to claim 2, characterized in that: The liquid injection device comprises: An infusion pump having an infusion port for sucking in or delivering liquid; The telescopic mechanism comprises a driving member and a moving member, wherein the driving member is fixedly connected to the injection pump, and an output end of the driving member is connected to the moving member, and is used to drive the moving member to move relative to the loading platform; A turntable assembly connected to the moving part and moving with the moving part; Wherein, the turntable assembly includes a rotating mechanism, a turntable base, an injection turntable, a mounting seat and a second pipe joint, the mounting seat is connected to the moving part, and the rotating mechanism and the turntable base are respectively arranged on the mounting seat; the injection turntable is rotatably connected to the turntable base and is connected to the rotating end of the rotating mechanism; the second pipe joint has a first end and a second end that are relatively arranged, the second pipe joint is passed through the injection turntable, and the first end is connected to the infusion port through a hose, and the second end can abut against the edge of the liquid inlet to connect the second pipe joint with the liquid inlet.

6. The exosome extraction device according to claim 5, characterized in that: The rotating mechanism comprises a driving motor and a transmission member, wherein the driving motor drives the transmission member to rotate, and the transmission member is in driving connection with the injection turntable; The driving motor and the turntable base are respectively fixedly connected to the mounting base.

7. The exosome extraction device according to claim 6, characterized in that: The transmission member is a first gear, the first gear is connected to the output end of the drive motor, and the injection turntable is provided with a second gear meshing with the first gear; or The transmission member includes a first pulley and a belt, the first pulley is connected to the output end of the driving motor, a second pulley is arranged on the injection turntable, and the belt transmission is connected to the first pulley and the second pulley.

8. The exosome extraction device according to claim 5, characterized in that: The turntable base is provided with a step hole, and the injection turntable is provided with a third bearing, and the third bearing is located on the side of the step hole close to the moving part; and / or, the injection turntable is also provided with a fourth bearing, and the fourth bearing is located on the side of the step hole away from the moving part.

9. The exosome extraction device according to claim 5, characterized in that: An annular groove is formed on the end surface of the second end, and the annular groove is used for clamping with the probe gun head.

10. The exosome extraction device according to claim 9, characterized in that: The second pipe joint is provided with a vent hole communicated with the annular groove.

11. The exosome extraction device according to any one of claims 6 to 10, characterized in that: The exosome extraction device also includes a transverse moving mechanism, which is arranged on the rotating member. The injection pump and the driving member are arranged on the transverse moving mechanism through a slider, and the transverse moving mechanism is used to drive the slider to move along the length direction of the base.

12. The exosome extraction device according to claim 11, characterized in that: The lateral movement mechanism includes a bracket, a driving part and a transmission part. The bracket is installed on the rotating member, the driving part and the transmission part are installed on the bracket respectively, and the driving part is used to drive the transmission part to rotate. The slider is arranged on the transmission part.

13. The exosome extraction device according to claim 11, characterized in that: The exosome extraction device also includes a sample consumables rack, on which at least one first mounting position and at least one second mounting position are provided, wherein the first mounting position is used to place a test tube, and the second mounting position is used to place a probe gun tip.

14. The exosome extraction device according to claim 13, characterized in that: The sample consumables rack is also provided with a waste box with an opening, a gun head scraper is provided above the waste box, and a notch matching the probe gun head is opened on the gun head scraper.

15. The exosome extraction device according to claim 1, characterized in that: A filtering cavity is formed between the upper cover and the filtering membrane.

16. The exosome extraction device according to any one of claims 1 to 10, characterized in that: The vibration device is any one of an ultrasonic instrument, an ultrasonic transducer, a vortex vibrator, a high-frequency vibration motor, a pneumatic vibrator or an exciter.

17. A method for extracting exosomes, characterized in that: The use of the exosome extraction device according to claim 14 comprises the following steps: The injection device extracts a probe gun tip on the sample consumables rack through the second pipe joint, moves to the wetting reagent tube through the lateral moving mechanism to take the wetting agent and temporarily stores it in the injection pump, and after sucking the wetting agent, moves to the gun tip scraper to scrape off the probe gun tip; The injection turntable is moved onto the filter assembly through the telescopic mechanism and pressed and sealed, and the absorbed wetting agent is injected into the filter assembly and the filter paper is wetted; The injection device extracts another probe gun tip on the sample consumable rack through the second pipe joint, and moves to the blood sample test tube through the lateral moving mechanism to take the blood sample and temporarily store it in the injection pump, and after sucking the blood sample, moves to the gun tip scraper to scrape off the probe gun tip; The injection turntable is moved to the filter assembly through the telescopic mechanism and pressed and sealed. The injection device and the loading platform are rotated 90° counterclockwise through the rotating member, so that the second pipe joint on the injection turntable is in the lower position and the first pipe joint on the loading platform is in the upper position. The injection device injects the blood sample into the filter assembly at a constant speed for filtration. During the filtration process, the vibration device starts to work; The injection device extracts another probe gun tip on the sample consumables rack through the second pipe joint, and moves to the cleaning agent tube through the lateral moving mechanism to take the cleaning agent and temporarily store it in the injection pump, and after sucking the cleaning agent, moves to the gun tip scraper to scrape off the probe gun tip; The injection turntable is moved to the filter assembly through the telescopic mechanism and pressed and sealed. The injection device and the loading platform are rotated 90° counterclockwise through the rotating parts. At the same time, the rotating mechanism of the injection device drives the filter assembly on the loading platform to rotate together, so that the second pipe joint on the injection turntable is in the lower position and the first pipe joint on the loading platform is in the upper position. The injection device injects the cleaning agent into the filter assembly at a constant speed for filtration. During the filtration process, the vibration device starts to work; The liquid injection device extracts another probe gun head on the sample consumables rack through the second pipe joint, and moves the probe gun head to the liquid inlet on the filter assembly to absorb the filtered sample; The obtained sample was injected into the exosome collection reagent tube to complete the filtration experiment.

18. The extraction method according to claim 17, characterized in that The following steps are also included: Pre-treating the blood sample in the blood sample tube to be placed on the sample consumables rack; The pretreatment includes: centrifuging, diluting and filtering the blood sample.

19. The extraction method according to claim 18, characterized in that The pretreatment further comprises enzymatic hydrolysis of the blood sample.

20. The extraction method according to claim 18, characterized in that The pore size of the filtering filter is 0.15 μm to 0.22 μm.

21. The extraction method according to claim 18, characterized in that The centrifugal force of the centrifugation is 9000g to 12000g; the centrifugal time is 15min to 20min.

22. The extraction method according to claim 19, characterized in that The temperature of the enzymolysis is 36° C. to 40° C.; the time of the enzymolysis is 10 min to 30 min.

23. The extraction method according to any one of claims 17 to 22, characterized in that The vibration frequency of the vibration device is 20 Hz to 100 kHz.

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