A rapid processing device for plasma samples for ctDNA methylation detection

Abstract

The utility model discloses a kind of plasma sample rapid processing device for ctDNA methylation detection, by setting blood collection cavity plate and bottom liftable blood collection tube base in the top of blood separation box after centrifugation, multiple blood collection tubes are uniformly detachably arranged on base, multiple groups of evenly distributed and synchronous operation electric telescopic link are connected between blood collection cavity plate and blood collection tube base, each blood collection tube top is communicated to blood collection bin, and the plasma collected is transferred to blood storage bin by connecting pipe, transmission pipe with miniature pump body and sleeve pipe.This device realizes the precise collection of the upper plasma after centrifugation in multiple areas synchronously pumped, avoids the pollution to white blood cell layer and improves the collection speed and efficiency, and can realize multi-way synchronization, abnormal self-checking and safety interlocking.

Description

Technical Field

[0001] This utility model relates to the technical field, specifically to a rapid plasma sample processing device for ctDNA methylation detection. Background Technology

[0002] The rapid plasma sample processing device for ctDNA methylation detection mainly consists of five core modules; (1) Sample input and recognition module Input interface: Supports direct loading of standard blood collection tubes (such as EDTA-K2 tubes and cfDNA-specific Streck tubes) or loading of pre-separated plasma transfer pipette tips.

[0003] Intelligent identification system: Built-in barcode / RFID scanner to automatically identify and associate sample information.

[0004] Temperature-controlled zone: While samples are awaiting processing, they are kept in a 4°C environment to prevent nucleic acid degradation.

[0005] (2) High-speed centrifugation and plasma separation module Centrifuge: If whole blood samples are accepted, this module can complete blood stratification within minutes.

[0006] Plasma aspiration unit: aspirates the upper layer of plasma.

[0007] (3) Rapid ctDNA extraction and purification module Magnetic bead purification system: It is easy to automate, fast, and has high purity.

[0008] Reagent compartment: Built-in refrigeration area (4-8°C) for storing critical reagents such as lysis buffer, washing buffer, and elution buffer.

[0009] Magnetic rod / magnetic sleeve system: Capture and transfer magnetic bead-ctDNA complexes by moving a magnetic rod (or moving a magnetic base).

[0010] Efficient lysis and binding: The lysis chamber is rapidly heated to a specified temperature (e.g., 55°C) to accelerate the lysis of cell debris and the release of ctDNA, which then binds efficiently to functionalized magnetic beads.

[0011] Rapid washing: Through the rapid suspension and adsorption of magnetic beads, 2-3 washes are completed within minutes to remove impurities such as proteins and salt ions.

[0012] Low-volume elution: Use small volumes (e.g., 20-50 μL) of ultrapure water or low-concentration Tris-EDTA buffer for elution to ensure a high concentration of ctDNA.

[0013] (4) Bisulfite conversion pretreatment module Integrated reaction chamber: ctDNA methylation detection usually requires bisulfite treatment to convert unmethylated cytosine into uracil, while methylated cytosine remains unchanged.

[0014] Rapid conversion technology: The integrated rapid bisulfite conversion kit protocol, based on high temperature and specific pH control, can reduce the processing time from 3-4 hours to 60-90 minutes.

[0015] Purification Unit: The transformed DNA contains a miniaturized purification unit for rapid removal of bisulfite reagent and to obtain purified DNA that can be used for downstream detection (such as qPCR, digital PCR, sequencing).

[0016] (5) Control system and user interface Central Processing Unit: Controls all mechanical movements, temperature control, fluid flow, and timing.

[0017] Touchscreen interface: Provides an intuitive user interface, allowing users to select programs, monitor progress, and view processing logs.

[0018] Data output: Automatically generates processing reports, including sample ID, processing time, ctDNA concentration, etc.

[0019] The general workflow of this device is as follows: Sample loading: The operator places the plasma sample (or whole blood sample) on the sample rack, scans the barcode, and closes the hatch; Startup: Select the "ctDNA Methylation Rapid Extraction and Transformation" program on the touchscreen and click Run.

[0020] Automatic operation (fully enclosed): Step 1: High-speed centrifugation to separate plasma; Step 2: Precisely pipette a specific volume of plasma into the lysis chamber; Step 3: Add lysis buffer, incubate, release ctDNA and bind to magnetic beads; Step 4: Magnetic bead capture, multiple washes, and drying; Step 5: Elute to obtain preliminarily purified ctDNA; Step 6: The eluent is automatically transferred to the BS transformation chamber. Add BS transformation reagent and perform a rapid transformation reaction under precise temperature control; Step 7: The transformed sample undergoes rapid purification to remove the BS reagent; Step 8: Final elution to obtain BS-treated ctDNA suitable for downstream analysis; Output: Transfer the microcentrifuge tubes or 96-well plates containing the final product to the output area and prompt the user to pick them up.

[0021] In the plasma aspiration unit of the high-speed centrifugation and plasma separation module, simple pipettes are often used for aspiration. This makes it difficult to accurately control the distance between the bottom of the pipette and the lower layer of white blood cells, which can easily contaminate the white blood cell layer. At the same time, it is not possible to fully aspirate the upper plasma. Moreover, aspiration can only be performed on local areas, which is slow and inefficient. To address this, this technical solution designs a rapid plasma sample processing device for ctDNA methylation detection. Utility Model Content

[0022] The purpose of this invention is to provide a rapid plasma sample processing device for ctDNA methylation detection, so as to solve the problems mentioned in the background art.

[0023] To achieve the above objectives, this utility model provides the following technical solution: A rapid plasma sample processing device for ctDNA methylation detection includes a blood separation box, which stores centrifuged blood, i.e., upper plasma and lower white blood cells. A blood collection chamber plate is snap-fitted onto the top of the blood collection chamber plate, and a blood collection tube base is lifted and lowered at the bottom of the blood collection chamber plate. The blood collection tube base is used for multi-region synchronous pumping and precise collection of the upper plasma inside the blood separation box. The bottom of the blood collection chamber plate and the top of the blood collection tube base are connected by multiple sets of evenly distributed electric telescopic rods. The multiple sets of electric telescopic rods are initially in the same state and operate synchronously to maintain high-precision and stable lifting and lowering of the blood collection tube base. Multiple blood collection tubes are detachably and evenly installed at the bottom of the blood collection tube base. As the blood collection tube base is raised and lowered, the absorption port at the bottom of the blood collection tube extends into the plasma and is placed above the white blood cells. At the same time, the top of each blood collection tube is connected to a blood collection chamber located inside the blood collection tube base. The top of the blood collection chamber is connected to a transmission pipe installed on the outside of the top of the blood collection tube base via a connecting pipe. A micro pump is installed on the connecting pipe to apply an upward suction force to the blood collection tube at the bottom, that is, to control the blood collection tube to collect plasma from the corresponding area. The transfer tube is sealed with a sleeve on the outside. The top of the sleeve is connected to the middle of the bottom of the blood collection chamber plate. At the same time, the sleeve corresponding to the top of the transfer tube has a through structure that connects non-contactly to a blood storage chamber opened inside the blood collection chamber plate. When the electric telescopic rod controls the blood collection tube base to move up and down, the transfer tube slides synchronously along the inside of the sleeve. With the pumping action of the micro pump, the blood collection tube at the bottom is controlled to collect plasma from the corresponding location simultaneously. Then, the plasma is transferred to the blood storage chamber through the blood collection chamber, connecting tube, transfer tube, and sleeve. This enables the simultaneous collection of plasma from multiple locations, increasing the collection speed while ensuring adequate protection of white blood cells.

[0024] Compared with the prior art, the beneficial effects of this utility model are: by using multiple synchronously controllable blood collection tubes and micro pumps, multiple plasma collection sites can be collected in parallel at the same time, which significantly improves the collection speed and throughput.

[0025] By controlling the extension of the blood collection tube to the plasma layer and placing it above the leukocyte layer, the risk of disturbance to the leukocyte layer and nucleic acid contamination can be reduced, thereby improving the purity of downstream ctDNA.

[0026] The blood collection tubes and blood collection chamber are connected by a detachable male / female valve, which facilitates replacement and maintenance while ensuring closed-loop transfer and reducing cross-contamination. Attached Figure Description

[0027] Figure 1 This is a partial three-dimensional structural diagram of the blood separation box in a rapid plasma sample processing device for ctDNA methylation detection.

[0028] Figure 2 This is a partial front view schematic diagram of the blood separation box in a rapid plasma sample processing device for ctDNA methylation detection.

[0029] Figure 3 This is a three-dimensional structural diagram of the blood collection tube base in a rapid plasma sample processing device for ctDNA methylation detection.

[0030] Figure 4 This is a front view schematic diagram of the internal structure of the blood collection tube base in a rapid plasma sample processing device for ctDNA methylation detection.

[0031] Figure 5 for Figure 2 A magnified structural diagram of A in the middle.

[0032] Figure 6 for Figure 4 A magnified structural diagram of B in the diagram.

[0033] Figure 7 for Figure 4 A magnified structural diagram of C.

[0034] The system includes a blood separation box 10, a blood collection chamber plate 11, a clamping plate 12, an internal clamping plate 13, a blood collection tube base 14, a blood collection tube 15, a male connecting valve 16, a female connecting valve 17, a blood collection chamber 19, a micro pump body 20, a transmission tube 21, a sleeve 22, an electric telescopic rod 23, a blood storage chamber 24, and a control panel 25. Detailed Implementation

[0035] It should be noted that, unless otherwise specified, the embodiments and features described in these embodiments of the present invention can be combined with each other.

[0036] In the description of this utility model, it should be understood that the terms "center," "longitudinal," "lateral," "upper," "lower," "front," "rear," "left," "right," "vertical," "horizontal," "top," "bottom," "inner," and "outer," etc., indicating orientation or positional relationships based on the orientation or positional relationships shown in the accompanying drawings, are only for the convenience of describing this utility model and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation, and therefore should not be construed as a limitation of this utility model. Furthermore, the terms "first," "second," etc., are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of indicated technical features. Thus, features defined with "first," "second," etc., may explicitly or implicitly include one or more of that feature. In the description of this utility model, unless otherwise stated, "a plurality of" means two or more.

[0037] In the description of this utility model, it should be noted that, unless otherwise explicitly specified and limited, the terms "installation," "connection," and "joining" should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral connection; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; and they can refer to the internal connection of two components. Those skilled in the art can understand the specific meaning of the above terms in this utility model based on the specific circumstances.

[0038] The present invention will now be described in detail with reference to the accompanying drawings and embodiments.

[0039] Please see Figures 1-4 , Figure 6 A rapid plasma sample processing device for ctDNA methylation detection includes a blood separation box 10, which stores centrifuged blood, i.e., upper plasma and lower white blood cells. A blood collection chamber plate 11 is snap-fitted onto the top and a blood collection tube base 14 is lifted and lowered at the bottom of the blood collection chamber plate 11. The blood collection tube base 14 is used for multi-region synchronous pumping and precise collection of the upper plasma inside the blood separation box 10. The bottom of the blood collection chamber plate 11 and the top of the blood collection tube base 14 are connected by multiple sets of evenly distributed electric telescopic rods 23. The multiple sets of electric telescopic rods 23 are initially in the same state and operate synchronously to maintain the high-precision and stable lifting and lowering of the blood collection tube base 14. Multiple blood collection tubes 15 are detachably and evenly installed at the bottom of the blood collection tube base 14. Under the lifting and lowering of the blood collection tube base 14, the absorption port at the bottom of the blood collection tube 15 is extended into the plasma and placed above the white blood cells. At the same time, the top of each blood collection tube 15 is connected to a blood collection chamber 19 located inside the blood collection tube base 14. The top middle of the blood collection chamber 19 is connected to a transmission pipe 21 installed on the outside of the top of the blood collection tube base 14 through a connecting pipe. At the same time, a micro pump 20 is set on the connecting pipe to apply an upward suction force to the bottom blood collection tube 15, that is, to control the blood collection tube 15 to collect plasma from the corresponding area. The outer side of the transfer tube 21 is sealed with a sleeve 22. The top of the sleeve 22 is connected to the bottom center of the blood collection chamber plate 11. At the same time, the sleeve 22 corresponding to the top of the transfer tube 21 has a through structure and a non-contact connection to the blood storage chamber 24 opened inside the blood collection chamber plate 11. When the electric telescopic rod 23 controls the blood collection tube base 14 to move up and down, the transfer tube 21 slides synchronously along the inside of the sleeve 22. With the help of the pumping action of the micro pump body 20, the blood collection tube 15 at the bottom is controlled to collect plasma from the corresponding location synchronously. Then, it is transferred to the blood storage chamber 24 through the blood collection chamber 19, connecting tube, transfer tube 21, sleeve 22, etc., so as to realize the function of synchronous collection of plasma from multiple locations at the same time, increasing the collection speed while ensuring sufficient protection of white blood cells.

[0040] In this embodiment of the utility model, a control panel 25 is provided on one side of the top of the blood collection chamber plate 11. The control panel 25 is connected to the electric telescopic rod 23 and the micro pump body 20, which is used to control the synchronous operation of multiple sets of electric telescopic rods 23 and the start and stop of the micro pump body 20. See Figure 7 The top of the blood collection tube 15 is provided with a male valve 16, and the bottom of the blood collection tube base 14 corresponding to the male valve 16 is provided with a female valve 17. The top of the female valve 17 is connected to the blood collection chamber 19, that is, the male valve 16 and the female valve 17 are detachably connected, which makes it easy to replace the blood collection tube 15 as needed. Preferably, the male valve 16 typically has external threads and a tapered head, while the female valve 17 typically has internal threads and an elastic sealing ring. The tapered head of the male valve is inserted into the female valve, and then the outer nut on the male valve is rotated to engage and lock with the threads of the female valve. This can usually be done by "half a turn" or "one turn". Alternatively, plug-in quick connectors can be used.

[0041] In one embodiment of this utility model, see reference Figure 5Multiple "Z"-shaped clamping plates 12 are evenly installed in a ring around the bottom edge of the blood collection chamber plate 11. The clamping plates 12 are engaged with the top edge of the blood separation box 10. The weight of the blood collection chamber plate 11, the blood collection tube base 14, etc., keeps the blood collection chamber plate 11 and the blood collection tube base 14 stably placed on the upper side of the blood separation box 10. At the same time, in order to increase the installation stability between the blood collection chamber plate 11 and the blood separation box 10, an internal clamping plate 13 is engaged with the inner side of each set of clamping plates 12. The internal clamping plate 13 contacts the inner wall of the blood separation box 10, and the inner side of the clamping plate 12 contacts the outer wall of the blood separation box 10, thereby firmly inserting the blood collection chamber plate 11 into the top of the blood separation box 10. Preferably, buffer pads are installed at the contact points between the built-in clamping plate 13, the clamping plate 12 and the outer wall of the blood separation box 10, in order to increase the clamping force and reduce the degree of wear and tear from mutual contact.

[0042] Preferably, the blood separation box 10 is made of disposable injection-molded polypropylene or polycarbonate, which needs to be resistant to centrifugal force and chemical reagents, and can be coated with a low adsorption coating (such as a hydrophobic / low protein adsorption coating) to reduce ctDNA adsorption loss.

[0043] The blood collection chamber plate 11, the card plate 12, and the internal clamping plate 13 are made of fatigue-resistant engineering plastics, such as ABS, polyamide, or aluminum alloy frames; the buffer pads are made of silicone or EPDM rubber.

[0044] Preferably, the micro pump body 20 is a micro peristaltic pump, which propels fluid by compressing a flexible tube, and the pump body does not directly contact the liquid (the liquid only contacts the inner diameter of the tube). Multiple sets of liquid level detection sensors are evenly arranged at the bottom of the blood collection tube base 14 to monitor the distance between the blood collection tube 15 and the white blood cells in real time, so as to ensure accurate sampling.

[0045] It should be noted that the control panel 25 serves as the human-machine interface and the external display of the central controller. It sends operation instructions to the PLC / microcontroller and receives position signals from the electric telescopic rod 23, the liquid level detection sensor, and the flow / status feedback from the micro pump 20. Through closed-loop control logic, it synchronously drives the telescopic rod to rise and fall, pump to suck / stop, and the liquid level detection sensor to detect according to a predetermined sequence. This ensures that suction is only started after the blood collection tube 15 is in position, achieving multi-channel synchronization, abnormal self-checking, and safety interlocking. The above is only a brief introduction. The operating principle is a conventional technology and will not be elaborated on here.

[0046] The working principle of this utility model is as follows: place the blood collection tube 15 or the blood separation box 10 containing blood sample in the designated position of the device, scan the sample barcode and input / confirm the sample information on the touch control panel 25, and separate the whole blood sample into layers in the centrifugation module to obtain the upper plasma layer and the lower white blood cell layer. This process assumes that the centrifugation has been completed and the blood separation box has been returned to the working position. The blood collection chamber plate 11 is snapped into the top clamping plate 12, the built-in clamping plate 13, and the buffer pad of the blood separation box 10 for auxiliary positioning. The blood collection tube base 14 is in the initial upper position. The control panel 25 issues a descent command, and the electric telescopic rod 23 synchronously drives the blood collection tube base 14 to descend, controlling the bottom end of the blood collection tube 15 to extend to the upper plasma layer and be above the white blood cell layer. The micro pump 20 is activated, applying upward suction to each blood collection tube 15. The blood collection tubes transfer the plasma synchronously to the sleeve 22 through the blood collection chamber 19, connecting tube, and transmission tube 21 and deliver it to the blood storage chamber 24. After the predetermined collection volume is reached, the micro pump 20 stops suction, and the electric telescopic rod 23 drives the blood collection tube base 14 back to the initial position. At the same time, abnormality detection and recording are performed.

[0047] It should be understood that in this application, all rotating, sliding, meshing, belt-driven and other moving parts are well lubricated and not prone to slippage or wear, and each part is provided with a corresponding protective shell. However, in the accompanying drawings of this application, the connection state of each moving part is not shown. It should also be understood that all parts in this application are made of metal or plastic materials with suitable strength in the relevant field to ensure that their structural rigidity meets the actual requirements.

[0048] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of the present invention.

Claims

1. A rapid plasma sample processing device for ctDNA methylation detection, characterized in that, Includes a blood separation box (10); a blood collection chamber plate (11) is provided on the top of the blood separation box (10), and a blood collection tube base (14) is provided at the bottom of the blood collection chamber plate (11). Multiple blood collection tubes (15) are detachably and evenly installed at the bottom of the blood collection tube base (14), and the bottom of the blood collection chamber plate (11) and the top of the blood collection tube base (14) are connected by multiple sets of evenly distributed electric telescopic rods (23). Multiple sets of liquid level detection sensors are evenly distributed at the bottom of the blood collection tube base (14). Each blood collection tube (15) is connected to the top of the blood collection chamber (19), and the blood collection chamber (19) is connected to the transmission tube (21) through a connecting tube. The transmission tube (21) is provided with a sleeve (22) on the outside, and a micro pump body (20) is provided on the transmission tube (21). The transmission tube (21) and the sleeve (22) are connected to the blood storage chamber (24).

2. The rapid plasma sample processing device for ctDNA methylation detection according to claim 1, characterized in that, The micro pump body (20) is a micro peristaltic pump.

3. The rapid plasma sample processing device for ctDNA methylation detection according to claim 2, characterized in that, A male valve (16) is provided at the top of the blood collection tube (15), and a female valve (17) is provided at the corresponding position of the blood collection tube base (14).

4. The rapid plasma sample processing device for ctDNA methylation detection according to claim 3, characterized in that, The bottom edge of the blood collection chamber plate (11) is uniformly equipped with multiple "Z"-shaped clamping plates (12). Each set of clamping plates (12) has an internal clamping plate (13) inside. The clamping plates (12) and the internal clamping plate (13) are used to securely clamp the blood collection chamber plate (11) to the top of the blood separation box (10).

5. The rapid plasma sample processing device for ctDNA methylation detection according to claim 4, characterized in that, Buffer pads are installed at the positions where the built-in clamp (13), the card plate (12) contacts the outer wall of the blood separation box (10).

6. The rapid plasma sample processing device for ctDNA methylation detection according to claim 5, characterized in that, The transmission tube (21) slides inside the sleeve (22), and the top of the sleeve (22) is connected to the bottom center of the blood collection chamber plate (11). The sleeve (22) is provided with a through structure to connect non-contactly to the blood storage chamber (24) inside the blood collection chamber plate (11).

Images