An on-line magnetic capture device for a flow cytometer

Abstract

This application relates to an online magnetic capture device for flow cytometers, comprising a magnetic frame and a capture tube. The magnetic frame includes a frame body and a magnet disposed on the frame body. A slot for limiting the capture tube is provided on the frame body. The capture tube has a top interface at its top and a bottom interface at its bottom. The magnet is located near the middle of the capture tube along its length. This application offers the advantages of online treatment of flow cytometer waste liquid, simple operation, and no need for filter membranes, thus eliminating clogging issues and facilitating the capture of magnetic bead complexes. The capture tube and magnetic frame are completely independent of each other. The capture tube serves as both the sample flow path and reaction vessel, and can be used once or easily cleaned; the magnetic frame serves as a reusable magnetic field generating unit, achieving functional separation and reducing operating costs.

Description

Technical Field

[0001] This application relates to the field of flow cytometer auxiliary equipment, and more particularly to an online magnetic capture device for flow cytometers. Background Technology

[0002] In trace pollutant screening analyses (such as mycotoxins, pesticide residues, and veterinary drug residues) based on immunomagnetic beads and flow cytometry, when a positive result is obtained, mass spectrometry (such as LC-MS / MS) confirmation of the target analyte is often required. Current confirmation procedures typically require reprocessing the original sample, which is cumbersome and may result in insufficient sample volume. The waste liquid after flow cytometry analysis actually contains a complete "magnetic bead-antibody-toxin-fluorescent label" complex, making it an ideal material for confirmation. However, the waste liquid volume is large, the concentration of the target complex is extremely low, and direct processing is inefficient.

[0003] In related technologies, a filter membrane is usually simply installed at the end of the waste liquid output pipe, which has the problem of easy clogging of the filter membrane, making it difficult to separate the target complex. Utility Model Content

[0004] To address the difficulty in separating magnetic bead complexes from waste liquid, this application provides an online magnetic capture device for flow cytometers.

[0005] This application provides an online magnetic capture device for flow cytometers, which adopts the following technical solution:

[0006] An online magnetic capture device for flow cytometer includes a magnetic frame and a capture tube. The magnetic frame includes a frame body and a magnet disposed on the frame body. The frame body is provided with a slot for limiting the capture tube. The top of the capture tube is provided with a top interface and the bottom of the capture tube is provided with a bottom interface. The magnet is located near the middle of the capture tube along its length.

[0007] By employing the above technical solution, during online capture, the waste liquid output tube of the flow cytometer is inserted into the capture tube. The bottom of the capture tube is open and aligned with the waste liquid container, and the capture tube is located within the magnetic field region corresponding to the magnetic rack. After starting the flow cytometer, the waste liquid containing the target magnetic bead complex flows into the capture tube. As the waste liquid flows through the tube wall region corresponding to the magnet, the magnetic bead complex is instantly and firmly adsorbed onto the inner wall of the capture tube under the influence of a strong radial gradient magnetic field. The clarified waste liquid continues to flow downwards and is discharged directly into the waste liquid container below through the open bottom of the tube.

[0008] This allows for online treatment of waste liquid from flow cytometers. It employs specific adsorption, is simple to operate, and eliminates the need for filter membranes, thus avoiding clogging issues and facilitating the capture of magnetic bead complexes.

[0009] Optionally, it also includes a tube cap with a central through-hole, the tube cap being connected to the top interface, the central through-hole being used to connect to the waste liquid output tube of the flow cytometer.

[0010] By adopting the above technical solution, the waste liquid output pipe is connected through a central through-hole. This ensures a stable connection and prevents the waste liquid output pipe from easily detaching from the capture pipe.

[0011] Optionally, a bottom closure is also included, which is a bottom cover used to close the bottom interface.

[0012] By adopting the above technical solution, after the online capture operation is completed, the bottom interface is sealed with the bottom cap, the tube cap is opened, eluent is added to the capture tube, the capture tube is removed from the magnetic rack, the tube cap is closed, and the tube is vortexed for several seconds to fully disperse the magnetic beads in the eluent. Then, the capture tube is reinserted into the magnetic rack and allowed to stand for about 1 minute to allow the magnetic beads to be re-adsorbed onto the tube wall. The clear eluent containing the target analyte in the capture tube is transferred to a sample vial. The eluent can be directly used for subsequent LC-MS / MS and other mass spectrometry confirmatory analyses.

[0013] Optionally, it also includes a bottom closure, which is a centrifuge tube with a centrifuge interface that connects to the bottom interface.

[0014] By adopting the above technical solution, after the online capture operation is completed, the centrifuge tube is connected to the bottom interface through the centrifuge interface, eluent is added to the capture tube, the capture tube is removed from the magnetic rack, the tube cap is closed, and the tube is vortexed for several seconds to allow the magnetic beads to be fully dispersed in the eluent and enter the centrifuge tube. Then, the capture tube is separated, the centrifuge tube is inserted into the magnetic rack, and left to stand for about 1 minute to allow the magnetic beads to be re-adsorbed onto the centrifuge tube wall. The clear eluent containing the target analyte in the capture tube is transferred to a sample vial. The eluent can be directly used for subsequent LC-MS / MS and other mass spectrometry confirmatory analyses.

[0015] Optionally, the frame includes a base frame, a connecting plate, and a top frame. The slot is disposed on the top frame, the base frame is provided with a drain hole, the connecting plate connects the base frame and the side of the top frame, and the magnet is disposed on the connecting plate.

[0016] By adopting the above technical solution, the base frame is used to support the platform to make the support stable. The connecting plate connects the base frame and the side of the top frame, so that the middle of the capture tube is exposed for easy observation. At the same time, the magnet is set on the connecting plate, and the side wall of the connecting plate is exposed to facilitate the installation of the magnet.

[0017] Optionally, the outer wall of the capture tube is provided with an anti-detachment ring, the outer diameter of which is larger than the inner diameter of the slot.

[0018] By adopting the above technical solution, the anti-detachment ring prevents the capture tube from falling out of the slot.

[0019] Optionally, the top interface is provided with internal or external threads, and the pipe cap is threaded to the top interface.

[0020] By adopting the above technical solution, it is easy to connect the top interface and the pipe cap, and it is not easy to leak liquid.

[0021] Optionally, the bottom interface is provided with internal or external threads, and the bottom cover is threadedly connected to the bottom interface.

[0022] By adopting the above technical solution, it is easy to connect the bottom interface and the bottom cover, and it is not easy to leak liquid.

[0023] Optionally, the centrifuge interface is provided with internal or external threads, and the centrifuge interface is threadedly connected to the bottom interface.

[0024] By adopting the above technical solution, it is easy to connect the centrifuge interface and the bottom interface, and it is not easy to leak liquid.

[0025] In summary, this application includes at least one of the following beneficial technical effects:

[0026] 1. The proposed solution achieves reliable online capture: During flow cytometry, the capture tube is first inserted into the slot of the magnetic rack, and then the device is connected to the waste liquid output line of the flow cytometer. As the waste liquid flows through the capture tube, the magnetic bead complex is rapidly adsorbed onto the inner wall of the tube under the action of a high-gradient magnetic field generated by the magnet, thereby treating the waste liquid of the flow cytometer online. The operation is simple, and no filter membrane is required, thus eliminating the problem of easy clogging and facilitating the capture of the magnetic bead complex.

[0027] 2. The capture tube and magnetic rack are completely independent. The capture tube serves as the sample flow path and reaction vessel, and can be used once or easily cleaned; the magnetic rack, as a reusable magnetic field generating unit, is protected. This design achieves functional separation and reduces operating costs.

[0028] 3. It facilitates seamless switching of elution modes. After the test is completed, screw on the bottom tube cap, take out the capture tube from the magnetic rack, open the top tube cap, add acetonitrile for denaturation elution, and then reinsert the magnetic rack to adsorb magnetic beads to collect the pure supernatant. The operation is extremely flexible and helps to improve elution efficiency.

[0029] 4. Simple structure and strong compatibility: The device has no valves, no moving parts, and requires no power supply. The standard threaded interface at the bottom of the capture tube allows it to directly use common laboratory centrifuge tubes as bottom caps or collection tubes, greatly improving practicality and economy.

[0030] 5. High versatility: The device proposed in this application is not only suitable for small molecule pollutant detection systems, but can also be widely used in other flow cytometry detection and analysis scenarios based on immunomagnetic beads to recover target magnetic bead complexes. Attached Figure Description

[0031] Figure 1 This is a structural diagram of the online magnetic capture device for flow cytometer in Example 1.

[0032] Figure 2 This is a structural diagram of the online magnetic capture device for flow cytometer in Example 2.

[0033] Explanation of reference numerals in the attached diagram: 1. Magnetic frame; 2. Capture tube; 21. Top interface; 22. Bottom interface; 11. Frame; 111. Base frame; 112. Connecting plate; 113. Top frame; 114. Slot; 115. Drain hole; 12. Magnet; 3. Tube cap; 31. Central through hole; 4. Bottom closure; 41. Bottom cover; 42. Centrifuge tube; 421. Centrifuge interface; 5. Anti-detachment ring; 6. Waste liquid output tube. Detailed Implementation

[0034] The present application will be further described in detail below with reference to the accompanying drawings.

[0035] Example 1

[0036] 1. Embodiment 1 of this application discloses an online magnetic trapping device for flow cytometers. (Refer to...) Figure 1 An online magnetic capture device for flow cytometers includes a magnetic frame 1 and a capture tube 2, wherein the magnetic frame 1 is used to support the capture tube 2.

[0037] The magnetic frame 1 includes a frame body 11 and a magnet 12 disposed on the frame body 11. The frame body 11 includes a base frame 111, a connecting plate 112, and a top frame 113. The top frame 113 has a slot 114 for inserting a capture tube 2, and the base frame 111 has a drain hole 115. The connecting plate 112 connects the base frame 111 and the side of the top frame 113, and the magnet 12 is fixed to the side wall of the connecting plate 112, with the magnet 12 located between the top frame 113 and the base frame 111.

[0038] The capture tube 2 is a transparent tubular structure that runs vertically through the tube (e.g., made of polypropylene). The top of the capture tube 2 has a top interface 21, and the bottom of the capture tube 2 has a bottom interface 22. Both the top interface 21 and the bottom interface 22 are provided with internal or external threads; in this embodiment, the top interface 21 and the bottom interface 22 are provided with external threads. The drain hole 115 is directly below the bottom interface 22, and the magnet 12 is located near the middle of the length of the capture tube 2.

[0039] When the capture tube 2 is located inside the slot 114, each capture tube 2 has a corresponding magnet 12 on its sidewall to ensure that a high-intensity, high-gradient radial magnetic field is formed in the central region of the slot 114.

[0040] An online magnetic capture device for a flow cytometer also includes a cap 3 and a bottom closure 4. The cap 3 has a central through-hole 31 for connecting the waste liquid output tube 6 of the flow cytometer.

[0041] The bottom closure 4 is a bottom cover 41. The inner wall of the bottom cover 41 is provided with internal threads. The bottom cover 41 is used to connect the bottom interface 22 by threads.

[0042] In this application, the internal thread on the bottom cover 41, the external thread on the bottom interface 22, and the external thread on the top interface 21 are all standard threads, which are compatible with the commonly used threaded tube cover 3 or centrifuge tube 42 in the laboratory.

[0043] The outer wall of the capture tube 2 is integrally formed with an anti-detachment ring 5. The outer diameter of the anti-detachment ring 5 is larger than the inner diameter of the slot 114, so that the capture tube 2 will not fall out of the slot 114. The inner diameter of the drain hole 115 is larger than the outer diameter of the bottom cover 41, so that the bottom cover 41 is fixed to the capture tube 2 when it is not necessary to remove the capture tube 2 from the slot 114.

[0044] The implementation principle of an online magnetic capture device for flow cytometer according to Embodiment 1 of this application is as follows: Preparation and assembly stage: The capture tube 2 is vertically inserted into the slot 114 of the external magnetic frame 1 to ensure its stability. The top cap is screwed onto the top interface 21 at the top of the capture tube 2. The bottom interface 22 of the capture tube 2 remains open so that the bottom outlet of the capture tube 2 is directly aligned with the laboratory waste container.

[0045] Online capture phase: Pass the waste liquid output tube 6 of the flow cytometer through the central through-hole 31 of the tube cap 3 and insert it into the capture tube 2, ensuring that the end of the waste liquid output tube 6 is above the magnetic field region corresponding to the magnetic frame 1. Start the flow cytometer, and the waste liquid containing the target magnetic bead complex flows into the capture tube 2 in the direction of the magnetic field. When the waste liquid flows through the tube wall region corresponding to the permanent magnet, the magnetic bead complex is instantly and firmly adsorbed onto the inner wall of the capture tube 2 under the action of a strong radial gradient magnetic field. The clarified waste liquid continues to flow downward and is discharged directly into the waste liquid tank below through the open tube bottom.

[0046] The magnetic beads inside the tube are mainly affected by the magnetic field attraction force Fm and the fluid drag force Fd. In this embodiment, the magnet 12 uses a high-performance neodymium iron boron magnet that adheres closely to the outer wall of the capture tube 2, generating an extremely high magnetic field gradient exceeding 100 T / m in the near-wall region within the capture tube 2. For commercially available superparamagnetic microspheres with a diameter of 1-3 μm, the Fm generated under this gradient can reach hundreds of piconewtons to nanonewtons. However, the flow rate of flow cytometry wastewater is typically below 10 mL / min, resulting in a very low linear flow rate within the cross-sectional area of ​​the capture tube 2. Therefore, the Fd for a single micrometer-sized magnetic bead is only on the piconewton scale. Thus, the condition Fm >> Fd is always satisfied, ensuring that the magnetic beads can be reliably captured throughout the entire operating flow rate range of the instrument and will not be washed away.

[0047] Elution and Collection Stage: After the flow cytometry analysis is complete, remove the waste output tube 6 from the capture tube 2. Tighten a bottom cap 41 to the bottom interface 22 to the bottom of the capture tube 2. Open the cap 3 and add eluent (e.g., 200-500 μL of acetonitrile for denaturing and eluting the target analyte) to the capture tube 2. Remove the capture tube 2 from the magnetic rack 1, replace the cap 3, and vortex for several seconds to fully disperse the magnetic beads in the eluent. Then reinsert the capture tube 2 into the magnetic rack 1 and let it stand for about 1 minute to allow the magnetic beads to be re-adsorbed onto the tube wall. Transfer the clear eluent containing the target analyte from the capture tube 2 to a sample vial. The eluent can be directly used for subsequent LC-MS / MS and other mass spectrometry confirmatory analyses.

[0048] Example 2

[0049] Reference Figure 2 The difference between this embodiment and embodiment 1 is that the bottom sealing member 4 is a centrifuge tube 42, and the centrifuge tube 42 is provided with a centrifuge interface 421 that is connected to the bottom interface 22. The centrifuge interface 421 is provided with an internal thread, and the centrifuge interface 421 is threadedly connected to the bottom interface 22.

[0050] The implementation principle of Example 2 is as follows: After the flow cytometry analysis is completed, centrifuge tube 42 is connected to bottom interface 22 through centrifuge interface 421. Eluent (e.g., 200-500 μL of acetonitrile, used for denaturing and eluting the target analyte) is added to capture tube 2. Capture tube 2 is removed from magnetic rack 1, and tube cap 3 is placed on it. The tube is vortexed for several seconds to allow the magnetic beads to be fully dispersed in the eluent and enter centrifuge tube 42. Then, capture tube 2 is separated, centrifuge tube 42 is inserted into magnetic rack 1, and left to stand for about 1 minute to allow the magnetic beads to be re-adsorbed onto the wall of centrifuge tube 42. The clear eluent containing the target analyte in capture tube 2 is transferred to a sample vial using a disposable dropper. The eluent can be directly used for subsequent LC-MS / MS and other mass spectrometry confirmatory analyses.

[0051] The above are all preferred embodiments of this application, and are not intended to limit the scope of protection of this application. Therefore, all equivalent changes made in accordance with the structure, shape and principle of this application should be covered within the scope of protection of this application.

Claims

1. An on-line magnetic capture device for a flow cytometer, characterized by: The device includes a magnetic frame (1) and a capture tube (2). The magnetic frame (1) includes a frame (11) and a magnet (12) disposed on the frame (11). The frame (11) is provided with a slot (114) for limiting the capture tube (2). The top of the capture tube (2) is provided with a top interface (21), and the bottom of the capture tube (2) is provided with a bottom interface (22). The magnet (12) is located near the middle of the capture tube (2) in the length direction.

2. An on-line magnetic capture device for a flow cytometer according to claim 1, characterized in that: It also includes a tube cap (3) with a central through hole (31) connected to the top interface (21), and the central through hole (31) is used to connect to the waste liquid output tube (6) of the flow cytometer.

3. An on-line magnetic capture device for a flow cytometer according to claim 1, characterized in that: It also includes a bottom closure (4), which is a bottom cover (41) used to close the bottom interface (22).

4. An on-line magnetic capture device for a flow cytometer according to claim 1, characterized in that: It also includes a bottom closure (4), which is a centrifuge tube (42) and a centrifuge interface (421) connected to the bottom interface (22).

5. An on-line magnetic capture device for a flow cytometer according to claim 1, wherein: The frame (11) includes a base frame (111), a connecting plate (112), and a top frame (113). The slot (114) is provided on the top frame (113). The base frame (111) is provided with a drain hole (115). The connecting plate (112) connects the base frame (111) and the side of the top frame (113). The magnet (12) is provided on the connecting plate (112).

6. An on-line magnetic capture device for a flow cytometer according to claim 1, wherein: The outer wall of the capture tube (2) is provided with an anti-detachment ring (5), the outer diameter of which is larger than the inner diameter of the slot (114).

7. An on-line magnetic capture device for a flow cytometer according to claim 2, wherein: The top interface (21) is provided with internal or external threads, and the pipe cap (3) is threaded to the top interface (21).

8. An on-line magnetic capture device for a flow cytometer according to claim 3, wherein: The bottom interface (22) is provided with internal or external threads, and the bottom cover (41) is threaded to the bottom interface (22).

9. An on-line magnetic capture device for a flow cytometer according to claim 4, characterized in that: The centrifugal interface (421) is provided with internal or external threads, and the centrifugal interface (421) is threadedly connected to the bottom interface (22).

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