Flow cell electroporation device

Abstract

A flow cell electroporation device, comprising a first electrode sheet, a second electrode sheet, and an electroporation cavity body. The electroporation cavity body is internally provided with an electroporation cavity, an annular channel, a flow guide cavity, an inlet hole communicated with the electroporation cavity, and an outlet hole communicated with the flow guide cavity; two opposite ends of the annular channel are respectively communicated with the electroporation cavity and the flow guide cavity; the first electrode sheet and the second electrode sheet are respectively mounted on two opposite inner side walls of the electroporation cavity; and the axis of the electroporation cavity, the axis of the annular channel, and the center line of the inlet hole coincide. The cell electroporation device has the ability of continuously processing a large volume of cell fluid without the risk of channel blockage, and also ensures the uniformity of cell electroporation results, thereby improving the overall transfection efficiency and viability.

Description

A flow cytometry electroporation device

[0001] This application claims priority to Chinese Patent Application No. 202411905355.5, filed on December 23, 2024, entitled “A Flow Cytometry Electroporation Device”, the entire contents of which are incorporated herein by reference. Technical Field

[0002] This application belongs to the field of medical device technology, and in particular relates to a flow cytometry electroporation device. Background Technology

[0003] Electroporation, also known as cell electroporation, is a typical non-biological cell transfection technique. It utilizes relatively short (milliseconds to tens of milliseconds) high-intensity electrical pulses to directly act on the cell membrane, temporarily creating tiny pores (electroporations) that allow exogenous macromolecules (such as DNA, RNA, and proteins) to enter the cell. After a short period (milliseconds to seconds) of self-repair, the pores close, thus achieving electroporation.

[0004] Conventional electroporation typically utilizes two parallel plate electrodes for electroporation. By applying a short voltage to the two electrodes, an electric field is created in the region between them, allowing cells within this region to electroporate with exogenous substances. While this method can achieve some cell transfection, the voltage experienced by cells at different locations within the electric field varies, resulting in a relatively low overall transfection efficiency. Furthermore, conventional electroporation devices can only perform single-pass electroporation and cannot handle large volumes of solutions or continuous flow electroporation.

[0005] To address this, researchers improved upon the existing technology by constructing microchannels or microcavities between two parallel plate electrodes, achieving continuous flow electroporation processing capability (CN112639112A). Although this flow cytometry electroporation device can achieve continuous flow electroporation of a certain volume, its electroporation chamber volume is small, limiting the sample volume and throughput processed in continuous flow mode. Furthermore, during relatively high-flow-rate cell electroporation, the fluid distribution inside the electroporation chamber is uneven, especially in the horizontal direction perpendicular to the fluid flow direction. This results in significant differences in the time cells at different locations are subjected to the electric field during continuous flow, leading to inconsistent overall cell transfection results. Summary of the Invention

[0006] This application aims to solve the technical problems in the prior art, such as the inability to achieve high-throughput, large-volume cell electroporation and the non-uniform flow field in continuous flow electroporation, and provides a flow cytometry cell electroporation device.

[0007] To solve the above-mentioned technical problems, the first embodiment of this application provides a cell electroporation device, including a first electrode plate, a second electrode plate, and an electroporation chamber; the electroporation chamber is provided with an electroporation chamber, an annular channel, a flow guide chamber, an inlet hole communicating with the electroporation chamber, and an outlet hole communicating with the flow guide chamber; the opposite ends of the annular channel are respectively connected to the electroporation chamber and the flow guide chamber.

[0008] The first electrode plate and the second electrode plate are respectively mounted on the two opposite inner sidewalls of the electro-rotation cavity;

[0009] The axis of the electro-rotating cavity, the axis of the annular channel, and the center line of the inlet hole coincide.

[0010] Optionally, the annular channel includes multiple arc-shaped channels, which are distributed annularly around the axis of the electro-rotation cavity, and the central angle and width of each arc-shaped channel are equal.

[0011] Optionally, the flow guiding cavity is a frustum-shaped cavity, and the center line of the inlet, the axis of the electro-rotating cavity, the axis of the frustum-shaped cavity, and the center line of the outlet coincide; the inner diameter of the frustum-shaped cavity gradually increases from the outlet towards the annular channel.

[0012] Optionally, the first electrode plate includes a first electrode disc and a first electrode lead-out piece connected to the first electrode disc, wherein the first electrode disc is attached to the top inner wall of the electro-rotation cavity;

[0013] The second electrode plate includes a second electrode disc and a second electrode lead-out piece connected to the second electrode disc, the second electrode disc being attached to the bottom inner wall of the electro-rotation cavity;

[0014] The first electrode disc and the second electrode disc are arranged in parallel. The diameter of the first electrode disc is equal to the diameter of the second electrode disc, and the axis of the first electrode disc, the axis of the second electrode disc, and the axis of the electro-rotation cavity coincide.

[0015] Optionally, the electro-rotation cavity includes a first cover plate, an electro-rotation block, an intermediate block, and a second cover plate stacked sequentially; the inlet is disposed on the first cover plate, the electro-rotation cavity is disposed on the electro-rotation block, the annular channel is disposed on the intermediate block, and the guide cavity and the outlet are both disposed on the second cover plate.

[0016] Optionally, the first cover plate is further provided with a first receiving groove communicating with the inlet hole, and the first electrode sheet is installed in the first receiving groove;

[0017] The intermediate block is also provided with a second receiving groove, the annular channel is arranged around the second receiving groove, and the second electrode sheet is installed in the second receiving groove.

[0018] Optionally, the first cover plate is further provided with a first annular positioning groove surrounding the first electrode sheet, and the electro-rotating block is further provided with a first annular positioning protrusion surrounding the electro-rotating cavity; the first annular positioning protrusion is inserted into the first annular positioning groove.

[0019] The electro-rotating block has a second annular positioning groove at one end opposite to the first annular positioning protrusion. The second annular positioning groove surrounds the electro-rotating cavity. The middle block has a second annular positioning protrusion surrounding the annular channel. The second annular positioning protrusion is inserted into the second annular positioning groove.

[0020] The middle block has a third annular positioning protrusion at one end opposite to the second annular positioning protrusion. The third annular positioning protrusion is arranged around the annular channel. The second cover plate also has a third annular positioning groove arranged around the flow guide cavity. The third annular positioning protrusion is inserted into the third annular positioning groove.

[0021] Optionally, the first cover plate is further provided with a first threaded hole, the first threaded hole communicating with the end of the inlet hole away from the electro-rotation cavity; and / or

[0022] The second cover plate is also provided with a second threaded hole, which connects to the end of the outlet hole away from the guide cavity.

[0023] Optionally, the first electrode sheet is further provided with a through hole, and the inlet hole is connected to the electro-rotation cavity through the through hole.

[0024] Optionally, the electro-rotation cavity is cylindrical.

[0025] In this application, the electroporation chamber is provided with an inlet, an electroporation chamber, an annular channel, a guide cavity, and an outlet in sequence. The axis of the electroporation chamber coincides with the center line of the inlet. Cell fluid flows into the electroporation chamber through the inlet. The electroporation chamber is a cylindrical or frustum-shaped orifice. After the cell fluid flows vertically into the center of the electroporation chamber, it disperses evenly in all directions (360 degrees) in the radial direction of the electroporation chamber, and the flow velocity is the same in each radial direction of the electroporation chamber. Specifically, cell sap flows into the electroporation chamber through a narrow inlet. The inlet is relatively narrow, resulting in a high flow velocity of the cell sap within it. The electroporation chamber itself is more spacious, causing the flow velocity of the cell sap to decrease abruptly at the inlet, with the flow direction changing 90 degrees and becoming uniformly dispersed. As the cell sap flows away from the inlet, the flow velocity remains almost constant at different radial positions. Simultaneously, before flowing into the electroporation chamber from the inlet, the cell sap flows along the axial direction of the chamber. As the cell sap flows within the chamber, its flow direction changes to the radial direction, resulting in the same continuous flow velocity at different horizontal heights. Because this flow cytometry electroporation device employs an axisymmetric design, multiple coaxial cylindrical surfaces are cut around the axis of the electroporation chamber. Software simulation analysis shows that the flow velocity distribution and magnitude of the cell sap remain consistent on each cylindrical surface.

[0026] In addition, the first electrode and the second electrode are respectively mounted on the two opposite inner sidewalls of the electroporation chamber; the first electrode and the second electrode are arranged in parallel, which can form a uniformly distributed electric field in the electroporation chamber, and the direction of the electric field lines is the axis of the electroporation chamber and perpendicular to the flow direction of the cell fluid in the electroporation chamber. This ensures that the electric field intensity experienced by cells at different positions in the electroporation chamber is consistent at the same time, thereby improving the uniformity and stability of cell transfection in the electroporation chamber.

[0027] In this application, the cell fluid flows radially within the electroporation chamber. Due to the axially symmetric design, the flow rate of the cell fluid is equal in all directions on the same cylindrical surface. Furthermore, an electric field perpendicular to the flow direction of the cell fluid can be formed within the electroporation chamber, allowing for stable and uniform transfection. After transfection in the electroporation chamber, the cell fluid enters the guiding chamber through the annular channel. Since the axis of the guiding chamber coincides with the axis of the electroporation chamber, the flux of the cell fluid in the annular channel is equal in all directions. The cell fluid then converges in the guiding chamber and is discharged through the outlet.

[0028] In this application, the cell electrotransfection device has the ability to continuously process large volumes of cell fluid, supporting long-term, high-throughput cell electrotransfection without the risk of channel blockage. Simultaneously, the highly uniform electric field distribution and symmetrical flow field design within the electrotransfection chamber ensure that each cell in the continuous flow experiences the same electric field intensity. The continuous flow of cell fluid within the electrotransfection chamber exhibits the same velocity magnitude and consistent flow direction in both radial and vertical dimensions. This uniform flow direction is perpendicular to the uniform electric field direction, ensuring that each cell in the continuous flow experiences the same number of electric shocks for the same duration, guaranteeing the uniformity of cell electrotransfection results, thereby improving overall transfection efficiency and viability. Attached Figure Description

[0029] The present application will be further described below with reference to the accompanying drawings and embodiments.

[0030] Figure 1 is a schematic diagram of the structure of a cell electroporation device provided in an embodiment of this application;

[0031] Figure 2 is a cross-sectional view of a cell electroporation apparatus provided in an embodiment of this application;

[0032] Figure 3 is a schematic diagram of the structure of the first electrode sheet of the cell electrotransfection device provided in an embodiment of this application mounted on the first cover plate;

[0033] Figure 4 is a schematic diagram of the structure of the electrotransfer block of a cell electrotransfection device provided in an embodiment of this application;

[0034] Figure 5 is a schematic diagram of the structure of the second electrode sheet of the cell electrotransfection device provided in an embodiment of this application, which is mounted on the intermediate block;

[0035] Figure 6 is a schematic diagram of the structure of the second cover plate of the cell electroporation device provided in an embodiment of this application;

[0036] Figure 7 is an axial cross-sectional view of the electroporation chamber in this application, and a schematic diagram of the flow of cell fluid in the electroporation chamber;

[0037] Figure 8 is a radial cross-sectional view of the electroporation chamber in this application, and a schematic diagram of the flow of cell fluid in the electroporation chamber;

[0038] Figure 9 is a horizontal cross-sectional simulation diagram of the distribution of fluid velocity and flow direction when the cell flow enters the electroporation chamber;

[0039] Figure 10 is a simulation diagram of the fluid velocity distribution at three different locations on the curved surface where the cell flow enters the electroporation chamber;

[0040] Figure 11 is a simulation diagram of the uniform electric field distribution and electric field direction formed inside the electro-rotation cavity.

[0041] The reference numerals in the accompanying drawings are as follows: 1. First electrode plate; 11. First electrode disc; 12. First electrode lead-out plate; 13. Through hole; 2. Second electrode plate; 21. Second electrode disc; 22. Second electrode lead-out plate; 3. Electro-rotating cavity; 31. Electro-rotating cavity; 32. Annular channel; 321. Arc-shaped channel; 33. Guide cavity; 34. Inlet hole; 35. Outlet hole; 36. First cover plate; 361. First receiving groove; 362. First annular positioning groove; 363. First threaded hole; 37. Electro-rotating block; 371. First annular positioning protrusion; 372. Second annular positioning groove; 38. Intermediate block; 381. Second receiving groove; 382. Second annular positioning protrusion; 383. Third annular positioning protrusion; 39. Second cover plate; 391. Third annular positioning groove; 392. Second threaded hole. Detailed Implementation

[0042] To make the technical problems, technical solutions, and beneficial effects solved by this application clearer, the following detailed description is provided in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the scope of this application.

[0043] It should be understood that the terms "upper", "lower", "left", "right", "front", "rear", "middle", etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are only for the convenience of describing this application 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. Therefore, they should not be construed as limitations of this application.

[0044] As shown in Figures 1 and 2, the first embodiment of this application provides a cell electroporation device, including a first electrode plate 1, a second electrode plate 2, and an electroporation chamber 3. The electroporation chamber 3 is provided with an electroporation chamber 31, an annular channel 32, a flow guiding chamber 33, an inlet 34 connecting the electroporation chamber 31, and an outlet 35 connecting the flow guiding chamber 33. The opposite ends of the annular channel 32 are respectively connected to the electroporation chamber 31 and the flow guiding chamber 33. Preferably, the electroporation chamber 31 is cylindrical. Understandably, the inlet 34, the electroporation chamber 31, the annular channel 32, the flow guiding chamber 33, and the outlet 35 are sequentially connected.

[0045] The first electrode plate 1 and the second electrode plate 2 are respectively mounted on the two opposite inner sidewalls of the electro-rotation cavity 31; it can be understood that the first electrode plate 1 and the second electrode plate 2 are respectively attached to the upper and lower inner walls of the electro-rotation cavity 31; the first electrode plate 1 and the second electrode plate 2 are arranged in parallel, and one of the first electrode plate 1 and the second electrode plate 2 is a positive electrode plate and the other is a negative electrode plate.

[0046] The axis of the electro-rotating cavity 31, the axis of the annular channel 32, and the center line of the inlet hole 34 coincide.

[0047] In this application, the electroporation chamber 3 is provided with an inlet 34, an electroporation chamber 31, an annular channel 32, a guide chamber 33, and an outlet 35 connected in sequence. The axis of the electroporation chamber 31 coincides with the center line of the inlet 34. Cell fluid flows into the electroporation chamber 31 through the inlet 34. The electroporation chamber 31 is a cylindrical or frustum-shaped orifice. As shown in Figures 7 and 8, after the cell fluid flows vertically into the center of the electroporation chamber 31, it disperses evenly in all directions (360 degrees) in the radial direction of the electroporation chamber 31, and the flow velocity is the same in each radial direction of the electroporation chamber 31. Specifically, cell fluid flows into the electroporation chamber 31 through a narrow inlet 34. The inlet 34 is relatively narrow, resulting in a high flow velocity of the cell fluid within it. The electroporation chamber 31, with its wider space, experiences a sudden decrease in flow velocity at its inlet, a 90-degree turn in direction, and uniform dispersion. As the cell fluid flows away from the inlet, the flow velocity remains almost constant at different radial positions. Simultaneously, before flowing into the electroporation chamber 31 from the inlet 34, the cell fluid flows along the axial direction of the chamber. As the cell fluid flows within the chamber, its flow direction changes to the radial direction, maintaining the same continuous flow velocity at different horizontal heights. Because this flow cytometry electroporation device employs an axisymmetric design, multiple coaxial cylindrical surfaces are cut around the axis of the electroporation chamber 31. Software simulation analysis shows that the flow velocity distribution and magnitude of the cell fluid remain consistent on each cylindrical surface.

[0048] Furthermore, as shown in Figure 7, the first electrode plate 1 and the second electrode plate 2 are respectively installed on the two opposite inner sidewalls of the electroporation chamber 31. The first electrode plate 1 and the second electrode plate 2 are arranged in parallel, which can form a uniformly distributed electric field in the electroporation chamber 31. The direction of the electric field lines is the axial direction of the electroporation chamber 31 and the direction of the electric field lines is perpendicular to the flow direction of the cell fluid in the electroporation chamber 31. This ensures that the electric field intensity experienced by cells at different positions in the electroporation chamber 31 is consistent at the same time, thereby improving the uniformity and stability of cell transfection in the electroporation chamber 31.

[0049] In this application, the cell fluid flows radially within the electroporation chamber 31. Due to the axially symmetric design, the flow rate of the cell fluid is equal in all directions on the same cylindrical surface. Furthermore, an electric field perpendicular to the flow direction of the cell fluid can be formed within the electroporation chamber 31, allowing for stable and uniform transfection within the chamber. After transfection in the electroporation chamber 31, the cell fluid enters the guiding chamber 33 through the annular channel 32. Since the axis of the guiding chamber 33 coincides with the axis of the electroporation chamber 31, the flux of the cell fluid in the annular channel 32 is equal in all directions. The cell fluid then converges in the guiding chamber 33 and is discharged through the outlet 35.

[0050] In this application, the cell electrotransfection device has the ability to continuously process large volumes of cell fluid, supporting long-term, high-throughput cell electrotransfection without the risk of channel blockage. Simultaneously, the highly uniform electric field distribution and symmetrical flow field design within the electrotransfection chamber 31 ensure that each cell in the continuous flow experiences the same electric field intensity. The continuous flow of cell fluid within the electrotransfection chamber 31 exhibits the same velocity magnitude and consistent flow direction in both radial and vertical dimensions. The direction of this uniform flow is perpendicular to the direction of the uniform electric field, ensuring that each cell in the continuous flow experiences the same number of electric shocks for the same duration, thus guaranteeing the uniformity of the cell electrotransfection results and improving overall transfection efficiency and viability.

[0051] In one embodiment, as shown in FIG5, the annular channel 32 includes a plurality of arc-shaped channels 321, which are distributed annularly around the axis of the electroporation chamber 31, and the central angle and width of each arc-shaped channel 321 are equal. Understandably, the number of arc-shaped channels 321 can be set according to actual needs; for example, 4, 6, or 8 arc-shaped channels 321 may be provided. In this embodiment, the width and central angle of the plurality of arc-shaped channels 321 are equal, and the centers of the plurality of arc-shaped channels 321 coincide. When the cell fluid in the electroporation chamber 31 flows into the guide cavity 33 through each arc-shaped channel 321, the flux and velocity of the cell fluid in each guide cavity 33 are equal, thereby ensuring the stability of cell fluid accumulation in the guide cavity 33 and improving cell viability.

[0052] In one embodiment, as shown in Figures 2 and 6, the guide cavity 33 is a frustum-shaped cavity. The centerline of the inlet 34, the axis of the electro-rotation cavity 31, the axis of the frustum-shaped cavity, and the centerline of the outlet 35 coincide. From the outlet 35 towards the annular channel 32, the inner diameter of the frustum-shaped cavity gradually increases. Understandably, the maximum inner diameter of the frustum-shaped cavity is equal to the inner diameter of the annular channel 32, and the minimum inner diameter of the frustum-shaped cavity is equal to the inner diameter of the outlet 35. In this embodiment, the inner wall of the frustum-shaped cavity is connected to the inner wall of the annular channel 32. During the flow of cell fluid through the annular channel 32 into the frustum-shaped cavity, the frustum-shaped cavity guides the cell fluid to the outlet 35, ensuring the stability of the cell fluid discharge from the outlet 35.

[0053] In one embodiment, as shown in Figures 3 and 5, the first electrode plate 1 includes a first electrode disc 11 and a first electrode lead-out piece 12 connected to the first electrode disc 11. The first electrode disc 11 is attached to the top inner wall of the electro-rotation cavity 31. Understandably, the end of the first electrode lead-out piece 12 away from the first electrode disc 11 is located outside the electro-rotation cavity 3.

[0054] The second electrode plate 2 includes a second electrode disc 21 and a second electrode lead-out piece 22 connected to the second electrode disc 21. The second electrode disc 21 is attached to the bottom inner wall of the electro-rotation cavity 31. Understandably, the end of the second electrode lead-out piece 22 away from the second electrode disc 21 is located outside the electro-rotation cavity 3.

[0055] The first electrode disc 11 and the second electrode disc 21 are arranged in parallel. The diameter of the first electrode disc 11 is equal to the diameter of the second electrode disc 21, and the axes of the first electrode disc 11, the second electrode disc 21, and the electrotransfer chamber 31 coincide. In this embodiment, the first electrode disc 11 and the second electrode disc 21 are coaxially arranged and respectively attached to the upper and lower inner walls of the electrotransfer chamber 31. Therefore, after the first electrode disc 11 and the second electrode disc 21 are energized, a uniformly distributed electric field can be formed within the electrotransfer chamber 31. The direction of the electric field is the axial direction of the electrotransfer chamber 31 and perpendicular to the flow direction of the cell fluid within the electrotransfer chamber 31. This ensures that the electric field intensity experienced by cells at different locations within the electrotransfer chamber 31 is consistent at the same time, improving the uniformity and stability of cell transfection within the electrotransfer chamber 31.

[0056] In one embodiment, as shown in FIG1, the electro-rotation cavity 3 includes a first cover plate 36, an electro-rotation block 37, an intermediate block 38, and a second cover plate 39 stacked sequentially. An inlet 34 is disposed on the first cover plate 36, the electro-rotation cavity 31 is disposed on the electro-rotation block 37, the annular channel 32 is disposed on the intermediate block 38, and the guide cavity 33 and the outlet 35 are both disposed on the second cover plate 39. Understandably, the first cover plate 36 and the electro-rotation block 37, the electro-rotation block 37 and the intermediate block 38, and the intermediate block 38 and the second cover plate 39 can be fixed and sealed by curing with ultraviolet adhesive (UV adhesive), or by methods such as thermosetting bonding, laser bonding, ultrasonic bonding, and hydrophilic treatment bonding (plasma treatment, etc.). In this embodiment, the electroporation chamber 3 is designed as a split structure consisting of a first cover plate 36, an electroporation block 37, an intermediate block 38, and a second cover plate 39, which reduces the manufacturing difficulty and cost of the cell electroporation device.

[0057] In one embodiment, as shown in FIG3, the first cover plate 36 is further provided with a first receiving groove 361 communicating with the inlet hole 34, and the first electrode plate 1 is installed in the first receiving groove 361; in this embodiment, the first electrode plate 1 is embedded in the first receiving groove 361, thereby ensuring the stability of the first electrode plate 1 installed on the first cover plate 36.

[0058] In one embodiment, as shown in FIG5, the intermediate block 38 is further provided with a second receiving groove 381, and the annular channel 32 is arranged around the second receiving groove 381. The second electrode plate 2 is installed in the second receiving groove 381. The second electrode plate 2 is embedded in the second receiving groove 381, thereby ensuring the stability of the second electrode plate 2 installed on the intermediate block 38.

[0059] In one embodiment, as shown in Figures 2 to 4, the first cover plate 36 is further provided with a first annular positioning groove 362 surrounding the first electrode sheet 1, and the electro-rotating block 37 is further provided with a first annular positioning protrusion 371 surrounding the electro-rotating cavity 31; the first annular positioning protrusion 371 is inserted into the first annular positioning groove 362; in this embodiment, the first annular positioning protrusion 371 is adapted to the first annular positioning groove 362, and during the process of installing the first cover plate 36 on the electro-rotating block 37, the first annular positioning protrusion 371 is inserted into the first annular positioning groove 362, thereby achieving precise positioning between the first cover plate 36 and the electro-rotating block 37.

[0060] In one embodiment, as shown in Figures 2 and 5, the end of the electro-rotating block 37 opposite to the first annular positioning protrusion 371 is provided with a second annular positioning groove 372, which surrounds the electro-rotating cavity 31. The intermediate block 38 is provided with a second annular positioning protrusion 382 surrounding the annular channel 32. The second annular positioning protrusion 382 is inserted into the second annular positioning groove 372. In this embodiment, the second annular positioning protrusion 382 is adapted to the second annular positioning groove 372. During the process of installing the electro-rotating block 37 on the intermediate block 38, the second annular positioning protrusion 382 is inserted into the second annular positioning groove 372, thereby achieving precise positioning between the electro-rotating block 37 and the intermediate block 38.

[0061] In one embodiment, as shown in Figures 2 and 6, the middle block 38 has a third annular positioning protrusion 383 at one end opposite to the second annular positioning protrusion 382. The third annular positioning protrusion 383 surrounds the annular channel 32, and the second cover plate 39 also has a third annular positioning groove 391 surrounding the guide cavity 33. The third annular positioning protrusion 383 is inserted into the third annular positioning groove 391. In this embodiment, the third annular positioning protrusion 383 is adapted to the third annular positioning groove 391. During the installation of the middle block 38 on the second cover plate 39, the third annular positioning protrusion 383 is inserted into the third annular positioning groove 391, thereby achieving precise positioning between the middle block 38 and the second cover plate 39.

[0062] In one embodiment, as shown in FIG1, the first cover plate 36 is further provided with a first threaded hole 363, which connects to the end of the inlet hole 34 away from the electric rotary cavity 31; it can be understood that the first threaded hole 363 is located at the upper end of the inlet hole 34, and the design of the first threaded hole 363 facilitates the connection of the inlet hole 34 with the external pipe.

[0063] In one embodiment, as shown in FIG2, the second cover plate 39 is further provided with a second threaded hole 392, which connects to the end of the outlet 35 away from the guide cavity 33. Understandably, the second threaded hole 392 is located at the lower end of the outlet 35, and its design facilitates the connection of the outlet 35 to an external pipe.

[0064] In one embodiment, as shown in FIG2, the first electrode plate 1 is further provided with a through hole 13, and the inlet hole 34 communicates with the electro-rotation cavity 31 through the through hole 13. It can be understood that the through hole 13 is located at the center of the first electrode disc 11, and the inner diameter of the through hole 13 is equal to the inner diameter of the inlet hole 34. Further, the second electrode plate 2 comprises a complete electrode disc (i.e., the second electrode disc).

[0065] Furthermore, Figure 9 shows a simulation diagram of the velocity and flow direction distribution in the horizontal cross-section of the electro-conversion cavity. As shown in Figure 9A, the fluid in the central region of the circular electro-conversion cavity 31 with the horizontal cross-section initially enters through the inlet 32 ​​with a relatively high velocity. As the fluid diffuses into the electro-conversion cavity 31, the velocity rapidly decreases and gradually remains constant. Meanwhile, as shown in Figure 9B, since the fluid enters from the center of the circular electro-conversion cavity 31, it flows uniformly horizontally in all directions (360°) after entering the cavity, meaning the fluid velocity is the same in any direction on the horizontal plane. Taking the center of the cylindrical electro-conversion cavity 31 as the origin, three cylindrical surfaces with different radii are cut, as shown in Figures 9A-C. The simulation results show that the fluid velocity distribution and magnitude remain consistent on the three different cylindrical surfaces. In summary, the high-flux continuous cellular flow within the electro-conversion cavity 31 exhibits the same velocity magnitude in all directions in both the horizontal and vertical dimensions. Furthermore, the simulation diagrams of the electric field distribution and direction of the central vertical section of the electroporation chamber 31 are shown in Figures 10 and 11. The electric field distribution within the electroporation region is highly uniform, and the electric field direction is vertically downward, perpendicular to the horizontal direction of the continuous fluid within the electroporation region. This ensures that the electric field intensity experienced by cells at different locations within the electroporation chamber 31 is consistent at the same time, thereby improving the uniformity and stability of cell transfection efficiency within the electroporation chamber 31.

[0066] After electroporation, the cells flow from the edge of the electroporation chamber 31 through the annular channel 32 of the intermediate layer to the guide chamber 33 of the second cover plate 39. Because the four support strips of the intermediate layer are perpendicular to each other, the four arc-shaped channels 321 formed have the same shape and cross-sectional area, and each arc-shaped channel 321 carries the same amount of cells in the same amount of time. After entering the guide chamber 33, the four cell streams converge again and exit from the outlet 35.

[0067] This application proposes a flow cytometry electrotransfection device that supports long-term, high-throughput cell electrotransfection while reducing the risk of fluid blockage. Secondly, the cell flow injection direction is perpendicular to the fluid flow direction during electrotransfection, resulting in more uniform vertical fluid flow. Simultaneously, after entering the electrotransfection chamber, the fluid flows in a radial (360°) direction, ensuring the same flow velocity in all horizontal directions during electrotransfection. Furthermore, the first electrode plate 1 and the second electrode plate 2 are placed parallel to each other, forming a large-volume electrotransfection chamber 31 with a uniformly distributed electric field intensity inside. In summary, thanks to the unique chamber structure design, this application ensures the uniformity of the continuous fluid in both horizontal and vertical directions during high-throughput continuous flow electrotransfection, and generates a uniform electric field perpendicular to the continuous fluid flow direction within the large-volume electrotransfection chamber. This ensures that each cell in the continuous flow receives consistent electric shocks over a consistent duration, guaranteeing the uniformity of the cell electrotransfection results and thereby improving the overall cell transfection rate and viability.

[0068] The beneficial effects of this application include the following:

[0069] 1. The overall axisymmetric structural design ensures that after the fluid continuously enters the electro-rotating cavity 31 from the inlet, it can flow radially and radially in the horizontal direction within the electro-rotating cavity 31, thereby ensuring that the flow velocity and flow direction are the same in all directions, and that the continuous flow velocity is the same at different vertical positions in the same horizontal direction.

[0070] 2. The direction of the fluid entering the electro-rotation chamber 31 in a continuous flow is perpendicular to the direction of the fluid in the electro-rotation region. It enters vertically and flows out horizontally, with the flow direction changing by 90°. The vertical component of the flow direction is small.

[0071] 3. Two electrode plates (i.e., the first electrode plate 1 and the second electrode plate 2) are arranged in parallel inside the electro-rotation cavity 31. The electric field intensity formed after applying voltage is uniformly distributed throughout the electro-rotation region. More specifically, the electric field is distributed along the direction perpendicular to the fluid motion and is perpendicular to the horizontal direction of the continuous flow in the electro-rotation region.

[0072] 4. This application ensures that each cell in the continuous flow experiences the same number of electrical shocks and time when passing through the electroporation region in two ways: a highly uniform electric field distribution in the vertical direction within the electroporation region, and a uniform flow velocity distribution along different radial angles in the horizontal direction; this ensures the uniformity and stability of the performance of large-volume cells in flow cytometry electroporation, and improves the overall transfection efficiency and viability of the cell flow.

[0073] The above are merely embodiments of the cell electroporation device of this application and are not intended to limit this application. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. A flow cytometry electroporation device, wherein, It includes a first electrode plate, a second electrode plate, and an electro-rotation cavity; the electro-rotation cavity is provided with an electro-rotation chamber, an annular channel, a flow guide cavity, an inlet hole communicating with the electro-rotation cavity, and an outlet hole communicating with the flow guide cavity; the opposite ends of the annular channel are respectively connected to the electro-rotation cavity and the flow guide cavity; The first electrode plate and the second electrode plate are respectively mounted on the two opposite inner sidewalls of the electro-rotation cavity; The axis of the electro-rotating cavity, the axis of the annular channel, and the center line of the inlet hole coincide.

2. The flow cytometry electroporation apparatus according to claim 1, wherein, The annular channel includes multiple arc-shaped channels, which are distributed in a ring around the axis of the electro-rotation cavity, and the central angle and width of each arc-shaped channel are equal.

3. The flow cytometry electroporation apparatus according to claim 1, wherein, The flow guide cavity is a frustum-shaped cavity, and the center line of the inlet, the axis of the electric rotating cavity, the axis of the frustum-shaped cavity, and the center line of the outlet coincide; the inner diameter of the frustum-shaped cavity gradually increases from the outlet towards the annular channel.

4. The flow cytometry electroporation apparatus according to claim 3, wherein, The maximum inner diameter of the frustum-shaped cavity is equal to the inner diameter of the annular channel, and the minimum inner diameter of the frustum-shaped cavity is equal to the inner diameter of the outlet hole.

5. The flow cytometry electroporation apparatus according to claim 1, wherein, The first electrode plate includes a first electrode disc and a first electrode lead-out piece connected to the first electrode disc, and the first electrode disc is attached to the top inner wall of the electro-rotation cavity; The second electrode plate includes a second electrode disc and a second electrode lead-out piece connected to the second electrode disc, the second electrode disc being attached to the bottom inner wall of the electro-rotation cavity; The first electrode disc and the second electrode disc are arranged in parallel. The diameter of the first electrode disc is equal to the diameter of the second electrode disc, and the axis of the first electrode disc, the axis of the second electrode disc, and the axis of the electro-rotation cavity coincide.

6. The flow cytometry electroporation apparatus according to claim 1, wherein, The electro-rotation cavity includes a first cover plate, an electro-rotation block, an intermediate block, and a second cover plate stacked sequentially; the inlet is disposed on the first cover plate, the electro-rotation cavity is disposed on the electro-rotation block, the annular channel is disposed on the intermediate block, and the flow guide cavity and the outlet are both disposed on the second cover plate.

7. The flow cytometry electroporation apparatus according to claim 6, wherein, The first cover plate is also provided with a first receiving groove that communicates with the inlet hole, and the first electrode sheet is installed in the first receiving groove; The intermediate block is also provided with a second receiving groove, the annular channel is arranged around the second receiving groove, and the second electrode sheet is installed in the second receiving groove.

8. The flow cytometry electroporation apparatus according to claim 6, wherein, The first cover plate is also provided with a first annular positioning groove surrounding the first electrode sheet, and the electric rotating block is also provided with a first annular positioning protrusion surrounding the electric rotating cavity; the first annular positioning protrusion is inserted into the first annular positioning groove; The electro-rotating block has a second annular positioning groove at one end opposite to the first annular positioning protrusion. The second annular positioning groove surrounds the electro-rotating cavity. The middle block has a second annular positioning protrusion surrounding the annular channel. The second annular positioning protrusion is inserted into the second annular positioning groove. The middle block has a third annular positioning protrusion at one end opposite to the second annular positioning protrusion. The third annular positioning protrusion is arranged around the annular channel. The second cover plate also has a third annular positioning groove arranged around the flow guide cavity. The third annular positioning protrusion is inserted into the third annular positioning groove.

9. The flow cytometry electroporation apparatus according to claim 6, wherein, The first cover plate is also provided with a first threaded hole, which is connected to the end of the inlet hole away from the electric rotary cavity.

10. The flow cytometry electroporation apparatus according to claim 6, wherein, The second cover plate is also provided with a second threaded hole, which connects to the end of the outlet hole away from the guide cavity.

11. The flow cytometry electroporation apparatus according to any one of claims 1 to 10, wherein, The first electrode plate is also provided with a through hole, and the inlet hole is connected to the electro-rotation cavity through the through hole.

12. The flow cytometry electroporation apparatus according to any one of claims 1 to 10, wherein, The electro-rotation cavity is cylindrical.

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