A multifunctional cell culture multiwell plate for electroporation

Abstract

This application relates to the field of cell electroporation technology, and provides a multifunctional cell culture multiwell plate for electroporation, including a consumable cover, a consumable interlayer, and a consumable base plate. The top surface of the base plate has multiple wells for containing cell solutions. The consumable interlayer is located on top of the base plate, and a cavity for containing cell solutions is formed between the consumable interlayer and the base plate. Electrode slots are provided on the consumable interlayer at positions corresponding to both sides of each well. The consumable cover is located on top of the interlayer, and a removable electrode is provided on the cover at a position corresponding to each electrode slot. An impedance detection electrode assembly is provided on the bottom surface of the base plate at a position corresponding to each well. This application improves the practicality of the multifunctional cell culture multiwell plate.

Description

Technical Field

[0001] This application relates to the field of cell electroporation technology, and in particular to a multifunctional cell culture multiwell plate for electroporation. Background Technology

[0002] Most existing technical solutions employ a consumable structure design based on electroporation cups. This consumable structure limits the low-throughput electroporation to a very small number (one or a few) of electroporation cups at a time, which is unsuitable for the high-throughput, large-scale applications required in the biopharmaceutical field.

[0003] Existing electroporation consumables only function as electroporation containers; cells cannot be cultured within them, and they have no other practical value. Due to their structure, current electroporation consumables cannot perform real-time monitoring before and after electroporation. Electrodes can ionize metal ions, leading to cell death. Furthermore, most existing electrodes are parallel plate electrodes, resulting in electrode polarization at the corners. Therefore, current containers for electroporation lack practicality. Summary of the Invention

[0004] This application provides a multifunctional cell culture multiwell plate for electroporation, which can solve the problem of insufficient practicality of containers for electroporation.

[0005] This application provides a multifunctional cell culture multiwell plate for electroporation, comprising: The consumable cover, consumable interlayer, and consumable base plate are provided. The top surface of the consumable base plate is provided with multiple holes for containing cell solution. The consumable interlayer covers the top of the consumable base plate, and a receiving cavity for containing cell solution is formed between the consumable interlayer and the consumable base plate. Electrode slots are provided on both sides of each hole on the consumable interlayer. The consumable cover is located on the top of the consumable interlayer, and a detachable electrode is provided on the consumable cover at the position corresponding to each electrode slot. An impedance detection electrode group is provided at the position corresponding to each hole on the bottom surface of the consumable base plate.

[0006] Optionally, a set of pin through holes is provided on the consumable cover at the position corresponding to each hole.

[0007] Optionally, the consumable base plate is made of conductive silver paste.

[0008] Optionally, the impedance sensing electrode assembly may include multiple impedance sensing electrodes.

[0009] Optionally, the outer surface of the consumable cover is provided with an insulating protective layer.

[0010] Optionally, a mounting structure is provided on the consumable cover at the position corresponding to the electrode, and the electrode is detachably connected to the mounting structure.

[0011] Optionally, the apex of the electrode has a chamfered structure.

[0012] Optionally, the electrode is an elliptical electrode.

[0013] Optionally, the consumable interlayer and the consumable base plate are an integrated structure.

[0014] The above-mentioned solution in this application has the following beneficial effects: In the embodiments of this application, a multifunctional cell culture multiwell plate for electroporation includes a consumable cover, a consumable interlayer, and a consumable base plate. The top surface of the base plate has multiple wells for containing cell solutions. The consumable interlayer covers the top of the base plate, and a cavity for containing cell solutions is formed between the interlayer and the base plate. Electrode slots are provided on the interlayer at positions corresponding to both sides of each well. The consumable cover is located on top of the interlayer, and a removable electrode is provided on the cover at a position corresponding to each electrode slot. An impedance detection electrode group is provided on the bottom surface of the base plate at a position corresponding to each well. By providing multiple wells, the high-throughput, high-volume electroporation requirements can be met, improving the practicality of the multifunctional cell culture multiwell plate. Cell culture can be performed directly in the wells without inserting electrodes, while avoiding cell death during culture due to metal ion ionization, further enhancing the practicality of the multifunctional cell culture multiwell plate.

[0015] Other beneficial effects of this application will be described in detail in the following detailed description section. Attached Figure Description

[0016] To more clearly illustrate the technical solutions in the embodiments of this application, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0017] Figure 1 This is a schematic diagram of the structure of the top surface of the consumable cover provided in one embodiment of this application; Figure 2 This is a schematic diagram of the structure of the consumable interlayer and consumable base plate provided in an embodiment of this application; Figure 3 This is a schematic diagram of the structure of a consumable base plate provided in one embodiment of this application; Figure 4 This is a schematic diagram of the structure of the bottom surface of the consumable cover provided in an embodiment of this application; Figure 5 This is a schematic diagram of the specific structure of an impedance detection electrode assembly provided in an embodiment of this application; Figure 6 Electrode schematic provided for an embodiment of this application Figure 1 ; Figure 7 Electrode schematic provided for an embodiment of this application Figure 2 ; Figure 8 This is a schematic diagram of an electric field simulation provided in an embodiment of this application; Figure 9 This is a schematic diagram of a single well of a multifunctional cell culture multiwell plate provided in an embodiment of this application.

[0018] Figure 10 This is a schematic diagram of the overall structure of the consumable interlayer and consumable base plate provided in an embodiment of this application.

[0019] [Explanation of Labels in the Attached Image] 1—Consumable cover, 2—Consumable interlayer, 3—Consumable base plate, 11—Electrode, 12—Ejector pin through hole, 21—Hole position, 22—Electrode slot, 31—Impedance detection electrode group, 32—Impedance detection electrode. Detailed Implementation

[0020] In the following description, specific details such as particular system architectures and techniques are set forth for illustrative purposes and not for limitation, in order to provide a thorough understanding of the embodiments of this application. However, those skilled in the art will understand that this application may also be implemented in other embodiments without these specific details. In other instances, detailed descriptions of well-known systems, apparatuses, circuits, and methods have been omitted so as not to obscure the description of this application with unnecessary detail.

[0021] It should be understood that, when used in this application specification and the appended claims, the term "comprising" indicates the presence of the described features, integrals, steps, operations, elements and / or components, but does not exclude the presence or addition of one or more other features, integrals, steps, operations, elements, components and / or a collection thereof.

[0022] It should also be understood that the term “and / or” as used in this application specification and the appended claims means any combination of one or more of the associated listed items and all possible combinations, and includes such combinations.

[0023] As used in this application specification and the appended claims, the term "if" may be interpreted, depending on the context, as "when," "once," "in response to determination," or "in response to detection." Similarly, the phrase "if determined" or "if detected [the described condition or event]" may be interpreted, depending on the context, as "once determined," "in response to determination," "once detected [the described condition or event]," or "in response to detection [the described condition or event]."

[0024] Furthermore, in the description of this application and the appended claims, the terms "first," "second," "third," etc., are used only to distinguish descriptions and should not be construed as indicating or implying relative importance.

[0025] References to "one embodiment" or "some embodiments" as described in this specification mean that one or more embodiments of this application include a specific feature, structure, or characteristic described in connection with that embodiment. Therefore, the phrases "in one embodiment," "in some embodiments," "in other embodiments," "in still other embodiments," etc., appearing in different parts of this specification do not necessarily refer to the same embodiment, but rather mean "one or more, but not all, embodiments," unless otherwise specifically emphasized. The terms "comprising," "including," "having," and variations thereof mean "including but not limited to," unless otherwise specifically emphasized.

[0026] To address the issue of poor practicality in existing containers for electroporation, this application provides a multifunctional cell culture multiwell plate for electroporation, comprising a consumable cover, a consumable interlayer, and a consumable base plate. The top surface of the base plate has multiple wells for containing cell solutions. The consumable interlayer covers the top of the base plate, forming a cavity for containing cell solutions between the interlayer and the base plate. Electrode slots are located on both sides of each well on the interlayer. The consumable cover is located on top of the interlayer, and a removable electrode is located on the cover corresponding to each electrode slot. An impedance detection electrode assembly is located on the bottom surface of the base plate corresponding to each well. By providing multiple wells, the high-throughput, high-volume electroporation requirements can be met, improving the practicality of the multifunctional cell culture multiwell plate. Cell culture can be performed directly in the wells without electrodes being inserted, while avoiding cell death during culture due to metal ion ionization, further enhancing the practicality of the multifunctional cell culture multiwell plate.

[0027] The following is an exemplary description of the multifunctional cell culture multiwell plate provided in this application.

[0028] like Figures 1 to 4 As shown, the multifunctional cell culture multiwell plate provided in this application includes: a consumable cover 1, a consumable interlayer 2, and a consumable base plate 3. The top surface of the consumable base plate 3 is provided with a plurality of wells 21 for containing cell solution. The consumable interlayer 2 covers the top of the consumable base plate 3, and a receiving cavity for containing cell solution is formed between the consumable interlayer 2 and the consumable base plate 3.

[0029] Electrode slots 22 are provided on both sides of each hole 21 on the consumable interlayer 2. The consumable cover 1 is located on the top of the consumable interlayer 2, and a removable electrode 11 is provided on the consumable cover 1 at the position corresponding to each electrode slot 22 (e.g., Figure 4 (As shown).

[0030] An impedance detection electrode group 31 is provided on the bottom surface of the consumable base plate 3 at the position corresponding to each hole 21.

[0031] It should be noted that in the aforementioned multifunctional cell culture multiwell plate, the consumable interlayer 2 is provided with an m×n array of wells, enabling the consumable interlayer 2 to hold a large number of cell solutions at once. Correspondingly, each well 21 has an electrode slot 22 on both sides, that is, there are 2×m×n electrode slots 22 on the consumable interlayer 2. For the consumable cover 1, an electrode 11 is provided at the top of each electrode slot 22, and the electrode 11 is inserted into the electrode slot by insertion. For the consumable base plate 3, an impedance detection electrode group 31 is provided at the bottom of each well 21.

[0032] During electroporation, cell solution is filled into well 21, and the consumable cover 1 is connected to the consumable interlayer 2, so that electrode 11 is inserted into the corresponding electrode slot 22 and comes into contact with the cell solution in the corresponding well 21. The power supply and other equipment are electrically connected to the electrode 11 to make the electrode 11 energized. The two electrodes 11 in each well 21 simultaneously apply voltage to the cell solution to perform the electroporation process. At the same time, the multimeter and other equipment are electrically connected to the impedance detection electrode group 31 to collect data such as current and impedance of the cell solution.

[0033] An exemplary consumable interlayer 2 has an insulating material such as polyethylene on its inner surface to prevent electrical conductivity between the holes.

[0034] In some optional embodiments, a set of pin through holes 12 are provided on the consumable cover 1 at the position corresponding to each hole.

[0035] It should be noted that the pin through hole 12 is specifically set at the corresponding position of the electrode 11, so that the wire, metal pin, etc. can be connected to the electrode 11 through the pin through hole 12. The other end of the wire or metal pin is connected to the power supply or other equipment, so that the electrode 11 is energized.

[0036] In some optional embodiments, the aforementioned consumable base plate 3 is made of conductive silver paste.

[0037] It should be noted that the conductive silver paste can conduct electricity, enabling the impedance detection electrode group 31 on the consumable base plate 3 to be electrically connected to the cell solution in the well position 21.

[0038] In some optional embodiments, the impedance detection electrode group 31 includes a plurality of impedance detection electrodes 32 (e.g., Figure 5 (As shown).

[0039] It should be noted that the number of impedance detection electrodes 32 can be four, preferably arranged in a rectangular or symmetrical pattern, for connecting wires, etc., which in turn connect to devices such as meters to achieve impedance detection. The impedance detection method can be a four-electrode measurement with two-point excitation and two-point sampling (to reduce polarization error), or multiple combination measurements can be achieved through circuit switching.

[0040] Impedance detection is used for, but is not limited to: confirming the liquid loading, conductivity and contact status before electroporation, and confirming the adaptive voltage adjustment of the load impedance; collecting impedance comparisons before electroporation to identify abnormalities such as bubbles, local drying, short circuits / open circuits, and arcs; and performing real-time detection after electroporation to assess cell damage before and after electroporation, as well as to evaluate cell electroporation efficiency and healing efficiency.

[0041] In some optional embodiments, the outer surface of the consumable cover 1 is provided with an isolation protective layer, which can be an insulating material such as polyethylene, for electrical isolation to prevent electrical conduction between electrodes or between other electrodes and the cell solution during electrotransfection.

[0042] In some optional embodiments, a mounting structure is provided on the consumable cover 1 at the position corresponding to the electrode 11, and the electrode 11 is detachably connected to the mounting structure.

[0043] For example, the above-mentioned installation structure can be a snap-on type, a slide rail type, a threaded type, or a magnetic connection structure. When it is a snap-on type structure, the consumable cover 1 is provided with a slot, and the electrode 11 is provided with a snap. The snap and the slot fit together to snap and fix the electrode 11 onto the consumable cover 1. When it is a slide rail type, the consumable cover 1 is provided with a guide rail, and the electrode 11 is provided with a slider. The slider and the guide rail slide together to fix the electrode 11 onto the consumable cover 1. When it is a threaded type, the consumable cover 1 is provided with an internal thread, and the electrode 11 is provided with an external thread. The external thread and the internal thread fit together to fix the electrode 11 onto the consumable cover 1. When it is a magnetic type, both the consumable cover 1 and the electrode 11 are provided with magnetic elements. The electrodes are fixed onto the consumable cover 1 by the magnetic attraction force of the magnetic elements.

[0044] In some optional embodiments, the apex of electrode 11 has a chamfered structure (e.g., Figure 6 As shown in the figure, part a is a top view of a pair of electrodes, part b is a side view, and part c is a bottom view.

[0045] For example, in addition to the chamfered structure described above, electrode 11 can also be an elliptical electrode (such as...). Figure 7As shown in the figure, part a is a front view of a pair of electrodes, part b is a side view, and part c is a top view. In actual production, the structure of electrode 11 is selected according to requirements, and in the same multifunctional cell culture multiwell plate, all electrodes 11 have the same structure.

[0046] It should be noted that setting electrode 11 as a chamfered or elliptical structure, or a double-elliptical or continuously curvature transition structure, is used to smooth the field line distribution, reduce the tip effect, and increase the solution volume; the chamfer / rounded corners are used to reduce the concentration of field strength at the edges and improve the uniformity of the field strength within the pore. The above structures can be used individually or in combination. Figure 8 As shown, Figure 8 a is a schematic diagram of the electric field simulation of the flat plate electrode. Figure 8 b is a schematic diagram of the electric field simulation of the chamfered electrode. Figure 8 c is a schematic diagram of the electric field simulation of the elliptical electrode. As can be seen from the figure, the electric field distribution at the edge of the flat plate electrode is uneven, while the electric field of the chamfered electrode and the electric field strength at the edge of the elliptical electrode are concentrated, thus achieving uniform pressure application to the cell solution when voltage is applied.

[0047] When the vertex of electrode 11 is chamfered, the electrode slot 22 has the same shape as electrode 11 to accommodate the insertion of electrode 11, which is a rectangular groove with a chamfered bottom vertex.

[0048] When electrode 11 is an elliptical electrode, electrode slot 22 has the same shape as electrode 11 to accommodate the insertion of electrode 11, which is an elliptical groove.

[0049] It should be noted that a microporous membrane and / or polymer coating can be applied to the electrode surface. This interface layer, by controlling the dielectric properties, thickness, and microporous structure of the material, allows the pulsed electric field to still effectively couple into the pore medium, while reducing direct contact between the electrode material and the solution, thereby reducing the impact of ion release and reaction products on cell activity.

[0050] For example, the single well of the multifunctional cell culture multiwell plate of this application is as follows: Figure 9 As shown, the bottom is the consumable base plate 3, the two sides are electrodes 11, and the dots represent impedance detection electrodes 32.

[0051] In some embodiments of this application, the dimensions of the multi-well cell culture plate conform to the SBS standard, facilitating practical production use. Furthermore, by making the electrode 11 a detachable structure, the consumable cover 1 can be replaced with an electrode-free version (i.e., the electrode 11 is removed), to meet the need for simplified operation and achieve integrated functions of electroporation, impedance detection, and cell culture.

[0052] For example, such as Figure 10As shown, the consumable interlayer 2 and the consumable base plate 3 can be an integrated structure, that is, the consumable interlayer 2 and the consumable base plate 3 can be directly combined to form a whole, thus constituting the double-layer structure of the consumable cover 1 and the combined whole. In actual production, the required structure can be selected according to the needs.

[0053] It should be noted that when using this multifunctional cell culture multiwell plate, insert the electrodes into the corresponding electrode slots, connect the electrodes to the metal pins through the pin holes, and electrically connect the metal pins to the power supply, signal generator, etc., to energize the electrodes. The electrodes then apply high voltage to the cell fluid in the wells to perform the electrotransfection process. The impedance detection electrode at the bottom connects to a multimeter, signal generator, etc., allowing the multimeter to collect the impedance of the cell fluid. Before cell electrotransfection begins, a low-voltage signal can be output to the impedance detection electrode via the signal generator to detect the impedance of the wells and identify any abnormalities such as wells not being filled with fluid. If no abnormalities are found, the electrodes are energized through the pins, applying high voltage to the cell fluid in the wells to perform the electrotransfection process. When not performing electrotransfection, the electrodes on the consumable cover can be removed, and the wells in the consumable compartment can be used as cell culture space.

[0054] It is worth mentioning that by setting multiple well positions, the high-throughput and large-volume use of electroporation can be met, improving the practicality of multifunctional cell culture multiwell plates. Cell culture can be carried out directly in the well positions without the electrode being inserted, while avoiding cell death caused by metal ion ionization during culture, further improving the practicality of multifunctional cell culture multiwell plates.

[0055] Furthermore, this application has the following advantages: 1. High throughput and standardization compatibility: High-throughput consumables suitable for industrial production and compliant with SBS specifications, adaptable to automated pipetting, robotic arms and plate reading / imaging workflows.

[0056] 2. The electroporation process can be monitored: Impedance detection can be achieved using the four electrodes at the bottom. Before electroporation, abnormal liquid loading, bubbles, arcs, poor contact, etc. can be identified, improving repeatability and reliability. After electroporation, real-time detection can be performed to assess cell damage before and after electroporation, and the cell electroporation efficiency and healing efficiency can be assessed in real time after electroporation.

[0057] 3. Direct culture on the same plate after electroporation: The whole structure is made of biocompatible material, and the vertical high-voltage electrode is detachable. After removal, the space inside the well is released and the adhesion area for cell growth is increased, reducing transfer loss and the risk of contamination.

[0058] 4. Reduce the impact of side reactions: Microporous membranes and / or polymer coatings reduce the impact of metal ion release while ensuring the effective effect of the electric field, thereby improving cell activity and culture stability.

[0059] 5. Improve electric field uniformity: The double elliptical or chamfered structure suppresses the concentration of the electric field strength at the edge, improves the uneven distribution of the electric field at the edge of the square hole, enhances the consistency within the same hole, and reduces the risk of excessive local electroporation.

[0060] 6. Modular structure, easy to mass production and maintenance: The high voltage application electrode and the detection electrode have clear division of labor, the electrode components are replaceable / reusable, and the plate can be mass-produced.

[0061] The above are preferred embodiments of this application. It should be noted that, for those skilled in the art, several improvements and modifications can be made without departing from the principles of this application, and these improvements and modifications should also be considered within the scope of protection of this application.

Claims

1. A multi-functionalized cell culture multi-well plate for electroporation, characterized in that, The multifunctional cell culture multiwell plate includes a consumable cover (1), a consumable interlayer (2) and a consumable base plate (3). The top surface of the consumable base plate (3) is provided with a plurality of holes (21) for containing cell solution. The consumable interlayer (2) covers the top of the consumable base plate (3), and a cavity for containing cell solution is formed between the consumable interlayer (2) and the consumable base plate (3). Electrode slots (22) are provided on the consumable interlayer (2) at positions corresponding to both sides of each hole (21). The consumable cover (1) is located on the top of the consumable interlayer (2). A detachable electrode (11) is provided on the consumable cover (1) at a position corresponding to each electrode slot (22). Impedance detection electrode groups (31) are provided at the positions corresponding to each of the holes (21) on the bottom surface of the consumable base plate (3).

2. The multi-functionalized cell culture multi-well plate of claim 1, wherein, The consumable cover (1) has a set of pin through holes (12) at the position corresponding to each hole.

3. The multi-functionalized cell culture multi-well plate of claim 1, wherein, The consumable base plate (3) contains conductive silver paste.

4. The multi-functionalized cell culture multi-well plate of claim 1, wherein, The impedance detection electrode group (31) includes multiple impedance detection electrodes (32).

5. The multi-functionalized cell culture multi-well plate of claim 2, wherein, The outer surface of the consumable cover (1) is provided with an isolation protective layer.

6. The multi-functionalized cell culture multi-well plate of claim 2, wherein, The consumable cover (1) is provided with an installation structure at the position corresponding to the electrode (11), and the electrode (11) is detachably connected to the installation structure.

7. The multi-functionalized cell culture multi-well plate of claim 1, wherein, The vertex of the electrode (11) has a chamfered structure.

8. The multi-functionalized cell culture multi-well plate of claim 1, wherein, The electrode (11) is an elliptical electrode.

9. The multi-functionalized cell culture multi-well plate of claim 1, wherein, The consumable interlayer (2) and the consumable base plate (3) are an integrated structure.

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