ELECTRODE AND ELECTRODE KIT
20250014777 ยท 2025-01-09
Inventors
Cpc classification
C12M35/02
CHEMISTRY; METALLURGY
International classification
Abstract
An object is to provide an electrode that is easily arranged at a predetermined position when electroporation is performed. The object is achieved by an electrode used for performing electroporation, the electrode includes: at least two or more conductors; and a member configured to hold each conductor and expose at least one end of each of the conductors to outside, the member has a first face where the one end of each of the conductors is exposed, and at least one or more protruding parts provided to the first face, and the length by which the one or more protruding parts protrude in an axis direction from the first face is longer than the length by which the one end of each of the conductors is exposed from the first face.
Claims
1. An electrode used for performing electroporation, the electrode comprising: at least two or more conductors; and a member configured to hold at least a portion of each conductor inside and protrude at least one end of each of the conductors to outside, wherein the member has a first face where the one end of each of the conductors is protruded, and at least one or more protruding parts provided to the first face, and a recess provided in the first face, and wherein the recess is recessed in an axis direction of the member from the first face and penetrates through in a side face direction of the member, wherein the length by which the one or more protruding parts protrude in an axis direction from the first face is longer than the length by which the one end of each of the conductors is protruded from the first face.
2. The electrode according to claim 1, wherein the one or more protruding parts comprise two or more protruding parts, and the two or more protruding parts are spaced apart from each other.
3. The electrode according to claim 2, wherein when, in an outer circumferential portion of the first face, a portion interposed between virtual lines extending from the conductors facing each other is defined as an outer circumferential virtual region, a clearance, which is provided by the protruding parts being spaced apart from each other, and the outer circumferential virtual region at least partially overlap each other.
4. The electrode according to claim 1, wherein a slope part is provided to the first face.
5. The electrode according to claim 2, wherein a slope part is provided to the first face.
6. The electrode according to claim 3, wherein a slope part is provided to the first face.
7. The electrode according to claim 1, wherein the one or more protruding parts are arranged on the outer circumferential side from each of the conductors protruded in the first face.
8. The electrode according to claim 1, wherein the member has a through hole, wherein the through hole penetrates between the first face and a face other than the first face.
9. The electrode according to claim 7, wherein the member has a through hole, wherein the through hole penetrates between the first face and a face other than the first face.
10. An electrode kit comprising: the electrode according to claim 1; and a holder connected to a power supply, wherein the other end of each of the conductors is electrically connected to the holder, and power is supplied to each of the conductors from the power supply.
11. An electrode kit comprising: the electrode according to claim 2; and a holder connected to a power supply, wherein the other end of each of the conductors is electrically connected to the holder, and power is supplied to each of the conductors from the power supply.
12. An electrode kit comprising: the electrode according to claim 3; and a holder connected to a power supply, wherein the other end of each of the conductors is electrically connected to the holder, and power is supplied to each of the conductors from the power supply.
13. An electrode kit comprising: the electrode according to claim 4; and a holder connected to a power supply, wherein the other end of each of the conductors is electrically connected to the holder, and power is supplied to each of the conductors from the power supply.
14. The electrode according to claim 2, wherein the one or more protruding parts are arranged on the outer circumferential side from each of the conductors protruded in the first face.
15. The electrode according to claim 3, wherein the one or more protruding parts are arranged on the outer circumferential side from each of the conductors protruded in the first face.
16. The electrode according to claim 4, wherein the one or more protruding parts are arranged on the outer circumferential side from each of the conductors protruded in the first face.
17. The electrode according to claim 5, wherein the one or more protruding parts are arranged on the outer circumferential side from each of the conductors protruded in the first face.
18. The electrode according to claim 6, wherein the one or more protruding parts are arranged on the outer circumferential side from each of the conductors protruded in the first face.
19. The electrode according to claim 2, wherein the member has a through hole, wherein the through hole penetrates between the first face and a face other than the first face.
20. The electrode according to claim 3, wherein the member has a through hole, wherein the through hole penetrates between the first face and a face other than the first face.
Description
BRIEF DESCRIPTION OF DRAWINGS
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DETAILED DESCRIPTION OF EMBODIMENTS
[0057] Embodiments of an electrode and an electrode kit will be described below in detail with reference to the drawings. Note that, in the present specification, members having similar functions are labeled with the same or similar references. Further, for the members labeled with the same or similar references, duplicated description may be omitted.
[0058] Further, the position, the size, the range, or the like of each configuration illustrated in the drawings do not always represent the actual position, the actual size, the actual range, or the like for easier understanding. Thus, the disclosure in the present application is not necessarily limited to the position, the size, the range, like disclosed in the drawings.
First Embodiment of Electrode
[0059] Electrodes 1A to 1C according to a first embodiment will be described with reference to
[0060] The electrode 1A according to the first embodiment is used when electroporation is performed on cells. The electrode 1 at least has at least two or more conductors 2 (in the following description, two or more conductors may be simply referred to as conductors) and a member 3.
[0061] The conductors 2 are held in the member 3 described later. Further, the conductors 2 are electrically connected to an external power supply and used to perform electroporation on cells by using electric pulses supplied from the power supply. The conductor 2 may be any conductor as long as it can conduct electric pulses supplied from the power supply, and the shape or the material of the conductor 2 is not particularly limited. The shape of the conductor 2 may be, for example, a plate shape, a bar shape, or the like. In the example illustrated in
[0062] The member 3 holds the conductors 2 and exposes at least one ends of the conductors 2 from the surface of the member 3 so that the conductors 2 can perform electroporation on cells. Further, the other ends of the conductors 2 held by the member 3 may be in any form as long as they can be electrically connected to the external power supply and may or may not be exposed from the surface of the member 3. Note that, in the present specification, the face where one ends of the conductors 2 that perform electroporation on adherent cells of the member 3 (hereafter, which may be referred to as end(s) 21) is exposed is defined as the first face 31, and the face opposite to the first face 31 is defined as a second face 32.
[0063] In the example illustrated in
[0064] Although the member 3 has a cylindrical shape in the example illustrated in
[0065] Further, the conductors 2 held by the member 3 are required to have a positive polarity and a negative polarity when electroporation is performed. Therefore, while the member 3 holds at least two conductors 2, this does not limit that the member 3 holds three or more conductors 2. The number of conductors 2 held by the member 3 can be, for example, two, three, four, or the like. Note that, in the conventional electrode with legs, the three conductors 81 are coated with the insulator 82, respectively, as illustrated in
[0066] The length by which the end 21 of each conductor 2 is exposed from the first face 31 (in other words, the length of the conductor 2 protruding from the first face 31) is not particularly as limited long as electroporation can be performed on cells. While electroporation can be performed as long as the first face 31 and the end faces of the ends 21 of the conductors 2 are on the same level, it is preferable that the ends 21 of the conductors 2 protrude from the first face 31. When electroporation is performed, the efficiency thereof varies in accordance with the amount of electroporation buffer (hereafter, which may be referred to as buffer), the surface area of the conductor 2 exposed from the first face 31, or the like. Therefore, to achieve desired efficiency of electroporation in accordance with the situation where the electrode 1A according to the first embodiment is used, the surface area of the conductor 2 exposed from the first face 31 can be designed as appropriate.
[0067] The member 3 has at least one or more protruding parts 34. Each protruding part 34 is provided to the first face 31 of the member 3 and protrudes in the axis L direction of the member 3. In the present specification, the axis (L) direction of the member 3 means the direction of the center axis of a prism when the member 3 has a prism shape such as a circular prism shape or a polygonal prism shape, for example. Note that, when the shape of the member 3 is not a prism, the direction substantially perpendicular to the first face 31 may be the axis direction. Further, when two or more conductors 2 are held by the member 3 so as to be substantially parallel to each other, it can also be said that the axis direction is a direction parallel to the conductors 2.
[0068] Further, the length by which the protruding part 34 protrudes in the axis L direction from the first face 31 is longer than the length by which the end 21 of each conductor 2 is exposed from the first face 31. Thus, when the electrode 1A according to the first embodiment is used to perform electroporation on adherent cells adhered to a substrate or the like, since the top of the protruding part 34 is located on the substrate side from the ends 21 of the conductors 2 exposed in the first face 31, the ends 21 of the conductors 2 are not in contact with the adherent cells. Therefore, this prevents adherent cells from being significantly damaged due to supply of electric pulses. Note that the electrode 1A disclosed in the present application can be suitably used for electroporation on adherent cells in an adhered state and can also be used for electroporation on other cells such as floating cells. In such a case, the length by which the protruding part 34 protrudes in the axis L direction from the first face 31 can be adjusted as appropriate.
[0069] Further, in the electrode 1A according to the first embodiment, because the member 3 has the protruding parts 34, the protruding parts 34 come into contact with a substrate or the like and keep the attitude of the electrode 1A when the electrode 1A is installed to the substrate or the like. Therefore, this makes it easier to maintain the conductors 2 exposed in the first face 31 of the member 3 at a predetermined position. In the conventional electrode with legs, it is not possible to provide legs to a part other than the tip of the electrode because of the structure thereof. In contrast, in the electrode 1A, since the member 3 holds the conductors 2, it is possible to provide the protruding parts 34 to the first face 31. In the example illustrated in
[0070] The number of protruding parts 34, the shape of the protruding part 34, and the place to arrange the protruding parts 34 are not particularly limited as long as the conductors 2 are maintained stably at places where the electrode 1A is installed after installation of the electrode 1A. Since the protruding parts 34 protrude from the first face 31, when the electrode 1A is put into the buffer, air may remain in the space surrounded by the first face 31 and the protruding part 34. If air remains, this may cause a problem with electrical conduction between the conductors 2 facing each other. It is therefore preferable to design the protruding parts 34 so that the air around the first face 31A of the member 3 does not remain when the electrode 1A is put into the buffer. In the example illustrated in
[0071] Further, a slope part 36 may be provided to the first face 31 to facilitate air to move from the first face 31 to outside.
[0072] Note that, in the example illustrated in
[0073] Further, in terms of easier adjustment of the surface area of the conductors 2 exposed from the first face 31, the slope part 36 may be formed to a portion spaced apart from the conductors 2 in the first face 31 interposed between the conductors 2 facing each other. While the slope part 36 is formed in contact with the conductors 2 in the example illustrated in
[0074] The protruding parts 34 may be arranged between a plurality of conductors 2. However, since electric pulses are supplied to the conductors 2, it is preferable not to arrange the protruding parts 34 between the conductors 2 exposed in the first face 31 so as to less affect the electric field. Further, it is more preferable to arrange the protruding parts 34 on the outer circumferential side from the conductors 2 exposed in the first face 31. Because the protruding parts 34 are arranged outside the conductors 2, when the electrode 1A is installed to a substrate or the like, a contact portion between the protruding parts 34 and the substrate or the like is increased, and the conductors 2 can be maintained stably in the place where the electrode 1A is installed. Further, when the protruding parts 34 are arranged outside the conductors 2, it is possible to increase the distance between any points of the protruding parts 34. Thus, when the electrode 1A is installed to the substrate or the like, the conductors 2 can be maintained stably in the place where the electrode 1A is installed.
[0075] Note that, as described above, in terms of electrical conduction between the conductors 2 facing each other, it is preferable that no air remain between the conductors 2 facing each other. Therefore, as illustrated in
[0076] The electrodes 1A to 1C according to the first embodiment achieve the following advantageous effects.
[0077] (1) Since the member 3 of the electrodes 1A to 1C has the protruding parts 34 on the first face 31, when each of the electrodes 1A to 1C is installed to a substrate or the like, the conductors 2 can be maintained stably in the place where each of the electrodes 1A to 1C is installed. Further, the conventional electrode is fabricated by first coating the conductor 81 with the insulator 82 and then holding the conductor 81 coated with the insulator 82 in a holding portion. Thus, as illustrated in
[0078] (2) The length by which the protruding part 34 protrudes in the axis L direction from the first face 31 is longer than the length by which the end 21 of the conductor 2 is exposed from the first face 31. Therefore, when each of the electrodes 1A to 1C is used for electroporation on adherent cells, the electroporation can be performed without the conductors 2 being in contact with the adherent cells.
[0079] (3) When the slope part 36 is provided to the first face 31 interposed between the conductors 2 facing each other, air moves to the outer circumferential side of the member 3 along the slope part 36 even when the electrode is put into the buffer. Therefore, a likelihood of air remaining between the conductors 2 is reduced.
[0080] (4) Since each of the electrodes 1A to 1C can be handled as a separate member from a power supply, the electrodes 1A to 1C can also be used as a disposable electrode.
Second Embodiment of Electrode
[0081] Electrodes 1D to 1F according to the second embodiment will be described with reference to
[0082] The electrodes 1D to 1F according to the second embodiment differ from those of the first embodiment in that the member 3 has the through hole 38 and/or the recesses 37 in the first face 31. Therefore, for the electrodes 1D to 1F according to the second embodiment, features different from those of the first embodiment will be mainly described, and duplicated description for the features that have already been described in the first embodiment will be omitted. Thus, it is apparent that, even when not explicitly described in the second embodiment, the features that have already been described in the first embodiment can be employed.
[0083] The recess 37 is provided in the first face 31, is recessed in the axis L direction of the member, and penetrates through in the side face 33 direction of the member 3. Further, the through hole 38 penetrates through the first face 31 and a face other than the first face 31. Because the member 3 has the recesses 37 and/or the through hole 38, the space surrounded by the first face 31 and the protruding parts 34 is connected to outside of the member 3 via the recesses 37 and/or the through hole 38. Therefore, when each of the electrodes 1D to 1F is put into the buffer, air of the space surrounded by the first face 31 and the protruding parts 34 can be more reliably discharged through the recesses 37 and/or the through hole 38.
[0084] The number of recesses 37 and through holes 38 and the arrangement of the recesses 37 and the through holes 38 provided in the first face 31 are not particularly limited as long as they can connect the space surrounded by the first face 31 and the protruding parts 34 to outside, and the member 3 may have any one of or both of the recess 37 and the through hole 38.
[0085] In the example of the electrodes 1D and 1E illustrated in
[0086] The electrodes 1D to 1F according to the second embodiment synergistically achieve the following advantageous effects in addition to the advantageous effects achieved by the electrodes 1A to 1C according to the first embodiment.
[0087] (1) The member 3 has the recess(s) 37 and/or the through hole 38, and this enables more reliable discharge of air of the space surrounded by the first face 31 and the protruding parts 34. Therefore, electroporation can be performed without any contact of the conductors 2 exposed in the first face 31 with air.
[0088] (2) In the case of the electrodes 1A to 1C according to the first embodiment, the length by which the protruding part 34 protrudes in the axis L direction from the first face 31 is required to be designed in accordance with use. For example, when electroporation is performed on adherent cells, the length by which the protruding part 34 protrudes in the axis L direction from the first face 31 is relatively shorter. Accordingly, the size of each clearance 35, which is formed by the protruding parts 34 being spaced apart from each other, depends on the width of the protruding parts 34. In contrast, in the electrodes 1D to 1F according to the second embodiment, the size of each recess 37 and/or each through hole 38 can be decided regardless of the length by which the protruding part 34 protrudes, and this improves flexibility of design.
Embodiment of Electrode Kit
[0089] An electrode kit has at least any one of the electrodes 1A to 1F and a holder. The electrodes 1A to 1F of the electrode kit have already been described in the above embodiments. Thus, duplicated description will be omitted for the electrodes 1A to 1F.
[0090] The holder electrically connects an external power supply to the conductors 2 held by the member 3 of the electrodes 1A to 1F and is used to supply power from the external power supply to the conductors 2. The holder is not particularly limited as long as it can electrically connect the external power supply and the conductors 2 of the member 3 to each other. For example, in the example of the electrode 1A illustrated in
[0091] The electrode kit according to the embodiment is characterized in that the electrode kit includes the electrode 1 according to the above embodiments. Therefore, the same advantageous effects as those of the electrodes 1A to 1F according to the above embodiment are achieved.
[0092] Note that the present invention is not limited to the embodiment described above. Within the scope of the present invention, any combination of respective embodiments described above, modification of any component of each embodiment, or omission of any component of each embodiment is possible.
EXAMPLES
Example 1
[0093] Electroporation was performed on adherent cells by using the electrode 1. The materials, devices, and procedures used will be described below. Further,
TABLE-US-00001 TABLE 1 size (mm) a 12.5 b 14.5 c 5.5 d 2.5 e 3 f 1.75 g 2 h 0.1 i 0.6 j 8 k 22
Procedure
[0101] 1. Plasmid DNA was added to the buffer to give plasmid DNA concentration of 5 g/L.
[0102] 2. The wells to which cells were adhered were washed by the buffer.
[0103] 3. The buffer of 250 L including the plasmid DNA prepared in 1. was added to the wells.
[0104] 4. The electrode 1 was arranged in the well so that the cells were not detached therefrom.
[0105] 5. Electroporation was performed. In the electroporation, the electrode 1 was connected to the electroporator such that the center conductor 2 was positive, and two conductors on both sides were negative, and electric pulses were supplied to the cells in a damped wave (Decay (V)) mode. The poration pulse (Pp) and the driving pulse (Pd) in the damped wave mode were set as follows. [0106] Pd setting value [0107] Voltage: 250V [0108] Pulse duration (Pon): 10 ms [0109] Pulse interval (Poff): 50 ms [0110] Pd setting value [0111] Voltage: 30V [0112] Pulse duration (Pon): 50 ms [0113] Pulse interval (Poff): 50 ms [0114] Number of pulses: 5 [0115] Pattern: +/ [0116] Capacitance: 940 F
[0117] 6. The electrode 1 was taken out of the well, and the well plate was incubated for 10 minutes in a CO.sub.2 incubator for recovering the cells.
[0118] 7. The buffer was removed from the well, and a pre-incubated cell culture medium for post-culture was gently added thereto.
[0119] 8. The well plate was moved to the CO.sub.2 incubator, and post-culture was started.
[0120] 9. After 48 hours of the post-culture, the well plate was taken out of the CO.sub.2 incubator, and the cells were observed by using the fluorescent microscope.
Comparative Example 1
[0121] Comparative example 1 is the same as Example 1 except that the electrode used in electroporation was the conventional electrode with legs (LF513-5, BEX CO., LTD.).
[0122] With respect to the electrode 1 used in Example 1, when the electrode 1 was arranged in the well, the conductors 2 was maintained at a predetermined position stably due to the protruding parts 34 of the member 3. Further, since the tips of the protruding parts 34 protrude more than the ends 21 of the conductors 2 exposed in the first face 31 of the member 3 of the electrode 1, the conductors 2 did not come into contact with cells. Therefore, in Example 1, with the use of the electrode 1, the conductors 2 was maintained at a predetermined position, and the electroporation on adherent cells was stably performed. In contrast, in Comparative example 1, since the conventional electrode with legs was used, it was difficult to stably maintain the electrode with legs at a predetermined position.
[0123] After the electroporation was performed, the post-cultured adherent cells were observed by using the fluorescent microscope.
INDUSTRIAL APPLICABILITY
[0124] The use of the electrode and the electrode kit disclosed in the present application makes it easier to arrange the electrode at a predetermined position when performing electroporation on cells. Therefore, the electrode and the electrode kit are useful for business entities that handle an electrode used for performing electroporation on cells.
LIST OF REFERENCES
[0125] 1, 1A to 1F [0126] electrode [0127] 2 conductor [0128] 21, 22 end [0129] 3 member [0130] 31 first face [0131] 32 second face [0132] 33 side face [0133] 34 protruding part [0134] 35 clearance [0135] 36 slope part [0136] 36a slope face [0137] 37 recess [0138] 38 through hole [0139] 81 conductor [0140] 82 insulator [0141] 83 leg [0142] IL virtual line [0143] IR outer circumferential virtual region [0144] L axis