WO2014194859A1 - 一种高密度分布式立体电极装置 - Google Patents

一种高密度分布式立体电极装置 Download PDF

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Publication number
WO2014194859A1
WO2014194859A1 PCT/CN2014/079391 CN2014079391W WO2014194859A1 WO 2014194859 A1 WO2014194859 A1 WO 2014194859A1 CN 2014079391 W CN2014079391 W CN 2014079391W WO 2014194859 A1 WO2014194859 A1 WO 2014194859A1
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electrode
electrodes
polarity
electrode array
array
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PCT/CN2014/079391
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English (en)
French (fr)
Inventor
戴爱德华
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苏州文曲微生物系统有限公司
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Priority to EP14806985.9A priority Critical patent/EP3006552B1/en
Priority to US14/896,902 priority patent/US20160137966A1/en
Publication of WO2014194859A1 publication Critical patent/WO2014194859A1/zh
Priority to US16/192,077 priority patent/US11421196B2/en
Priority to US17/862,707 priority patent/US20220340855A1/en

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
    • C12M35/02—Electrical or electromagnetic means, e.g. for electroporation or for cell fusion
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/87—Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation

Definitions

  • the present invention relates to cell electroporation techniques, and more particularly to a stereoelectrode device for electroporation.
  • electropores are areas of increased permeability caused by local rupture of the cell membrane caused by high voltage electric fields. These pores are present for a short period of time, but are sufficient for the entry of macromolecules such as plasmid DNA molecules into cells. Although cells can tolerate the formation of these pores, if the pores formed are too large, the process of production and the molecules introduced thereby may also kill the cells.
  • Electroporation was first performed with the simplest parallel-plate capacitors, and a substantially uniform electric field was generated between the electrodes opposite each other.
  • the cell suspension ready for electroporation is mixed with the molecules that the operator wishes to introduce into the cells, placed between the two electrodes, and one or more short-time high-voltage electric field pulses are passed through the electrodes to achieve molecular passage.
  • the spacing between the parallel plate electrodes is large, and the required voltage is usually as high as several thousand volts, which inevitably causes a cathodic effect and causes great damage to the cells.
  • the planar electrode that appears later solves the negative effects of excessive voltage, but the amount of cells that can be processed each time is small and is not suitable for high-throughput experimental operations.
  • Stereoscopic electrodes are easy to penetrate into tissues and living bodies, and are often used for electroporation in clinical directions such as tumors or living tissues. The electroporation efficiency is not high, and cell electroporation in vitro, such as for suspension cells or adherent cells, has not been used.
  • the technical problem to be solved by the present invention is to provide a high-density distributed stereo electrode device which is simple in structure and easy to manufacture and manufacture.
  • the present invention provides a high-density distributed stereoscopic electrode device comprising an electrode array and an electrode fixing assembly, the electrode array being fixed on the electrode fixing assembly, the electrode array comprising a plurality of electrodes,
  • the plurality of electrodes are divided into at least two groups, and the two groups of electrodes respectively apply an electric pulse of a first polarity and an electric pulse of a second polarity on a time period, wherein the first polarity is different from the second polarity, and An electrode corresponding to the second polarity electrical pulse is distributed around the electrode corresponding to the one polarity electrical pulse.
  • the plurality of electrodes in the electrode array are arranged in a regular polygon, and the distance between adjacent two electrodes in the electrode array is equal.
  • the shape of the electrode array is a regular hexagon composed of a plurality of equilateral triangles, the electrodes being located at the vertices of the equilateral triangle.
  • the first polarity is a positive electrode and the second polarity is a negative electrode.
  • the first polarity is a negative electrode and the second polarity is a positive electrode.
  • the diameter of the electrode is 0.01-1.2 mm, the distance between the center points of the two adjacent electrodes is 0.1-2.4 mm, and the number of the electrodes is greater than 5.
  • the roots are preferably 19 or more.
  • the material of the electrode is preferably stainless steel.
  • the electrode has a diameter of 0.1-0.4 mm, the distance between the center points of the two adjacent electrodes is 0.2-1.5 mm, and the number of the electrodes is greater than 36 .
  • the diameter of the electrode is preferably 0.3 mm, the distance between the center points of two adjacent electrodes is preferably 1 mm, and the number of electrodes is preferably 37.
  • the electrode fixing assembly includes an electrode connecting circuit board and an electrode positioning board, and the electrode connecting circuit board connects the electrodes to which electric pulses of the same polarity are applied through a line thereon.
  • the electrode is inserted into the electrode positioning plate.
  • the electrodes in the electrode array are divided into groups, and the electrodes of the same group are only applied with electric pulses of the same polarity, and one of the electrode arrays is applied to the positive electrode.
  • the electric pulse, the other group applies an electric pulse to the negative electrode, and the other group applies an electric pulse to the positive electrode, and the other group applies an electric pulse to the negative electrode, so that the obtained electric field is superposed to obtain a uniform electric field.
  • the invention has the beneficial effects of the high-density distributed stereo electrode device of the invention, which adopts the group-multiplexed electrode, can make up for the electric field unevenness caused by the stereo electrode to the greatest extent, and can process the milliliter-level cells in a single time. It can be used in both orifice plate and flow device.
  • the electrode spacing is small, the required electroporation voltage is small, avoiding high voltage damage to cells, low cost, and it is a high-throughput, high-efficiency cell electroporation. Device.
  • FIG. 1 is a schematic structural view of a preferred embodiment of a high density distributed stereoscopic electrode device in accordance with the present invention
  • Figure 2 is a front elevational view of the high density distributed stereoscopic electrode assembly of Figure 1;
  • Figure 3 is a plan view of the high density distributed stereoscopic electrode device of Figure 1;
  • FIG. 4 is a topological structural view of an electrode array in the high-density distributed stereoscopic electrode device shown in FIG. 1;
  • Electrode array The components corresponding to the respective marks in the drawing are as follows: 1. Electrode array, 2. Electrode positioning plate, 3. Electrode connection circuit board.
  • Embodiment 1 is a diagrammatic representation of Embodiment 1:
  • the present invention provides a high-density distributed electrode device comprising an electrode array 1 and an electrode fixing assembly, the electrode array 1 being fixed on the electrode fixing assembly.
  • the electrode array 1 is composed of solid 37 cylindrical electrodes arranged in such a manner that all the electrodes are arranged in a regular hexagonal structure, and the spacing between adjacent two electrodes is equal, between adjacent two electrodes.
  • the spacing is the length of the side, dividing the interior of the regular hexagon into a number of small equilateral triangle elements, and placing an electrode at the apex of each equilateral triangle, that is, all the electrodes are divided into I, II, III
  • the vertices of each equilateral triangle unit belong to I, II, and III, respectively.
  • the inner diameter of the regular hexagon of the electrode array 1 matches the hole of the perforated plate, and the distance between the center points of the two adjacent electrodes is 1 mm.
  • the spacing between the two electrodes affects the voltage at the time of electroporation and can be adjusted as needed.
  • the distance from the bottom of the orifice plate is 0.1 mm-lmm.
  • the diameter of the electrode is 0.3 mm, and the diameter of the electrode is too large or too small to affect the electroporation effect.
  • the diameter is too large, the effective electric field area of electroporation is reduced, and the amount of cells capable of electroporation is reduced, which is disadvantageous for high-throughput cell electroporation. If the diameter is too small, the electrode is easily bent, which greatly increases the manufacturing cost.
  • the material of the electrode can be selected from any conductive metal and other conductive materials, wherein the stainless steel electrode is an excellent material as an electrode.
  • the stainless steel material has good biocompatibility, is easy to clean, is not easily oxidized, is easy to be made into a long electrode, can be mass-produced and reused many times without affecting its conductive properties.
  • the electrode fixing assembly includes an electrode positioning plate 2, an electrode connection circuit board 3, the electrodes are positioned through the electrode positioning plate 2, and are connected to the electrode connection circuit board 3, and the electrode connection circuit board 3 is arranged through the line arrangement thereon Electrodes applying the same polarity are connected together. Since the electrode is relatively long and the electroporation technique requires high electrode pitch accuracy, the electrode positioning plate 2 is used to position the electrode during the electroporation experiment. Since the electrode positioning plate 2 has a thickness of about 1 cm, the electrode can be restrained and positioned for a long distance, so that the position of the electrode reaching the bottom of the orifice plate can be accurately controlled.
  • the support structure of the high-density distributed stereo electrode device can be easily expanded, and can form an electrode network of any combination array of 2*2, 1*4, 12*8, etc., so that the flexible combination can maximize the compatibility with the porous plate structure. User-friendly.
  • the electrode connection circuit board 3 connects the same group of electrodes together by soldering, conductive glue or other electrical connection. It can be a PCB board or any device that can perform the specified line connection.
  • the electrode positioning plate 2 is used for electrode positioning, and the electrode is elongated and easily bent. Therefore, in the embodiment shown in Figs. 1-2, the electrode positioning plate 2 is arranged to position the electrode at a substantially central position of the electrode. The use of the electrode positioning plate 6 can reduce the inconsistency of the electrode spacing, thereby improving the electric field uniformity.
  • the high-density distributed stereoscopic electrode device may be a single-hole device, and may be combined into a group of four or a group of 96 to match the commonly used porous plate structure of the organism, and even more.
  • the electroporation method of the high-density distributed stereo electrode device in this embodiment is: when electroporation, I first uses I as the positive electrode, II, III as the negative electrode, and then II as the positive electrode, and I and III as the negative electrode. The pulse is then applied with III as the positive electrode and I, II as the negative electrode.
  • the uniform electric field strength is controlled to be the optimal electroporation voltage for electroporation
  • cells in the entire effective region can be electroporated to the greatest extent. It can be seen that the packet multiplexing compensates for the electric field unevenness caused by a single packet and improves the electroporation efficiency. It can be judged that the electroporation efficiency of the combined electric field is much higher than that of the conventional electroporation.
  • the high density distributed stereoelectrode device is capable of suspending or adhering electroporation of many cell lines.
  • the electroporation rate that is, the greater the fluorescence intensity of cells of the same density, the higher the electroporation efficiency.
  • the high density distributed stereoscopic electrode device according to the present invention can be applied to a flow device.
  • the high-density distributed stereoscopic electrode device is placed in a flow environment, and the cells uniformly distributed with the flowing liquid pass through the electrode array, and are controlled by flow rate and pulse stimulation to receive the optimal electroporation stimulation.
  • Conditions, electroporation stimulation conditions include pulse voltage amplitude, pulse width, pulse interval, number of pulses, and electrode exchange control. In a continuous flow system, there is no need for "period processing" coordination between pulse and cell flow, except that the time at which the pulse is applied to the electrode is coordinated with the time at which the cell begins to flow between the electrodes.
  • the high-density distributed stereoscopic electrode device has the advantages of continuous flow electroporation, in particular, high-throughput electroporation of cells in a sterile closed system, and ensuring that each cell receives the optimal number of times.
  • Embodiment 2 is a diagrammatic representation of Embodiment 1:
  • the high-density distributed electrode device in this embodiment is basically the same as the first embodiment, and the difference is in the arrangement of the electrode arrays.
  • the arrangement of the electrode arrays is such that all the electrodes are arranged in a regular quadrilateral structure, the spacing between the adjacent two electrodes is equal, and the spacing between adjacent two electrodes is the side length, and the regular quadrilateral interior is Divided into a number of small square cells, each electrode is placed at the apex of the square, that is, all the electrodes are divided into I, II, III, IV 4 groups, the vertices of each square unit belong to
  • the arrangement of the electrode arrays of the present example can be expressed in a slight variation in connection with the case of the first embodiment shown in FIG. 4, and thus will not be illustrated by the drawings.
  • Embodiment 3 is a diagrammatic representation of Embodiment 3
  • the present invention provides a high-density distributed electrode device.
  • the other structures are the same as those in the first embodiment.
  • the arrangement of the electrode arrays is such that all the electrodes are arranged in a regular hexagonal structure, and the spacing between the adjacent two electrodes is equal.
  • the spacing between two adjacent electrodes is the side length, and the plane is formed into a number of small regular hexagonal units, and the apex of each regular hexagonal unit is placed with one electrode, that is, all the electrodes are divided into I, II, III, IV, V, VI
  • the vertices of each regular hexagonal unit belong to I, II, III, IV, V, and VI, respectively, and the electrodes of one group are used as the positive poles, and the electrodes of the other group are used as the negative electrodes, and electric pulses are alternately applied.
  • the arrangement of the electrode arrays of the present example can be expressed in a slight variation in connection with the case of the first embodiment shown in FIG. 4, and thus will not be illustrated by the drawings.

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Abstract

本发明公开了一种高密度分布式立体电极装置，包括电极阵列和电极固定组件，电极阵列固定在电极固定组件上，所述电极阵列包括多个电极，所述多个电极上按时间周期分别施加第一极性的电脉冲和第二极性的电脉冲，其中，所述第一极性的电脉冲对应的电极的周围分布的都是第二极性的电脉冲对应的电极。通过上述方式，本发明提供的一种高密度分布式立体电极装置，所述装置采用分组复用的电极，能够最大程度的弥补立体电极造成的电场不均，能够单次处理毫升级的细胞，在孔板和流式装置中均能使用，能避免了高电压对细胞的伤害，成本低，是一种高通量、高效率的细胞电穿孔装置。

Description

一种高密度分布式立体电极装置
技术领域
本发明涉及细胞电穿孔技术，特别是涉及一种用于电穿孔的立体电极装置。
背景技术
自从1970年代电穿孔技术就被用于将分子插入到动物细胞或植物细胞内，研究者证实，细胞暴露于短暂持续的高压电场中能使胞膜上形成通道，大分子如蛋白质和DNA可通过该通道进入细胞内。这些通道被称为电孔洞，是由高压电场造成的细胞膜局部破裂引起的通透性升高区。这些孔洞的存在时间虽然短暂，但足以满足大分子如质粒DNA分子进入细胞的需要。细胞虽然能够耐受这些孔洞的形成，但如果形成的孔洞过多过大，其产生的过程以及由此导入的分子也可能会杀死细胞。
最早是用最简单的带平行板电容器进行电穿孔，彼此相对的电极之间可产生基本均匀的电场。将准备进行电穿孔的细胞悬液与操作者希望将其导入到细胞内的分子混合置于两个电极之间，在电极上通以一次或多次短时间的高压电场脉冲，从而达到分子通过电穿孔导入到细胞内的效果。但平行板电极之间的间距较大，所需要的电压通常都高达数千伏，无可避免的会产生阴极效应，对细胞产生巨大伤害。
之后出现的平面电极虽然解决了电压过高带来的负面作用，但是每次能够处理的细胞量很小，不适合于高通量的实验操作。立体式电极容易刺入组织和活体，多用于肿瘤或活体组织等临床方向的电穿孔，其电穿孔效率并不高，对于体外的细胞电穿孔，如针对悬浮细胞或贴壁细胞，都没有过相关的报道。
发明内容
本发明主要解决的技术问题是提供一种结构简单、易于加工制造的高密度分布式立体电极装置。
为解决上述技术问题，本发明提供一种高密度分布式立体电极装置，包括电极阵列和电极固定组件，所述电极阵列固定在所述电极固定组件上，所述电极阵列包括多个电极，所述多个电极分为至少两组，两组电极上按时间周期分别施加第一极性的电脉冲和第二极性的电脉冲，其中，第一极性与第二极性不同，并且第一极性的电脉冲对应的电极的周围分布第二极性的电脉冲对应的电极。
在本发明的一个较佳实施例中，所述电极阵列中的多个电极按正多边形排列，所述电极阵列中相邻的两个电极之间的距离相等。
在本发明的一个较佳实施例中，所述电极阵列的形状为由若干等边三角形组成的正六边形，所述电极位于所述等边三角形的顶点上。
在本发明的一个较佳实施例中，所述第一极性为正极，所述第二极性为负极。
在本发明的一个较佳实施例中，所述第一极性为负极，所述第二极性为正极。
在本发明的一个较佳实施例中，所述电极的直径为0.01-1.2mm,所述两个相邻电极中心点之间的距离为0.1-2.4mm，所述电极的个数为大于5根，优选为19根以上。电极的材料优选为不锈钢。
在本发明一个更优选的实施例中，述电极的直径为0.1-0.4mm,所述两个相邻电极中心点之间的距离为0.2-1.5mm，所述电极的个数为大于36根。电极的直径优选为0.3mm，两个相邻电极中心点之间的距离最好为1mm，电极的个数优选为37根。
在本发明的一个较佳实施例中，所述电极固定组件包括电极连接电路板和电极定位板，电极连接电路板通过其上的线路将被施加同一极性的电脉冲的电极连接到一起，所述电极插入到所述电极定位板中。
在本发明的一个较佳实施例中，所述电极阵列中的电极分为若干组，所述同一组的电极只施加同一极性的电脉冲，以所述电极阵列的其中一组为正极施加电脉冲，其余组为负极施加电脉冲，再以另一组为正极施加电脉冲，其余组为负极施加电脉冲，如此交替，将得到的电场叠加得到均匀的电场。
本发明的有益效果是:本发明的高密度分布式立体电极装置，该电极装置采用分组复用的电极，能够最大程度的弥补立体电极造成的电场不均，能够单次处理毫升级的细胞，在孔板和流式装置中均能使用，电极间距小，所需电穿孔电压就很小，避免了高电压对细胞的伤害，成本低，是一种高通量、高效率的细胞电穿孔装置。
附图说明
图1是根据本发明的高密度分布式立体电极装置的一较佳实施例的结构示意图；
图2是图1所示高密度分布式立体电极装置的正视图；
图3是图1所示高密度分布式立体电极装置的俯视图；
图4是图1所示高密度分布式立体电极装置中电极阵列的拓扑结构图；
附图中各标记对应的部件如下:1、电极阵列，2、电极定位板，3、电极连接电路板。
具体实施方式
下面结合附图对本发明的较佳实施例进行详细阐述，以使本发明的优点和特征能更易于被本领域技术人员理解，从而对本发明的保护范围做出更为清楚明确的界定。
实施例一:
请参阅图1、图2、图3和图4，本发明提供一种高密度分布式电极装置，包括电极阵列1和电极固定组件，所述电极阵列1固定在所述电极固定组件上。
所述电极阵列1由实心的37根圆柱体电极组成，排列规则为所有的电极排列成正六边形结构，相邻的两个电极之间的间距都相等，以相邻两个电极之间的间距为边长，将正六边形内部分成若干小的等边三角形单元，每个等边三角形的顶点放置一个电极，即所有的电极分为I、II、III 3组，每个等边三角形单元的顶点分别属于I、II、III。
所述电极阵列1的正六边形的内径与多孔板的板孔相匹配，两个相邻电极中心点之间的距离为1mm。两个电极之间的间距影响电穿孔时的电压，可以根据需要调整。在电极插入孔板中，离孔板底部的距离为0.lmm-lmm。
所述电极的直径为0.3mm，直径过大或者过小都会影响电穿孔效果。直径过大时，电穿孔的有效电场面积会减少，使能够电穿孔的细胞量减少，不利于高通量的细胞电穿孔。直径过小，电极容易弯曲，使制作成本大大增加。
所述电极的材料可选择任意导电金属和其他导电材料，其中不锈钢电极是作为电极的极好的材料。所述不锈钢材料具有良好的生物兼容性，容易清洗，不易氧化，容易做成较长的电极，能够大批量生产并多次重复使用而不影响其导电特性。
所述电极固定组件包括电极定位板2、电极连接电路板3，电极穿过电极定位板2而被定位，并连接至电极连接电路板3，电极连接电路板3通过其上的线路安排将被施加同一极性的电极连接到一起。由于所述电极比较长，而电穿孔技术对于电极间距精度要求很高，因此在进行电穿孔实验时采用了电极定位板2对所述电极进行定位。由于电极定位板2的厚度约1cm，能够对所述电极进行长距离的约束和定位，因此能够很准确的控制电极到达孔板底部的位置。高密度分布式立体电极装置的支撑结构能够轻松的扩展，可以组成2*2、1*4、12*8等任意组合阵列的电极网络，这样灵活多样的组合能够最大程度的兼容多孔板结构，方便用户使用。
所述电极连接电路板3将同一组的电极通过焊接方式、导电胶或其他电学连接方式连在一起。可以采用PCB线路板，也可以是任何能够完成规定线路连接的器件。所述电极定位板2用于电极定位，电极细长容易弯曲，因而在图1-2所示的实施例中，在电极的大致中部位置安排电极定位板2对电极进行定位。使用所述电极定位板6能降低电极间距的不一致性，从而提高电场均匀性。
所述高密度分布式立体电极装置可以为单孔的器件，可以组合成4个一组，也可以96个一组以配合生物常用的多孔板结构，甚至更多来使用。
本实施例中的高密度分布式立体电极装置的电穿孔方法为:在电穿孔时先以I作为正极，II、III作为负极施加电脉冲，再以II作为正极，I、III作为负极施加电脉冲，然后以III作为正极，I、II作为负极施加电脉冲。
对于某种特定的细胞，所加电场高于某一阈值的时候，细胞膜会出现开孔。当电场逐渐增加，细胞的死亡率会出现上升。为了保证细胞的高电穿孔率和低死亡率，需要将电场精确控制在电穿孔阈值电场。
只要控制所述均匀电场强度为电穿孔的最佳电穿孔电压，整个有效区域内的细胞都能最大程度的进行电穿孔。由此可见，分组复用弥补了单一分组造成的电场不均，提高了电穿孔效率，可以判断这种组合电场的电穿孔效率会比传统电穿孔效率高很多。
所述高密度分布式立体电极装置能够对很多细胞系进行悬浮或者贴壁的电穿孔。我们选用 7 HEK-293A、Hela, MCF-7、A-375、Neuro-2A、U251、C2C12、3T3-L1、CHO, MDCK,HL-60,HUVEC 12种细胞进行电穿孔，使用GFP分子作为标志物。如果细胞被电穿孔，GFP分子会进入细胞，并且在细胞内合成荧光物质，合成的荧光物质在荧光场下会发出绿色荧光，因此用荧光场中细胞的数量除以整个细胞数量可以得出细胞的电穿孔率，即同等密度细胞的荧光强度越大，电穿孔效率越高。
根据本发明的高密度分布式立体电极装置能应用在流式装置中。将所述高密度分布式立体电极装置置于流式的环境中，随流动液体均匀分布流动的细胞经过所述电极阵列时，经流速和脉冲刺激的控制，使之接收最佳的电穿孔刺激条件，电穿孔刺激条件包括脉冲的电压幅值、脉冲宽度、脉冲间隔、脉冲个数和电极交换控制。在持续流动系统中，除了在电极上施加脉冲的时间与细胞在电极之间开始流动的时间需要协调之外，脉冲与细胞流动之间不需要“期间处理”的协调。
根据本发明的高密度分布式立体电极装置既有持续流式电穿孔的优点，特别是能在无菌封闭系统内对细胞进行高通量电穿孔，又能保证每个细胞都受到最佳次数的脉冲和最均匀的电场，以提高电穿孔的效率，并减少死亡率。
实施例二:
本实施例中的高密度分布式电极装置与实施例一基本相同，区别在于电极阵列的排列方式。本实施例中，电极阵列的排列规则为所有的电极排列成正四边形结构，相邻的两个电极之间的间距都相等，以相邻两个电极之间的间距为边长，将正四边形内部分成若干小的正方形单元，每个正方形的顶点放置一个电极，即所有的电极分为I、II、III、IV 4组，每个正方形单元的顶点分别属于
I、II、III、IV，以其中一组的电极作为正极，其余组的电极作为负极，轮流施加电脉冲。本实例的电极阵列的排列方式可结合图4所示的实施例一的情况稍加变动来表示，因此不再采用附图示出。
实施例三:
本发明提供一种高密度分布式电极装置，其他结构与实施例一相同，电极阵列的排列规则为所有的电极排列成正六边形结构，相邻的两个电极之间的间距都相等，以相邻两个电极之间的间距为边长，将平面成若干小的正六边形单元，每个正六边形单元的顶点放置一个电极，即所有的电极分为I、II、III、IV、V、VI 6组，每个正六边形单元的顶点分别属于I、II、III、IV、V和VI，以其中一组的电极作为正极，其余组的电极作为负极，轮流施加电脉冲。本实例的电极阵列的排列方式可结合图4所示的实施例一的情况稍加变动来表示，因此不再采用附图示出。
以上所述仅为本发明的实施例，并非因此限制本发明的专利范围，凡是利用本发明说明书及附图内容所作的等效结构或等效流程变换，或直接或间接运用在其他相关的技术领域，均同理包括在本发明的专利保护范围内。

Claims (9)

1.一种高密度分布式立体电极装置，其特征在于，该装置包括电极阵列和电极固定组件，所述电极阵列固定在所述电极固定组件上，所述电极阵列包括多个电极，所述多个电极分为至少两组电极，两组电极上按时间周期分别施加第一极性的电脉冲和第二极性的电脉冲，其中，第一极性的电脉冲对应的电极的周围分布第二极性的电脉冲对应的电极，第一极性和第二极性相反。
2.根据权利要求1所述的装置，其特征在于，所述电极的直径为0.01-1.2mm，所述两个相邻电极中心点之间的距离为0.1-2.4mm，所述电极的个数为大于5根。
3.根据权利要求2所述的装置，其特征在于，所述电极的直径为0.1-0.4mm，所述两个相邻电极中心点之间的距离为0.2-1.5mm，所述电极的个数为大于36根。
4.根据权利要求3所述的装置，其特征在于，所述电极的直径为0.3mm，所述两个相邻电极中心点之间的距离为1mm，所述电极的个数为37根。
5.根据权利要求1所述的装置，其特征在于，所述电极阵列中的多个电极按正多边形排列，所述电极阵列中每相邻两个电极之间的距离相等。
6.根据权利要求5所述的装置，其特征在于，所述电极阵列的形状为由若干等边三角形组成的正六边形，所述电极位于所述等边三角形的顶点上。
7.根据权利要求1-6中任一项所述的装置，其特征在于，所述电极的材料为不锈钢。
8.根据权利要求1 -6中任一项所述的装置，其特征在于，所述电极固定组件包括电极连接电路板和电极定位板，所述电极连接电路板将被施加同一极性的电脉冲的电极连接到一起所述电极插入到所述电极定位板中。
9.根据权利要求1-6中任一项所述的装置，其特征在于，所述电极阵列中的电极分为三组或更多，同一组的电极施加同一极性的电脉冲，多组电极以按时间周期交替施加电脉冲，将得到的电场叠加得到均匀的电场，交替的方式为：以所述电极阵列的其中一组电极为正极施加电脉冲，其余组为负极施加电脉冲，然后再以另一组为正极施加电脉冲，其余组为负极施加电脉冲。
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