200300450 五、發明說明（1) 發明所屬之技術領域 本發明是有關於一種用以在培養基中培養細胞的細胞 培養裝置以及劑量反應之量測方法。 先前技術 通常為了量測細胞對直流電或誘導電流的劑量反應， 所採用的方法是讓細胞内的電流流量不同，或是對細胞施 以不同的電場，以讓細胞内產生的誘導電流流量不同，藉 以量測細胞劑量反應的差異，細胞的劑量反應反映出細胞 數量的變化、細胞内基因的變化、細胞内染色體的變化、 細胞功能的改變、細胞内化合物的改變等，有一種細胞保 存系統被設計用來量測細胞的反應，此系統包含一個培養 皿、培養欄、以及具有相等面積的培養薄膜。一般細胞會 在培養皿或類似的物品中培養，而被培養出來的細胞會被 移出培養孤、稀釋、分成預定的量，並放入細胞保存系統 内，另外不同組的細胞會被放到細胞保存系統中，以進行 不同的劑量實驗，然後用不同的電流在不同的時間流通， 以量測電流的變化。 在上述的方法中，實驗的時間會被分成對不同的劑量 有不同的時間，除非同時作比較，否則細胞的本質與細胞 的數量上無法1 0 0 %的確定，也就是因為有可能培養的細胞 會隨著時間繁殖，且本質會隨著周圍的條件改變，因此假 如這些點沒有被納入考慮，在不同情況下量測到的細胞劑 量反應會得到不正確的量測結果。因此以往為了解決這樣 的缺點，必須增加實驗裝置的數量，所以幾組相同細胞會200300450 V. Description of the invention (1) Technical field to which the invention belongs The present invention relates to a cell culture device and a method for measuring a dose response for culturing cells in a culture medium. In the prior art, in order to measure the dose response of a cell to a direct current or an induced current, the method used is to make the current flow in the cell different or apply different electric fields to the cell to make the induced current flow in the cell different. By measuring the difference in the dose response of cells, the dose response of cells reflects changes in the number of cells, changes in genes in cells, changes in chromosomes in cells, changes in cell functions, changes in compounds in cells, etc. Designed to measure cell response, this system includes a petri dish, culture column, and a culture membrane of equal area. Generally, cells will be cultured in a petri dish or the like, and the cultured cells will be removed from the culture sol, diluted, divided into a predetermined amount, and placed in the cell preservation system, and different groups of cells will be placed in the cells Save the system to perform different dose experiments, and then use different currents to flow at different times to measure changes in current. In the above method, the time of the experiment will be divided into different times for different doses. Unless they are compared at the same time, the nature of the cells and the number of cells cannot be determined 100%, which is because it is possible to cultivate Cells will multiply over time, and the nature will change with the surrounding conditions, so if these points are not taken into account, the cell dose response measured in different situations will get incorrect measurement results. Therefore, in the past, in order to solve such a disadvantage, the number of experimental devices must be increased, so several groups of the same cells will
10330pif. ptd 第4頁 200300450 五、發明說明（2) 被保存在同樣組數的保存系統内，然後再用同樣組數的裝 置同時進行實驗。 發明内容 有鑑於此，本發明的目的之一在於提供一種細胞培養 裝置以及劑量反應的量測方法，可以不管實驗時間點，在 不同的電流值下準確的量測到細胞的劑量反應，在培養J3H 内的分裂細胞的數量與本質均可以被估量出來。 本發明之申請專利範圍第1項提到的細胞培養裝置包括 一個培養皿，用來透過由放置在相對端.的一對電極之間流 動的電流來培養放置在培養基内的細胞；以及電流密度控 制物件，用以對放置在培養孤内培養基中的細胞提供至少 兩個不同電流密度的電流，其中電流密度控制物件會依序 與電流電路系統相連接。 根據本發明申請專利範圍第2項之細胞培養裝置包括一 個培養孤，用來對放置在相對端的一對電極施以特定的電 壓以在電極之間產生電流，來培養放置在培養基内的細 胞；以及電流密度控制物件，用以對放置在培養皿内培養 基中的細胞提供至少兩個不同的電流密度，其中在電流密 度控制物件與電極之間會有一個電力傳送電池構件。 本發明之申請專利範圍第3項係依附於專利申請範圍第 1項或第2項，其中電流密度控制物件係由一個具有超越微 米等級的極小結構之可滲透薄膜構成，此薄膜之空隙不會 讓細胞通過，而在不同區域的複數個薄膜會用以特定間距 設置的支架支撐住。10330pif. Ptd Page 4 200300450 V. Description of the invention (2) It is stored in the storage system with the same number of groups, and then the same number of devices are used to conduct experiments simultaneously. SUMMARY OF THE INVENTION In view of this, one object of the present invention is to provide a cell culture device and a method for measuring dose response, which can accurately measure the dose response of cells under different current values regardless of the time point of the experiment. The number and nature of dividing cells in J3H can be estimated. The cell culture device mentioned in item 1 of the scope of patent application of the present invention includes a culture dish for culturing cells placed in a culture medium through a current flowing between a pair of electrodes placed at opposite ends; and a current density; A control object is used to provide at least two currents with different current densities to the cells placed in the culture medium, wherein the current density control object is sequentially connected with the current circuit system. The cell culture device according to item 2 of the scope of patent application of the present invention includes a culture cell for applying a specific voltage to a pair of electrodes placed at opposite ends to generate a current between the electrodes to cultivate cells placed in the culture medium; And a current density control object for providing at least two different current densities to the cells placed in the culture medium in the culture dish, wherein a power transmission battery component is provided between the current density control object and the electrode. The third item of the scope of patent application of the present invention is dependent on the first or second item of the scope of patent application. The current density control object is composed of a permeable film with a very small structure exceeding the micron level. The voids of this film will not Cells are allowed to pass, and multiple films in different areas are supported by scaffolds arranged at specific intervals.
10330pif. ptd 第5頁 200300450 五、發明說明（3) 根據本發明申請專利範圍第4項提到的劑量反應量測方 法包括下列步驟，對在相對端的一對電極施以特定的電 壓，以在電極之間產生電流；以及同時供應至少兩種不同 的電流密度到同一培養基中的細胞上。 為讓本發明之上述目的、特徵、和優點能更明顯易 懂，下文特舉一較佳實施例，並配合所附圖式，作詳細說 明如下： 實施方式 在本發明較佳實施例中提到的細胞培養裝置將會配合 圖示作進一步詳細的說明，第1圖繪示係依照本發明之較 佳實施例的一種細胞培養裝置。參考編號1 0表示細胞培養 裝置，此細胞培養裝置連接一個電流控制器1 4，可以用來 控制來自電源1 2的電流，並依序連接細胞培養裝置上的電 極以透過一個封閉電路提供特定值的電流。 細胞培養裝置1 0是一個内部空間具有特定的立體體積 的長方形結構，可以容納培養皿1 6，而培養皿1 6的設計可 以容納液體，此培養孤上的兩個相對端會有電極（1 8， 1 8 ’）（在相對壁的内側上），在培養皿内會放入個別的培養 基，透過此培養基可以量測細胞的反應，用特定的間距在 兩個電極之間設置複數個（比如為三個）支架（2 2， 24， 26) ° 支架22， 24， 26上分別放置有可滲透薄膜23, 25， 27, 每一個薄膜都有超越微米等級的極小結構，每一個可滲透 薄膜的孔隙直徑約為0 . 3 mm，厚度介於幾微米到1 0微米之10330pif. Ptd Page 5 200300450 V. Description of the invention (3) The dose response measurement method mentioned in item 4 of the patent scope of the present application includes the following steps, applying a specific voltage to a pair of electrodes at opposite ends to A current is generated between the electrodes; and at least two different current densities are simultaneously supplied to the cells in the same medium. In order to make the above-mentioned objects, features, and advantages of the present invention more comprehensible, a preferred embodiment is given below and described in detail with the accompanying drawings as follows: Embodiments are provided in the preferred embodiments of the present invention. The obtained cell culture device will be further described in detail with reference to the figure. FIG. 1 shows a cell culture device according to a preferred embodiment of the present invention. Reference number 10 indicates a cell culture device. This cell culture device is connected to a current controller 14 and can be used to control the current from the power source 12. It also sequentially connects the electrodes on the cell culture device to provide specific values through a closed circuit. Of current. The cell culture device 10 is a rectangular structure with a specific three-dimensional volume in the internal space, which can accommodate the culture dish 16 and the design of the culture dish 16 can accommodate the liquid. The two opposite ends of this culture isolate will have electrodes (1 8, 1 8 ') (on the inside of the opposite wall), an individual medium will be placed in the Petri dish. Through this medium, the response of the cell can be measured, and a plurality of ( For example, three) stent (2 2, 24, 26) ° Permeable films 23, 25, 27 are placed on the stent 22, 24, 26, each of which has a tiny structure beyond the micron level, each of which is permeable The pore diameter of the film is about 0.3 mm and the thickness is between several micrometers and 10 micrometers.
10330pif.ptd 第6頁 200300450 五、發明說明（4) 間。 在培養m與每個支架之間可能會有連接電力傳送電池 (28， 28’），像是洋菜膠（agarose)電池，洋菜膠可以用在 每個電極的表面上，使用洋菜膠可以減少在細胞上的電極 表面附近的化學反應。 每一個支架22， 24， 26上有可滲透薄膜，只有在這個 部分電流才可以通過，換句話說，大致圓形的可滲透薄膜 2 3， 2 5， 2 7只會放在特定的位置上，透過這些薄膜在電極 之間產生的電流可以流過，可滲透薄膜的面積被設定成 d 1， d 2， d 3， 隨著三個支架而改變。舉例來說，當面積比 隨著三個圓形的薄膜被設定成1 : 2 : 3時，相對應的電流密 度就會變成3 : 2 : 1 。 裝有細胞的培養基會分別被放置在以特定距離安置的 支架上，然後透過對兩個電極施以特定電壓以產生電流， 在培養基内的電流密度可以透過流過的電流量除以薄膜面 極計算出來，接著透過施加特定電壓在兩相對端的一對電 極上，同時提供至少兩種具有不同電流密度的電流到同一 培養基的細胞上以量測劑量反應。 在上述實施例中，那對電極是被安置在側邊，所以電 流會以水平方向流動，但是電流也可以設計成是垂直流 動，只要將電極垂直擺放即可，當電流以垂直方向流動的 時候，流到細胞内的電流會更穩定且均勻。 在提供電流達一段給定的時間以後，支架會被升起， 然後細胞會被用來量測反應，因此就可以得到同樣的細胞10330pif.ptd Page 6 200300450 V. Description of Invention (4). There may be a power transmission battery (28, 28 ') connected between the culture m and each support, such as agarose batteries. Agarose glue can be used on the surface of each electrode. Chemical reactions near the surface of the electrodes on the cells can be reduced. There is a permeable membrane on each of the brackets 22, 24, 26. Only in this part of the current can pass through. In other words, the approximately circular permeable membranes 2 3, 2 5, 2 7 will only be placed in specific positions. The current generated between the electrodes through these films can flow, and the area of the permeable film is set to d1, d2, d3, which changes with the three supports. For example, when the area ratio is set to 1: 2: 3 with three circular films, the corresponding current density becomes 3: 2: 1. The medium containing the cells will be placed on a stand placed at a specific distance, and then a specific voltage will be applied to the two electrodes to generate a current. The current density in the medium can be divided by the amount of current flowing through the membrane surface electrode. It is calculated, and then a specific voltage is applied to a pair of electrodes at two opposite ends, and at least two kinds of currents with different current densities are simultaneously provided to the cells of the same medium to measure the dose response. In the above embodiment, the pair of electrodes are placed on the side, so the current will flow in the horizontal direction, but the current can also be designed to flow vertically, as long as the electrodes are placed vertically, when the current flows in the vertical direction At this time, the current flowing into the cell will be more stable and uniform. After supplying a current for a given period of time, the scaffold will be raised, and the cells will be used to measure the response, so the same cells can be obtained.
10330pif.ptd 第7頁 200300450 五、發明說明（5) 在同樣的時間對不同電流密度的反應；其中薄膜的形狀並 沒有限定是圓形，也可以是方形；細胞是被放置在可滲透 薄膜上，此細胞包括有吸收力的細胞，像是内皮 (absorbent)細胞或成纖維細胞（fiberoblast)。 當採用本發明的細胞培養裝置與劑量反應測試方法 時，即使在因為分裂的時間控制上對細胞的時間控制不同 而使得細胞的數量與本質有變化的情況下，還是可以準確 的量測到細胞對於不同值的電流之劑量反應。10330pif.ptd Page 7 200300450 V. Description of the invention (5) Response to different current densities at the same time; the shape of the film is not limited to circular or square; cells are placed on the permeable film This cell includes absorptive cells, such as endothelial (absorbent) cells or fibroblasts. When the cell culture device and the dose-response test method of the present invention are used, even when the number and nature of cells are changed due to different time control of the cells in the time control of division, the cells can still be accurately measured Dose response to different values of current.
10330pif.ptd 第8頁 200300450 圖式簡單說明 第1圖繪示係依照本發明之較佳實施例的一種細胞培養 裝置。 圖示標記說明： 10 細胞培養裝置1 2 電源 14 電流控制器1 6 培養孤 18， 18’ 電極22， 24， 26 支架 23， 25， 27 薄膜28， 28’ 電力傳送電池 d 1， d 2， d 3 薄膜的面積比10330pif.ptd Page 8 200300450 Brief Description of Drawings Figure 1 shows a cell culture device according to a preferred embodiment of the present invention. Description of pictographs: 10 cell culture device 1 2 power supply 14 current controller 1 6 culture orphan 18, 18 'electrodes 22, 24, 26 holders 23, 25, 27 films 28, 28' power transmission batteries d 1, d 2, d 3 area ratio of film
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