JPH01174371A - Cell fusing chamber - Google Patents
Cell fusing chamberInfo
- Publication number
- JPH01174371A JPH01174371A JP62335700A JP33570087A JPH01174371A JP H01174371 A JPH01174371 A JP H01174371A JP 62335700 A JP62335700 A JP 62335700A JP 33570087 A JP33570087 A JP 33570087A JP H01174371 A JPH01174371 A JP H01174371A
- Authority
- JP
- Japan
- Prior art keywords
- cells
- electrodes
- space
- dividing
- cell
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000005192 partition Methods 0.000 claims description 37
- 239000006285 cell suspension Substances 0.000 claims description 19
- 230000007910 cell fusion Effects 0.000 claims description 14
- 238000007499 fusion processing Methods 0.000 claims description 5
- 239000012212 insulator Substances 0.000 claims description 2
- 239000011347 resin Substances 0.000 abstract description 7
- 229920005989 resin Polymers 0.000 abstract description 7
- 239000000725 suspension Substances 0.000 abstract 2
- 239000011810 insulating material Substances 0.000 abstract 1
- 230000001376 precipitating effect Effects 0.000 abstract 1
- 230000005684 electric field Effects 0.000 description 6
- 239000002244 precipitate Substances 0.000 description 2
- 230000006378 damage Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000004904 shortening Methods 0.000 description 1
- 230000000638 stimulation Effects 0.000 description 1
- 230000001052 transient effect Effects 0.000 description 1
Classifications
-
- 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
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Organic Chemistry (AREA)
- Biotechnology (AREA)
- Chemical & Material Sciences (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Microbiology (AREA)
- Sustainable Development (AREA)
- Physics & Mathematics (AREA)
- Biomedical Technology (AREA)
- Cell Biology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Electromagnetism (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
Abstract
Description
【発明の詳細な説明】
（産業上の利用分野）
本発明は電気刺激を利用して細胞とおしを融合させるた
めに、細胞懸濁液を収容する空間に対向電極を備え、そ
の電極間に電圧を印加して細胞に電界を印加する細胞融
合チャンバに関するものである。Detailed Description of the Invention (Field of Industrial Application) In order to fuse cells and cells using electrical stimulation, the present invention provides a space containing a cell suspension with opposing electrodes, and applies a voltage between the electrodes. The present invention relates to a cell fusion chamber that applies an electric field to cells by applying an electric field.
（従来の技術）
細胞融合チャンバでは細胞懸濁液を収容する空間に対向
電極を備えている。電極間に細胞懸濁液を入れ、細胞が
電極間の底部に沈澱した後に電界を印加する。細胞融合
をさせる場合は、初めに交流電界を印加して細胞に誘電
電気泳動を起こさせて細胞どおしを接触させ、その後、
高電圧直流パルス電界を印加することによって融合した
細胞に一過的膜破壊を生じさせて細胞どおしを融合させ
る。(Prior Art) A cell fusion chamber is equipped with a counter electrode in a space that accommodates a cell suspension. A cell suspension is placed between the electrodes, and an electric field is applied after the cells settle to the bottom between the electrodes. For cell fusion, first apply an alternating current electric field to cause dielectric electrophoresis in the cells to bring them into contact, and then
By applying a high-voltage direct current pulsed electric field, transient membrane destruction is caused in the fused cells, and the cells are fused together.
（発明が解決しようとする問題点）
従来の細胞融合チャンバでは電極間の底面は１つである
。電極間に入れられた細胞懸濁液の細胞が全てその底面
上に沈澱するには時間がかかる。(Problems to be Solved by the Invention) In the conventional cell fusion chamber, there is one bottom surface between the electrodes. It takes time for all the cells in the cell suspension placed between the electrodes to settle onto the bottom surface.
また、全細胞が狭い底面上に沈澱すると細胞濃度が高く
なりすぎて交流電界により細胞どおしを接触させたとき
多数の細胞が接触してマルチペアとなり、健全な融合細
胞が得られにくくなる。細胞融合は一対の細胞が融合し
てシングルペアとなる方が好ましい。Furthermore, if all the cells are precipitated on a narrow bottom surface, the cell concentration will become too high, and when cells are brought into contact with each other using an alternating electric field, many cells will come into contact and form multi-pairs, making it difficult to obtain healthy fused cells. In cell fusion, it is preferable that a pair of cells fuse to form a single pair.
本発明は、細胞懸濁液が収容される空間の電極間の底面
の面積を大きくすることによって、細胞が沈澱する時間
を短縮すると件もに、健全な融合細胞を多く得ることの
できる細胞融合チャンバを提供することを目的とするも
のである。The present invention improves cell fusion by increasing the area of the bottom surface between the electrodes in the space in which the cell suspension is accommodated, thereby shortening the time for cells to settle. The purpose is to provide a chamber.
（問題点を解決するための手段）
本発明では、対向電極を備えて細胞懸濁液を収容できる
空間に、細胞融合処理時に互いに平行な複数の底面を形
成する絶縁物製の仕切り部材を備えた。(Means for Solving the Problems) In the present invention, a space provided with opposing electrodes and capable of accommodating a cell suspension is provided with a partition member made of an insulator that forms a plurality of bottom surfaces parallel to each other during cell fusion processing. Ta.
（作用）
対向電極間に仕切り部材が存在し、細胞懸濁液が収容さ
れた状態で仕切り部材の底面が水平となるように静置す
る。仕切り部材の底面の面積の合計は対向電極間のチャ
ンバの底面の面積よりも大きい。細胞懸濁液中の細胞は
仕切り部材の底面上に沈澱する。(Function) A partition member exists between the opposing electrodes, and the cell suspension is left standing so that the bottom surface of the partition member is horizontal. The total area of the bottom surfaces of the partition members is larger than the area of the bottom surface of the chamber between the opposing electrodes. The cells in the cell suspension settle on the bottom surface of the partition member.
細胞沈澱後、所定の電気融合の処理を施す。After cell precipitation, a predetermined electrofusion treatment is performed.
（実施例）
第１図から第３図に一実施例を表わす。第１図、第２図
は垂直断面図であり、第３図は水平断面図である。第１
図は第３図のＡ−Ａ線位置で切断した状態を表わし、第
２図は第３図のＢ−Ｂ線位置で切断した状態を表わし、
第３図は第１図のＣ−Ｃ線位置で切断した状態を表わし
ている。(Example) An example is shown in FIGS. 1 to 3. 1 and 2 are vertical sectional views, and FIG. 3 is a horizontal sectional view. 1st
The figure shows the state cut at the line A-A in Fig. 3, and the figure 2 shows the state cut at the line B-B in Fig. 3.
FIG. 3 shows a state cut along the line C--C in FIG. 1.
１は円筒形の樹脂製ボディであり、その底部には透明ガ
ラス製の底板２が接着されている。底板２上には一対の
対向電極３−１．３−２が接着されている。Reference numeral 1 denotes a cylindrical resin body, to the bottom of which a transparent glass bottom plate 2 is adhered. A pair of opposing electrodes 3-1, 3-2 are bonded onto the bottom plate 2.
電極３−１．３−２はその対向面の底部側の面３ａがと
もに底板２の面に対して垂直になるように加工され、か
つ、互いに平行になるように位置決めされている。電極
３−１．．３−２の対向面の上部の面３ｂはともに外方
向に向って開くように傾斜している。The electrodes 3-1, 3-2 are processed so that the bottom side surfaces 3a of the opposing surfaces are both perpendicular to the surface of the bottom plate 2, and positioned so that they are parallel to each other. Electrode 3-1. ．． The upper surfaces 3b of the opposing surfaces 3-2 are both inclined so as to open outward.
電極３−１，３−２とボディ１の間は絶縁製樹脂４で充
填され、この樹脂４によって電極３−１゜３−２がボデ
ィｌに接着され、さらに底板２もボディ１に接着されて
いる。The space between the electrodes 3-1, 3-2 and the body 1 is filled with an insulating resin 4, and the electrodes 3-1 and 3-2 are bonded to the body 1 by this resin 4, and the bottom plate 2 is also bonded to the body 1. ing.
電極３−１．３−２にはそれぞれボディ１の外部に設け
られた端子５−１．５−２がボディ１を貫通して接続さ
れている。Terminals 5-1, 5-2 provided outside the body 1 penetrate through the body 1 and are connected to the electrodes 3-1, 3-2, respectively.
ボディｌの外壁の上部にはネジが設けられている。６は
ボディ１の上部開口を閉じる蓋であり、蓋６にもボディ
１のネジと噛み合うネジが設けられている。A screw is provided at the upper part of the outer wall of the body l. Reference numeral 6 denotes a lid that closes the upper opening of the body 1, and the lid 6 is also provided with a screw that engages with the screw of the body 1.
７は絶縁製樹脂にてなる仕切り部材であり、仕切り部材
７は第４図に示されるように下部に電極３−１．３−２
間の空間に挿入される仕切り板８が複数枚設けられてい
る。仕切り板８が設けられている仕切り部材の本体側は
、第４図で仕切り板８から本体側に向って広がるような
傾斜面８ａ’＋８８をもっている。Reference numeral 7 denotes a partition member made of insulating resin, and the partition member 7 has electrodes 3-1, 3-2 at the bottom as shown in FIG.
A plurality of partition plates 8 are provided to be inserted into the space between them. The main body side of the partition member on which the partition plate 8 is provided has an inclined surface 8a'+88 that widens from the partition plate 8 toward the main body side as shown in FIG.
仕切り部材７は、第２図に示されるように電極３−１．
３−２間に挿入される。仕切り板８の先端は底板２の面
に接触してその面と直交する。仕切り部材７の基端面７
ｂは蓋６をボディ１にネジ止めすることによって蓋６の
裏面で押しつけられ、仕切り板８の先端が底板４に押し
つけられるとともに、傾斜面７ａ、７ａが電極３−１．
３−２の傾斜面３ｂ、３ｂにそれぞれ押しつけられて電
極間３−１．３−２で細胞懸濁液を収容する空間を密閉
し、かつ、各仕切り板８でその空間を仕切る。As shown in FIG. 2, the partition member 7 includes the electrodes 3-1.
It is inserted between 3 and 2. The tip of the partition plate 8 contacts the surface of the bottom plate 2 and is perpendicular to that surface. Base end surface 7 of partition member 7
b is pressed against the back surface of the lid 6 by screwing the lid 6 to the body 1, the tip of the partition plate 8 is pressed against the bottom plate 4, and the inclined surfaces 7a, 7a are pressed against the electrodes 3-1.
The space accommodating the cell suspension is sealed between the electrodes 3-1 and 3-2 by being pressed against the inclined surfaces 3b and 3b of 3-2, respectively, and the space is partitioned by each partition plate 8.
次に、本実施例を用いた細胞融合操作を説明する。Next, a cell fusion operation using this example will be explained.
蓋６がボディ１から取り外され、仕切り部材７も取り外
された状態で、電極間３−１．３−２の空間に細胞懸濁
液９を入れる。With the lid 6 removed from the body 1 and the partition member 7 also removed, the cell suspension 9 is placed in the space between the electrodes 3-1, 3-2.
次に、第２図に示されるように仕切り部材７を電極３−
１．３−２間に入れ、蓋６をボディ１にネジ止めする。Next, as shown in FIG.
1. Place the cover between 3 and 2 and screw the lid 6 onto the body 1.
これにより、細胞懸濁液９はボディ１、電極３−１．３
−２、底板２及び仕切り部材７によって密閉され、かつ
、仕切り板８により複数の空間に区切られた状態となる
。As a result, the cell suspension 9 is transferred to the body 1 and the electrode 3-1.3.
-2, the space is sealed by the bottom plate 2 and the partition member 7, and divided into a plurality of spaces by the partition plate 8.
その後、仕切り板８が水平方向になるように、例えば第
１図で端子の設けられている側が上になるようにチャン
バを立てる。Thereafter, the chamber is erected so that the partition plate 8 is oriented horizontally, for example, so that the side on which the terminals are provided in FIG. 1 is facing upward.
その状態でチャンバを静置すると、各仕切り板８上に細
胞が沈澱してくる。細胞が沈澱して安定した頃を見計ら
って端子５−１〜５−２から、初めに交流電圧を印加し
て細胞どおしを接触させ、その後所定の高電圧直流パル
スを印加して細胞融合を起こさせる。When the chamber is left still in this state, cells will precipitate on each partition plate 8. When the cells have settled and become stable, first apply AC voltage from terminals 5-1 to 5-2 to bring the cells into contact, and then apply a predetermined high-voltage DC pulse to fuse the cells. cause to happen.
細胞融合処理終了後は、チャンバを図に示されるように
横方向に戻し、蓋６を取り外し、仕切り部材７も取り外
して細胞懸濁液９を回収する。After the cell fusion process is completed, the chamber is returned to the horizontal direction as shown in the figure, the lid 6 is removed, the partition member 7 is also removed, and the cell suspension 9 is collected.
第５図は本発明をセル型チャンバに適用した実施例を表
わす。FIG. 5 shows an embodiment in which the present invention is applied to a cell type chamber.
１０はチャンバのボディ、１１−１．１１−２は対向電
極、１４は電＠１ｌ−１，１１−２をボディ１ｏに接着
する絶縁製樹脂である。10 is the body of the chamber, 11-1, 11-2 is a counter electrode, and 14 is an insulating resin for bonding the electrodes 11-1 and 11-2 to the body 1o.
対向電極１１−１．１１−２間には互いに平行な仕切り
板を複数個有する仕切り部材１２が入れられている。電
極１１−１．１１−２の対向面は垂直方向を向き、仕切
り部材１２の仕切り板は水平方向を向いている。A partition member 12 having a plurality of mutually parallel partition plates is inserted between the opposing electrodes 11-1 and 11-2. The opposing surfaces of the electrodes 11-1 and 11-2 face in the vertical direction, and the partition plates of the partition member 12 face in the horizontal direction.
電極間１１−１．１１−２には細胞懸濁液９が入れられ
、細胞が仕切り部材１２の各仕切り板上に沈澱した後に
細胞融合のための所定の電気処理が施される。A cell suspension 9 is placed between the electrodes 11-1 and 11-2, and after the cells are precipitated on each partition plate of the partition member 12, a predetermined electrical treatment for cell fusion is performed.
仕切り部材１２は、例えば第６図に示されるように互い
に平行になるように一体化された、例えば樹脂製のもの
であり、細胞懸濁液はその仕切り部材１２と電極１１−
１．１１−２の隙間又は仕切り部材１２とボディ１０の
隙間を通って上部から注ぎ込むことができる。The partition members 12 are made of resin, for example, and are integrated so as to be parallel to each other, as shown in FIG.
1. It can be poured from above through the gap 11-2 or the gap between the partition member 12 and the body 10.
細胞懸濁液９を注入するために、又例えば第６図に示さ
れるように、各仕切り板に孔１３をあ番プ、その孔１３
を経て細胞懸濁液９を注入するようにしてもよい。In order to inject the cell suspension 9, a hole 13 is provided in each partition plate, as shown in FIG.
Alternatively, the cell suspension 9 may be injected through the steps.
（発明の効果）
本発明では、対向電極を備えて細胞懸濁液を収容できる
空間に、細胞融合処理時に互いに平行な複数の底面を形
成する絶縁物製の仕切り部材を備えたので、対向電極間
で細胞が沈澱することのできる底面の面積が増え、細胞
の沈澱に要する時間が短かくなって所定の融合処理時間
を短縮することができる。(Effects of the Invention) In the present invention, a space provided with a counter electrode and capable of accommodating a cell suspension is provided with an insulating partition member that forms a plurality of mutually parallel bottom surfaces during cell fusion processing. The area of the bottom surface on which cells can be precipitated increases, and the time required for cell precipitate becomes shorter, making it possible to shorten the predetermined fusion processing time.
また、細胞は底面上に適当な密度で安定して沈澱し、細
胞の融合数を高めることができる。In addition, cells are stably precipitated on the bottom surface at an appropriate density, and the number of cell fusions can be increased.
また、塩濃度が高いときの細胞懸濁液の対流を抑え、融
合を容易に行なうことができる。Furthermore, when the salt concentration is high, convection of the cell suspension can be suppressed and fusion can be easily performed.
第１図は一実施例を表わす垂直断面図、第２図も同実施
例を表わす垂直断面図、第３図は同実施例の水平断面図
であり、第１図は第３図のＡ−Ａ線位置で切断した状態
を表わし、第２図は第３図のＢ−Ｂ線位置で切断した状
態を表わし、第３図は第１図のＣ−Ｃ線位置で切断した
状態を表わしている。第４図は同実施例における仕切り
部材を示す斜視図、第５図は本発明をセル型チャンバに
適用した実施例を表わす垂直断面図、第６図は同実施例
における仕切り部材を示す斜視図である。
１．１０・・・・・・ボディ、
２・・・・・・底板、
３−１．３−２．１１−１．１１−２・・・・・・対向
電極、
７．１２・・・・・・仕切り部材、
８・・・・・・仕切り板、
９・・・・・・細胞懸濁液。
特許出願人 株式会社島津製作所FIG. 1 is a vertical sectional view showing one embodiment, FIG. 2 is a vertical sectional view showing the same embodiment, and FIG. 3 is a horizontal sectional view of the same embodiment. Figure 2 shows the state cut at line A, Figure 2 shows the state cut at line B-B in Figure 3, and Figure 3 shows the state cut at line C-C in Figure 1. There is. FIG. 4 is a perspective view showing a partition member in the same embodiment, FIG. 5 is a vertical sectional view showing an embodiment in which the present invention is applied to a cell type chamber, and FIG. 6 is a perspective view showing a partition member in the same embodiment. It is. 1.10...Body, 2...Bottom plate, 3-1.3-2.11-1.11-2...Counter electrode, 7.12... ... partition member, 8 ... partition plate, 9 ... cell suspension. Patent applicant: Shimadzu Corporation
Claims (1)
、前記空間にあって細胞融合処理時に互いに平行な複数
の底面を形成する絶縁物製の仕切り部材を備えた細胞融
合チャンバ。A cell fusion chamber comprising a counter electrode and a space capable of accommodating a cell suspension, and a partition member made of an insulator that is located in the space and forms a plurality of mutually parallel bottom surfaces during cell fusion processing.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP62335700A JPH01174371A (en) | 1987-12-29 | 1987-12-29 | Cell fusing chamber |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP62335700A JPH01174371A (en) | 1987-12-29 | 1987-12-29 | Cell fusing chamber |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH01174371A true JPH01174371A (en) | 1989-07-10 |
Family
ID=18291504
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP62335700A Pending JPH01174371A (en) | 1987-12-29 | 1987-12-29 | Cell fusing chamber |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH01174371A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1993002178A1 (en) * | 1991-07-22 | 1993-02-04 | Schmukler Robert E | Apparatus and methods for electroporation and electrofusion |
| JP2005520542A (en) * | 2002-03-20 | 2005-07-14 | バイオ−ラッド ラボラトリーズ，インコーポレイティド | Electroporation chamber |
-
1987
- 1987-12-29 JP JP62335700A patent/JPH01174371A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1993002178A1 (en) * | 1991-07-22 | 1993-02-04 | Schmukler Robert E | Apparatus and methods for electroporation and electrofusion |
| JP2005520542A (en) * | 2002-03-20 | 2005-07-14 | バイオ−ラッド ラボラトリーズ，インコーポレイティド | Electroporation chamber |
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