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Planar Patch Clamp Device, Electrodes For Said Device And Cell Ion Channel Current Measurement Method NEW

外国特許コード F170009216
整理番号 AF19-03WO
掲載日 2017年9月13日
出願国 中華人民共和国
出願番号 201280063562
公報番号 104024839
公報番号 104024839
出願日 平成24年12月5日(2012.12.5)
公報発行日 平成26年9月3日(2014.9.3)
公報発行日 平成28年11月16日(2016.11.16)
国際出願番号 JP2012081556
国際公開番号 WO2013094418
国際出願日 平成24年12月5日(2012.12.5)
国際公開日 平成25年6月27日(2013.6.27)
優先権データ
  • 特願2011-278445 (2011.12.20) JP
  • 特願2012-181786 (2012.8.20) JP
  • 2012WO-JP81556 (2012.12.5) WO
発明の名称 (英語) Planar Patch Clamp Device, Electrodes For Said Device And Cell Ion Channel Current Measurement Method NEW
発明の概要(英語) (CN104024839B)
In order to reduce a noise current by suppressing variations in an applied membrane potential in a planar patch clamp device, enabling an ion channel current to be accurately measured, this planar patch clamp device is provided with a liquid reservoir that has at least one fine through-hole disposed on an electrically insulative substrate, and holds a conductive liquid on surfaces on both sides of the through-hole; and electrodes, which are capable of being energized, are disposed in said liquid reservoir.
The electrodes are provided with: (a) an electrode vessel, wherein part of the vessel wall comprises an inorganic porous material; (b) an electrode, wherein a noble metal chloride (NmCl) layer is formed on the surface of the noble metal (Nm); and (c) an electrode solution, wherein the noble metal chloride and an alkali metal chloride are dissolved at a saturated concentration.
特許請求の範囲(英語) [claim1]
1. A planar patch clamp device, are formed with a nerve cell culture of nerve cells of the mesh-shaped planar patch clamp device is used, characterized in,
An (1) electrically insulating substrate 1 is set more fine through-holes, the electrical insulating substrate on both sides of the through hole surface and is in communication does not cause the cells to cause liquid to pass through but even though through the through hole of the 1st, (2) 2nd surface side are respectively provided with a front surface side: used to hold the conductive liquid in Reservoir section; and configured so as to affect this Reservoir section of the conductive liquid is energized to an electrode portion, (3) 1st Reservoir section configured to be as a cell-surface side of the Reservoir section 1st, (4) 2nd surface electrode portion of the side surface side and has the following (a) -(c) component:
(A) the electrode reservoir, when the conductive liquid has been introduced into the Reservoir section in contact with the conductive liquid at least a portion of the container wall by the electrode inside of the vessel to the exterior of the container of the liquid electrode solution mixing small conductive electrode to ignore an inorganic porous material;
(B) electrode, which is housed in a surface layer portion of the noble metal electrode holder and Nm are formed of the noble metal chlorides NmCl layer; and
(C) an electrode solution, which is filled in a container and dissolving the noble metal electrode at a saturating concentration of the alkali metal chloride or chlorides NmCl,
An (5) electrically insulating substrate is attached to the opening rim of the through hole 1st surface side of the extracellular matrix-forming material, the cells are arranged in the state where the through hole, having less than 10M Ω resistance of the seal.
[claim2]
2. The planar patch clamp device according to claim 1, characterized in,
1st surface by the 1st fluid sump Side reservoir portion, and the 2nd fluid sump and passing these The main reservoir Deputy reservoir communicating with the narrow passage from each of the flow of fluid, which is not optically transparent material by the 1st fluid sump and for the configuration cells, 1st 2nd fluid sump provided with electrode portion of the side surface.
[claim3]
3. The planar patch clamp device according to claim 1 or 2, characterized in,
2nd surface side of the Reservoir section and for moving the conductive liquid introduction and expulsion of flow passage in communication with, and is disposed on a portion of the 2nd surface inside the vent side of the electrode unit.
[claim4]
4. The planar patch clamp device according to claim 1 or 2, characterized in,
1st surface side at a plurality of Reservoir section, are formed in these Reservoir section having a width and is larger than that of the nerve cells can be prevented from cultured cells of the cell body of the depth of the concave portion of the moving state.
[claim5]
5. The planar patch clamp device according to claim 1 or 2, characterized in,
The apparatus comprises the following planar patch-clamp (A) /or well (B) a structure of:
(A) 2nd Reservoir section devices are connected to the suction surface side of the liquid, the liquid suction device is able to 2nd through Reservoir section of the negative pressure surface side;
(B) causing cells having an extracellular matrix-forming material adheres to the fixing force of the 1st through-hole in the substrate-side opening peripheries.
[claim6]
6. One method to measure current cell ion channel, characterized in,
Claim 1-5 to any one of the 2nd surface side of the Reservoir section is introduced into the planar patch clamp device electrically conductive liquid, and to be measured is introduced to the 1st surface side of the Reservoir section of the cell dispersion of the conductive liquid, 1st 2nd surface of the side surface of the Reservoir section in an electrically conductive manner to the side of the Reservoir section communicates with, the cells are arranged in a predetermined position of 1st surface side of the Reservoir section, 1st to 2nd surface side and on the basis of the voltage applied between electrodes on the surface side electrode portion, thereby serving as a measurement object of the cells is measured ion channel currents.
[claim7]
7. Ion channel current measurement method cell according to claim 6, characterized in,
1st surface side of the substrate in a planar patch clamp device providing a plurality of Reservoir section, are formed in these Reservoir section having a ratio of nerve cells can be cultured cells of the cell body width and body of the movement of the blocking state of the depth of the recess,
As the measurement objects are cells derived from neural cells,
A cell culture medium to neural cells of the conductive liquid.
[claim8]
8. Ion channel current measurement method cell according to claim 7, characterized in,
Ca introduced beforehand into a nerve cell Ca of the imaging probe for use,
As a means of measuring the Ca includes at least through imaging cell ion channel current, which is to observe during or at risk of cell Ca imaging that is caused by action potentials propagating action potential fluorescence.
[claim9]
9. Cell ion channel current measurement method according to claim 8, characterized in,
1st surface of the patch-clamp device in a plane arranged to correspond to a plurality of even more by introducing Ca probe of nerve cells, and form these are attached to each of the postsynaptic neurons in a neural cell network, to which a single nerve cells or nerve cells of the current injection is made a voltage is applied, thereby performing the plurality of even more based on the measurement of Ca in the nerve cell imaging.
  • 出願人(英語)
  • TAN FEI
  • JAPAN SCIENCE AND TECHNOLOGY AGENCY
  • 発明者(英語)
  • TAN FEI
  • LI HE
  • URISU TSUNEO
  • WANG ZHI-HONG
  • UNO HIDETAKA
  • OBULIRAJ SENTHIL KUMAR
  • NAGAOKA YASUTAKA
国際特許分類(IPC)
参考情報 (研究プロジェクト等) CREST Creation of Nanosystems with Novel Functions through Process Integration AREA
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