JP2008261833A - Flow device and flow method - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To prevent a phenomenon wherein, when driving a sample solution by an action of an electroosmotic flow or electrophoresis, a difference is generated in a flow rate distribution in a channel, and thereby flow of the sample solution becomes difficult. <P>SOLUTION: A pair of solution tanks and the channel 23 are formed in the recessed state on a channel substrate 1. A pressure sensor 7 and a micro-pipette are arranged in the solution tank on the downstream side, and crushing electrodes 56, 56 for applying a voltage for cell crushing and a narrow width part 55 are formed in the middle of the channel 23. When being used, the sample solution is introduced into the solution tank on the upstream side, and a driving voltage is applied to driving electrodes 51, 51, to thereby drive the sample solution. All this while, an inflow pressure of the sample solution into the solution tank on the downstream side is always measured by the pressure sensor 7, and when the inflow pressure falls below a threshold, a liquid flow adjusting device 6 is driven, and the sample solution in the solution tank is sucked from the micro-pipette. Hereby, the flow state of the sample solution can be maintained. <P>COPYRIGHT: (C)2009,JPO&INPIT

Description

  The present invention relates to a flow device in which a micro flow channel is formed on a flow channel substrate, and a flow method when the flow device is used.

In recent years, in biochemical analysis of cells, DNA (deoxyribonucleic acid), RNA (ribonucleic acid), and the like, a chip formed with a fine channel having a width of several hundred μm or less may be used (see Patent Document 1). ). In this chip, electrodes are formed at both ends in the extending direction of the flow path, and by applying a driving voltage to both electrodes, DNA or cells in the sample solution introduced into the flow path can be electroosmotic flow or electric It can be moved by electrophoresis, and operations such as mixing and separation of the DNA and cells can be performed.
JP 2004-286449 A

  However, in the chip having the above configuration, when the driving voltage is continuously applied, the pressure of the sample solution on the downstream side becomes higher than that on the upstream side due to the movement of the sample solution itself, and the phenomenon that it gradually becomes difficult to flow may occur. there were.

  This invention is made | formed in view of the above-mentioned subject, and it aims at providing the means which can maintain a fluid state by controlling the pressure balance in a flow path.

  In addition, since the flow path is fine and extends along the surface, it was introduced into the flow path depending on environmental conditions, for example, when illuminated by a microscope for a long time, or when it was hot or dry. In some cases, the liquid evaporates and falls below the required amount.

  The present invention has been made in view of the above-described problems, and an object thereof is to prevent evaporation and drying of liquid in the flow path of the flow path substrate.

  In order to solve the above-mentioned problem, a flow apparatus according to claim 1 of the present invention includes at least one pair of solution tanks and a flow path substrate in which a flow path communicating with the one pair of solution tanks is formed in a concave shape, and the one pair. Driving means for driving the sample solution introduced into the solution tank or the flow path from one side of the solution tank to the other by electroosmotic flow or electrophoresis, and the pair of solutions generated during the driving A liquid amount adjusting means for partially injecting or removing the sample solution so as not to cause a pressure that hinders the driving force of the driving means due to a difference in the amount of the sample solution in the tank and the flow path. It is characterized by.

  Here, in the description of the claims, “inhibiting the driving force” means reducing the driving force or increasing the driving force. As in the present invention, by removing the difference between the upstream and downstream liquid volume that causes the electric driving force to be obstructed and controlling the pressure balance between the two solution tanks, the sample solution is generated by the generation of the pressure gradient. Can be prevented from becoming difficult to flow. In the present invention, the sample is driven by purely electroosmotic flow or electrophoresis action by adjusting the amount of liquid instead of adding a flow rate to the sample solution. The material can be prevented from being destroyed by an excessive flow rate.

  According to a second aspect of the present invention, there is provided a fluidizing device according to the first aspect, further comprising determination means for determining whether or not a pressure that inhibits the driving force is generated. This allows more precise control.

  In addition, in the structure of Claim 1, it is not necessary to provide a determination means, and it is good also as a structure which makes a liquid amount adjustment means perform a predetermined procedure.

  The flow device according to claim 3 of the present invention is characterized in that, in claim 2, the determination means performs the determination based on the pressure of the sample solution detected in the pair of solution tanks or flow paths. And

  The flow device according to a fourth aspect of the present invention is the flow device according to any one of the first to third aspects, wherein the liquid amount adjusting means sucks a sample solution in the solution tank on the downstream side or the solution on the upstream side. The sample solution is injected into the tank.

  In this way, it is possible to prevent the pressure and flow rate during injection and suction from affecting the electroosmotic flow or the driving force of electrophoresis by performing the solution through the solution tank instead of directly injecting and suctioning into the flow path. Moreover, destruction of samples such as cells can be prevented.

  The flow device according to claim 5 of the present invention includes the cell crushing unit that crushes cells in the sample solution driven by the driving means in the flow channel according to any one of claims 1 to 4. It is characterized by.

  A flow device according to a tenth aspect of the present invention is characterized in that in any one of the first to fifth aspects, a cooling means for cooling the flow path substrate is provided. According to this, since the flow path substrate is cooled, evaporation of the liquid in the flow path is suppressed, and a long-time operation or the like is possible.

  The cooling means is preferably a Peltier element. Moreover, it is preferable that the Peltier element is disposed so that the endothermic surface thereof is in contact with the bottom surface of the flow path substrate. Moreover, it is preferable to provide the 2nd cooling means which cools the thermal radiation surface of the said Peltier element.

  A flow method according to claim 10 of the present invention includes at least one pair of solution tanks and a flow path substrate in which a flow path communicating with the one pair of solution tanks is formed in a concave shape, and the pair of solution tanks or the flow paths. And a driving means for driving the sample solution introduced into the solution tank from one side of the solution tank to the other by electroosmotic flow or electrophoresis, and a method of flowing the sample solution, The sample solution is partially injected or removed so that a pressure that hinders the driving force of the driving means does not occur due to the difference in the amount of the sample solution in the pair of solution tanks and flow paths generated during the driving. It is characterized by that.

  In the flow method, it is preferable that the flow path substrate is cooled by a Peltier element or the like to suppress evaporation of the sample solution.

  According to the present invention, the sample solution can be driven so as not to stay in the flow path.

  Next, embodiments of the present invention will be described with reference to the drawings.

  <First Embodiment>

  FIG. 1 is a functional block diagram of the cell disruption apparatus according to the first embodiment, FIG. 2A is a plan view of the flow path substrate 1 according to the present embodiment, and FIG. 1B is a flow of FIG. FIG. 3 is a cross-sectional view taken along the line II of FIG. 2A, FIG. 4 is a schematic view showing a liquid amount adjusting device according to the present embodiment, and FIG. 5 is a liquid amount adjusting device. FIG. 6 is a view showing a micropipette of the liquid amount adjusting device, and FIG. 7 is a view showing the arrangement positions of the micropipette and the pressure sensor 7.

  In the first embodiment, the flow device as an example of the present invention is configured as a cell crushing device for crushing a small amount of cells. As shown in FIG. 1, the cell disruption apparatus includes a flow path substrate 1, a drive voltage application device 52, a disruption voltage application device 53, a pressure sensor 7, a controller 8, and a liquid amount adjustment device 6. .

  As shown in FIG. 2B, the flow path substrate 1 is formed by sequentially laminating a film electrode 14 and a photosensitive resin layer 12 on a glass support 11.

  The support 11 is a plate-like support that supports the membrane electrode and the photosensitive resin layer 12. In the present embodiment, a commercially available plate-like borosilicate glass (trade name: Pyrex (registered trademark)) having a length of 20 mm, a width of 10 mm, and a thickness of 1 mm is used as the support 11.

  The membrane electrode 14 is formed in a predetermined pattern on the support 11, thereby constituting a pair of driving electrodes 51, 51 and a pair of crushing electrodes 56, 56 described later. The respective electrodes 51, 51, 56, 56 are formed so as to be separated from each other and extend to the end portion of the support 11 so as to be connectable to the external electrode at the end portion.

  In FIG. 2A, for the sake of easy understanding, the formation region of the film-like electrode is indicated by hatching. Among these, the hatched portion of the broken line indicates a portion formed under the photosensitive resin layer 12, and the hatched portion of the solid line indicates a portion where the film-like electrode is exposed. In this embodiment, after a mask pattern is formed on the support 11 using a resist or the like, a film electrode having a two-layer structure is formed by sputtering and covering the surface with a Pt reaction protective film (see FIG. 2 (b)). Thus, the adhesion with the support 11 can be improved by using Ti as a base, and the electrode reaction when exposed to the sample solution can be suppressed by covering with Pt.

  The photosensitive resin layer 12 is formed by photoreacting a photosensitive resin, and a predetermined two-dimensional pattern is formed on the surface of the support 11 with a predetermined thickness. Due to the difference in thickness between the formed portion of the photosensitive resin layer 12 and the unformed portion, a recess (described later) 2 for storing the sample solution is formed on the surface of the flow path substrate 1. In this embodiment, a negative photoresist (trade name: SU-8) manufactured by Kayaku Microchem Co., Ltd. is applied to the support 11 with a spin coater, exposed to ultraviolet light through a pattern mask, and undeveloped with a developer. The photosensitive resin layer 12 having a thickness of 25 μm is formed by dissolving and removing the cured portion.

  Next, the configuration of the recess 2 will be described. In FIG. 2 (a), the concave portion 2 is composed of solution tanks 21 and 21 having a substantially triangular shape in a plan view and symmetrically arranged on the paper surface, and a flow path 23 connecting these solution tanks 21 and 21. Yes.

  The solution tanks 21 and 21 are used for charging or collecting the sample solution. A pair of drive electrodes 51, 51 are formed in the formation regions of the solution tanks 21, 21, respectively, and are exposed on the surface, and constitute the bottom surfaces of the solution tanks 21, 21. The pair of driving electrodes 51 and 51 receives a power supply from the driving voltage applying device 52 and applies a driving voltage for forming an electric field serving as a driving force of the sample solution. A DC voltage (for example, 30V) is applied as the driving voltage.

  In the flow path 23, a narrow width portion 55 having a shape that narrows the flow path width is formed in the middle of the flow path direction. The narrow portion 55 shown in FIG. 2 is formed by projecting the flow path wall, and is smaller than the diameter of the cell to be crushed at the narrowest portion of the flow path width (FIG. 3). reference). Further, a pair of crushing electrodes 56 and 56 are formed side by side along the flow path direction in the vicinity of the narrow width portion 55 and upstream of the narrowest flow path width portion. In response to power supply from the disruption voltage application device 53, a high-frequency voltage for cell disruption (for example, 0.1 to 1 MHz) is applied.

  In the present embodiment, the width of the flow path 23 is set to 20 μm except for the narrow width portion 55, and the depth is 25 μm.

  The drive voltage application device 52 supplies drive voltage to the drive electrodes 51 and 51 as described above, and constitutes drive means together with the drive electrodes 51 and 51. The crushing voltage application device 53 supplies a high frequency voltage for cell crushing to the crushing electrodes 56 and 56, and constitutes a cell crushing unit together with the narrow width portion 55 and the crushing electrodes 56 and 56.

  In this embodiment, the pressure sensor 7 is provided inside the solution tank 21 on the downstream side where the sample solution flows, and detects the inflow pressure of the sample solution in the vicinity of the connection port with the flow path 23. In the present embodiment, a diaphragm as a pressure-sensitive elastic element is installed inside the solution tank 21, the mechanical signal is converted into an electric signal and output to the controller 8 described later, and the inflow pressure is calculated. FIG. 7 shows the arrangement position of the pressure-sensitive elastic element.

  The controller 8 determines based on the inflow pressure information of the sample solution input from the pressure sensor 7 whether or not the driving force generated by the application of the driving voltage is inhibited, and the driving force is inhibited. When the determination is made, a sample solution suction or injection command is output to the liquid amount adjusting device 6. Specifically, in the present embodiment, it is determined whether or not the inflow pressure is greater than or equal to a threshold value, and it is determined that the driving force is inhibited when it is less than the threshold value. And the liquid amount adjusting device for determining the suction amount or the injection amount based on the predetermined correspondence between the difference from the threshold value of the inflow pressure and the suction amount or the injection amount, and performing the suction or injection of the determined amount 6 is calculated (drive angle in the present embodiment), and a drive signal corresponding to the drive amount is output to the liquid amount adjusting device 6. The threshold value of the inflow pressure and the correspondence between the difference and the suction amount or the injection amount are obtained in advance by experiments or simulations.

  The above controller 8 corresponds to the determination means. In this embodiment, the controller 8 is configured by a computer, and stores the threshold value and the correspondence relationship as data such as a table. The series of programmed determination and calculation processes are performed while the sample solution is being driven. It is executed by timer interrupt processing at predetermined time intervals (for example, every 10 msec).

  The liquid amount adjusting device 6 receives the drive signal from the controller 8 and sucks the sample solution stored in the downstream solution tank 21. In the present embodiment, as shown in FIG. 4, a pump drive device 65, a coupling 64, a pump 63, a tube 62, and a micropipette 61 are provided.

  The pump driving device 65 includes a stepping motor and a driver for driving the stepping motor, and the driving shaft of the stepping motor rotates by a specified angle based on a driving signal input from the controller 8. The rotational driving force is transmitted to the pump 63 shown in FIG.

  In the pump 63 of FIG. 5, a connecting shaft 63 a coupled to the coupling 64 is rotatably supported inside the housing 63 b (not shown), and rotates according to the rotation of the pump driving device 65. The other end of the connecting shaft 63a is coaxially connected to the piston 63c via a screw mechanism inside the housing 63b. The piston 63c is retracted in the axial direction by the screw mechanism according to the rotation angle of the connection shaft 63a, thereby reducing the liquid pressure of the sample solution filled in the hollow portion of the cylinder 63d.

  A micropipette 61 is connected to the tip of the cylinder 63d through a tube 62, and the tip (capillary 61a in the figure is attached) is arranged toward the inside of the solution tank 21 on the downstream side ( (See FIG. 7). Therefore, when the hydraulic pressure in the cylinder 63d decreases as described above, the sample solution is sucked from the solution tank 21. In FIG. 5, reference numeral 63f denotes an injection hole that penetrates through the hollow portion of the cylinder 63d, and is used to fill the hollow portion with a sample solution or vent the air. Reference numeral 63g is a base for fixing the housing 63b. The liquid amount adjusting device 6 described above corresponds to the liquid amount adjusting means.

(About the effect of cell disruption device)
Next, the effect of the cell crushing device having the above configuration will be described.

  When the cell disrupting device is used, the driving electrodes 51 and 51 of the flow path substrate 1 are connected to the electrodes of the driving voltage applying device 52 so as to be operable. Then, a sample solution containing cells to be crushed is put into one solution tank 21 of the flow path substrate 1. In this state, the drive voltage application device 52 is powered on and a drive voltage is applied to the pair of drive electrodes 51 and 51. In this use example, the sample solution is introduced into the solution tank 21 on the left side of the paper surface, and the drive electrode 51 of the solution tank 21 is used as the positive electrode and the other is used as the negative electrode. Generate osmotic flow.

  By this electroosmotic flow, the cells in the sample solution are transported from the positive electrode side to the negative electrode side, but if the drive voltage is continuously applied, the liquid volume gradually increases between the positive electrode side and the negative electrode side due to the movement of the sample solution itself. A difference occurs, and the water level on the negative electrode side is slightly higher than that on the positive electrode side. Here, the water level on the negative electrode side does not increase as compared with the positive electrode side into which the sample solution is introduced in the state where the external force does not work. However, in the state where the driving voltage is applied, the driving force from the positive electrode side to the negative electrode side works against the force (pressure) that the sample solution moved to the negative electrode side tries to return to the positive electrode side. There remains a difference in the water level, which results in a pressure gradient that is higher on the negative electrode side and lower on the positive electrode side.

  And if a water level becomes high and this pressure and driving force balance, the flow velocity of an electroosmotic flow will fall and it will become difficult to flow a cell. This state is detected by the controller 8 as a decrease in the inflow pressure in the solution tank 21, and the sample solution in the solution tank 21 on the downstream side is sucked by the liquid amount adjusting device 6. As a result, the pressure of the sample solution on the downstream side is reduced, the driving force is increased, and the flow velocity of the electroosmotic flow is recovered.

  In this way, the electroosmotic flow is generated without stagnation, and the cells reach the narrow portion 55. When the cells are sandwiched between the narrow portions 55 and stop moving, the cells are energized and crushed by applying a high frequency voltage by the crushing voltage application device 53, and the inclusions (nucleic acid, protein, etc.) are transferred to the sample solution. It elutes and is conveyed to the solution tank 21 on the negative electrode side by electroosmotic flow.

  As described above, when a pressure that inhibits the driving force is generated, the influence of the pressure fluctuation can be removed by sucking the sample solution by the liquid amount adjusting device 6, and only the action of the electroosmotic flow is pure. Fluid control based on

(Modification)
The application of the present invention is not limited to the above embodiment. For example, the drive voltage is not limited to a DC voltage, and the present invention can also be applied to a case where an AC voltage is applied and a sample solution and cells contained therein are moved by dielectrophoresis.

  Further, whether or not the pressure that inhibits the driving force is generated is not limited to the case where the inflow pressure is measured by the pressure sensor 7 and determined based on this, for example, the determination based on the flow rate or the downstream side You may determine based on the water level in the solution tank 21, or a liquid quantity.

  Even when the pressure sensor 7 is used, it is not limited to always detecting the inflow pressure of the downstream solution tank 21 and determining whether or not the pressure is below the threshold. For example, the upstream and downstream solutions You may provide the pressure sensor 7 in both the tanks 21 and 21, measure inflow pressure and outflow pressure, and may determine based on the difference. Furthermore, without feeding back information such as pressure, for example, the suction operation is performed in a predetermined sequence according to information (driving conditions such as flow velocity and driving voltage, type of sample solution, flow amount, etc.) determined before driving. You may do it.

  Further, without using the pressure sensor 7 or the like, for example, a cell flowing in the flow path 23 is photographed with a camera attached to a microscope, and the moving speed of the cell is analyzed by image analysis based on the camera image (moving image information). By detecting and comparing with the theoretical value, it can be determined whether or not a pressure that inhibits the driving force is generated. In this case, for example, if the moving speed is slower than the theoretical value and out of the allowable range, the two micropipettes 61 provided on the upstream side and the downstream side may be driven to remove the inhibitory component due to pressure.

  Further, it is not necessary to fix the micropipette 61 while it is inserted in the solution tank 21 on the downstream side. For example, the micropipette 61 may be placed on an XY table so as to be movable. In this case, it is possible to move to the portion where the pressure that hinders the driving force is excessive and suck, and finer control becomes possible. In addition, the adjustment of the liquid amount is not limited to the case of sucking, and the sample solution may be injected into, for example, the portion of the recess 2 where the water level is low as long as the pressure balance between the upstream side and the downstream side can be controlled. In this case, a sample solution containing no cells is injected.

  Further, the driving electrodes 51 and 51 and the crushing electrodes 56 and 56 are not limited to being integrally formed on the support 11 as film-like electrodes, but are configured as wire-like electrodes, for example, and inserted into the flow path 23 for voltage. May be applied. Further, the crushing electrodes 56 and 56 are not necessarily required. Depending on the cells to be crushed or when the flow velocity is high, only the collision action to the narrow portion 55 is applied without applying a crushing voltage. The cells can also be crushed with. Further, the narrow width portion 55 is not limited to the shape described above, and may be, for example, a shape protruding from only one of the opposing side walls of the flow path 23 or formed separately from the side wall.

  The arrangement of the flow path 23 and the solution tanks 21 and 21 is not limited to that shown in the embodiment. For example, a part in which a plurality of the flow paths 23 are arranged side by side or a part that performs an action other than crushing cells. May be provided instead of or in addition to the crushing electrodes 56 and 56 and the narrow width portion 55.

  Moreover, the constituent material of the flow path substrate 1 is not limited to that shown in the above embodiment.

  For example, the support 11 may be made of any material having a necessary strength as a support. However, in addition to glass (borosilicate glass, quartz glass, etc.), for example, plastic (polystyrene, polymethyl methacrylate). , Polysulfone, polyester, etc.) and glass fiber and plastic composites can also be used.

  The film electrode may be a general electrode material as well as the above materials, but the surface portion has a relatively high standard electrode potential (such as a positive value) such as Pt, Au, or Ag. ) It is preferable to use a material because it can prevent electrolytic corrosion when exposed to the sample solution. In addition, when a transparent electrode such as ITO (Indium Tin Oxide) is used, the transparency of the flow path substrate 1 can be maintained, which is preferable when optical analysis of the flow path substrate 1 is performed. Moreover, although it can improve adhesiveness with the support body 11 by forming by sputtering, it can also form by not only this but chemical vapor deposition, ion plating, and other physical vapor deposition.

  The photosensitive resin is not limited to the negative photosensitive resin that cures the portion irradiated with light as shown in the above embodiment, and a positive photosensitive resin that dissolves the portion irradiated with light is also used. Can do. However, from the viewpoint of securing the thickness and strength of the layer, a negative photosensitive resin that becomes a cross-linked polymer by a polymerization reaction during photocuring is preferable. The polymerization reaction may be any of radical polymerization, anionic polymerization, cationic polymerization and the like. As the photosensitive resin forming the crosslinked polymer, a known photosensitive resin containing a monomer and / or oligomer as a main component and further containing a photopolymerization initiator and various additives (stabilizer, filler, pigment, etc.) should be used. Can do. As this monomer, for example, (meth) acrylic monomers such as diethylene glycol di (meth) acrylate and trimethylolpropane tri (meth) acrylate can be used. Moreover, as an oligomer, the (meth) acrylic acid ester of an epoxy resin, the (meth) acrylic acid ester of a polyether resin, and the polyurethane resin which has a (meth) acryloyl group in a molecule terminal can be used, for example. As the photopolymerization initiator, for example, a benzoin photopolymerization initiator (benzoin, benzoin methyl ether, etc.) or an acetophenone photopolymerization initiator (2-2′-diethoxyacetophenone, etc.) can be used.

  <Second Embodiment>

  In the second embodiment, the above-described flow path substrate is provided with cooling means such as a Peltier element in the flow device.

  FIG. 8 is a functional block diagram of the flow device according to the second embodiment, FIG. 9 is a view showing the appearance of the flow device ((a) is a front view, (b) is a plan view, and (c) is a side view). FIG. 10 is a diagram showing a schematic configuration of a Peltier element used in the present embodiment.

  As shown in FIGS. 9A to 9C, in the flow device of the present embodiment, a support plate 33b is horizontally attached to an L-shaped base 50, and the upper surface side of the support plate 33b. A flat Peltier element 31 is fixed to the upper surface of the Peltier element 31, and the flow path substrate 1 is placed so as to be in close contact with the upper surface of the Peltier element 31. A connector 41 is attached on the support plate 33b, and the flow path substrate 1 is inserted into a connection port of the connector 41. The connector 41 can be connected to the voltage application device 42 shown in FIG. 8 at the connection port 41b on the opposite side, and electrically connects the flow path substrate 1 and the voltage application device 42. A water cooler body 33a is attached below the support plate 33b.

  The flow path substrate 1 is formed with a flow path 23 along a plane, and is configured so that the sample solution introduced into the flow path 23 can flow by applying a voltage. The configuration is the same as in FIG.

  The Peltier element 31 is a thermoelectric conversion element that absorbs or dissipates heat by the Peltier effect, and has a configuration as shown in FIG. 10 in the present embodiment. A Peltier element is used as the surface. FIG. 10 shows a part of the Peltier element 31, and the Peltier element 31 includes a plurality of P-type thermoelectric semiconductors 31p and N-type thermoelectric semiconductors 31n arranged between a pair of substrates 31a and 31a, and these are arranged on the substrate 31a. The stacked copper electrodes 31c are alternately connected in series. As shown in FIG. 10, when a current is passed from the N-type thermoelectric semiconductor 31n toward the P-type thermoelectric semiconductor 31p, the surface facing the upper side of the paper becomes the heat absorbing surface and the surface facing the lower side of the paper becomes the heat radiating surface. On the other hand, when a current flows in the opposite direction, the surface facing the upper side of the paper becomes a heat radiating surface, and the surface facing the lower side of the paper becomes a heat absorbing surface. In the flow device, as described above, the Peltier element 31 is brought into close contact with the flow path substrate 1 to adjust the temperature of the flow path substrate 1.

  Further, a temperature sensor 34 is attached to the upper surface side of the Peltier element 31, as shown in FIG. 9B, so that the temperature of the contact surface with the flow path substrate 1 can be detected. The temperature sensor 34 uses, for example, a thermistor whose resistance changes according to temperature. In addition, the kind of temperature sensor to be used is not limited to this.

  Further, the Peltier element 31 is electrically connected to a Peltier controller 32 shown in FIG. 8 through a pair of conducting wires 31b and 31b. The drive current from the Peltier controller 32 is received and adjusted to a target temperature. The As shown in FIG. 8, the Peltier controller 32 receives the detection signal of the temperature sensor 34, and based on this, calculates the magnitude of the drive current for bringing the flow path substrate 1 to the target temperature. The drive current adjusted to the magnitude is output. This Peltier element 31 corresponds to a cooling means.

  As shown in FIGS. 9A to 9C, the water cooler 33 includes a metal support plate 33b and a water cooler body 33a attached to the lower surface side of the support plate 33b. The water cooler body 33a is provided with a coolant passage in the metal block, and the coolant introduced from the inlet 33c is discharged from the outlet 33d through the coolant passage. Yes. This cooling liquid may be drawn directly from the water supply, or a secondary cooling device provided with a pump or a radiator may be provided to circulate the cooling liquid cooled by the secondary cooling device. This water cooler 33 corresponds to the second cooling means.

  When using the flow device having the above configuration, the flow path substrate 1 is attached to the connector 41, the voltage application device 42 is turned on, and a drive voltage is applied to the drive electrodes 51, 51 of the flow path substrate 1. Then, the sample solution introduced into the recess 2 is driven. At the same time, the Peltier controller 32 is also turned on. When the Peltier controller 32 detects that the endothermic surface of the Peltier element 31 exceeds the preset temperature, the Peltier element 31 is driven to cool the flow path substrate 1. As a result, the flow path substrate 1 is kept at a preset temperature or lower, so that evaporation of the sample solution is suppressed. For this reason, when the work requires a long time, the sample solution does not dry up during the work, and the work can be performed while maintaining a certain amount of solution. In this case, the cells can be crushed by applying a high-frequency voltage to the crushing electrodes 56, 56 of the flow path substrate 1 from a device similar to the crushing voltage applying device 53 of FIG.

  Note that the present invention is not limited to the second embodiment. For example, the means for cooling the flow path substrate 1 is not limited to the Peltier element 31, and for example, a heat sink is disposed so as to contact the flow path substrate, and the fan is blown toward the heat sink to cool it. May be. However, there is a risk that the sample solution may be easily dried unless air is applied to the flow path substrate 1 during air cooling. In this respect, it is preferable to use the Peltier element 31. Further, when the Peltier element 31 is used, high-precision cooling is possible, so that it is possible to maintain a limited temperature range even when handling a biopolymer having a high temperature sensitivity, and the apparatus can be downsized and quiet. In addition, it is preferable because it can operate without vibration.

  9A to 9C has a configuration in which only one flow path substrate 1 is mounted, but a configuration in which a plurality of connectors 41 are provided and a plurality of flow path substrates 1 can be mounted simultaneously. It is good. In this case, a plurality of Peltier elements 31 may be provided and cooled for each flow path substrate 1, or a plurality of flow path substrates 1 may be mounted on a single Peltier element 31. If a plurality of Peltier elements 31 are provided and can be controlled for each Peltier element 31, each flow path substrate 1 can be cooled with high accuracy.

  Further, in order to enhance the adhesion between the Peltier element 31 and the flow path substrate 1, a mechanism for pressing the flow path substrate 1 downward may be provided in the connector 41, or silicon grease may be applied.

  Further, the flow path substrate 1 is not limited to the above configuration. For example, the number of the flow paths 23 may be two or more. In the second embodiment, the temperature sensor 34 is provided on the Peltier element 31 side. However, the temperature sensor 34 may be provided on the lower surface of the flow path substrate 1 or may be formed integrally with the recess 2 of the flow path substrate 1. Good. Further, the flow path substrate 1 and the Peltier element 31 may be integrally formed on one chip.

  Next, the present invention will be described more specifically with reference to examples, but the present invention is not limited to the examples.

  In this example, a flow path substrate having the same configuration as in FIG. 2 was prepared, and a DNA solution was introduced into the flow path to which silica beads were fixed, and DNA was detected. That is, an adsorption test using a DNA fragment solution sample that is fixed by hooking a porous silica bead having a diameter of 10 μm in a fine channel having a width of 20 μm on the channel substrate and dyed with a cyanine nucleic acid staining reagent (Y3601 manufactured by Invitrogen). Carried out. The state of the silica beads before and after the introduction of the DNA fragment solution was observed with a fluorescence microscope, and the brightness was measured from a gray scale. The brightness of the silica beads indicates the amount of DNA adsorbed on the bead surface.

  DNA with fragment lengths of (1) 48 kbp (bp is the number of base pairs in base pair and corresponds to the length of DNA), (2) 2 to 33 kbp, (3) 0.1 to 1.3 kbp of DNA FIG. 11 shows the results of testing by introducing an equal amount of each of the three types of samples including fragments into the flow path substrate. In FIG. 11, the horizontal axis represents the length of the introduced DNA fragment sample, the vertical axis represents the brightness of the beads after the introduction of each DNA fragment solution sample, and the bead in the case of introducing a blank solution containing no DNA fragments. Maximum fluorescence intensity ratio divided by brightness.

  According to FIG. 11, a rapid increase in sensitivity is observed in the sample containing the DNA fragment with the shortest DNA length (3), and the flow path substrate has a particularly short region (1.3 kbp or less). It was found to be suitable for the detection of DNA fragments.

  As described above, the best modes and examples for carrying out the present invention have been described, but the present invention is not limited to these, and various modifications are possible within the scope of the technical idea of the present invention. Such modifications are also within the scope of the present invention.

It is a functional block diagram of the cell crushing device concerning a 1st embodiment. FIG. 3A is a plan view of a flow path substrate according to the first embodiment and FIG. 2B is a diagram illustrating a stacked configuration of the flow path substrate of FIG. It is the II sectional view taken on the line of Fig.2 (a). It is the schematic which shows the liquid quantity adjustment apparatus which concerns on 1st Embodiment. It is a figure which shows the pump of the liquid quantity adjusting device of FIG. It is a figure which shows the micropipette of the liquid quantity adjustment apparatus of FIG. It is a figure which shows the arrangement position of the micropipette and pressure sensor in 1st Embodiment. It is a functional block diagram of the flow apparatus of a 2nd embodiment. It is a figure ((a) is a front view, (b) is a top view, (c) is a side view) which shows the external appearance of the flow apparatus of FIG. It is a figure which shows schematic structure of the Peltier device used by 2nd Embodiment. It is a graph which shows the relationship between the length of the DNA fragment sample tested in the present Example, and the maximum fluorescence intensity ratio.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Flow path board | substrate 2 Concave part 6 Liquid quantity adjusting device 7 Pressure sensor 8 Controller 11 Support body 12 Photosensitive resin layer 21 Solution tank 23 Channel 51 Driving electrode 52 Driving voltage application apparatus 53 Crushing voltage application apparatus 55 Narrow part 56 Crushing Electrode 61 Micropipette 62 Tube 63 Pump 64 Coupling 65 Pump drive device 31 Peltier device 33 Water cooler

Claims (11)

A flow path substrate in which at least one pair of solution tanks and a flow path communicating with the one pair of solution tanks are formed in a concave shape;
Driving means for driving the sample solution introduced into the pair of solution tanks or the flow path from one of the solution tanks to the other by electroosmotic flow or electrophoresis;
The sample solution is partially injected or removed so that a pressure that hinders the driving force of the driving means does not occur due to the difference in the amount of the sample solution in the pair of solution tanks and flow paths generated during the driving. And a fluid amount adjusting means.   The flow apparatus according to claim 1, further comprising a determination unit that determines whether or not a pressure that inhibits the driving force is generated.   The flow apparatus according to claim 2, wherein the determination unit performs the determination based on a pressure of the sample solution detected in the pair of solution tanks or flow paths.   4. The liquid quantity adjusting unit according to claim 1, wherein the liquid amount adjusting unit sucks the sample solution in the downstream solution tank or injects the sample solution into the upstream solution tank. 5. The fluidizing device as described.   The flow apparatus according to claim 1, further comprising a cell crushing unit that crushes cells in the sample solution driven by the driving unit in the flow path.   The flow device according to any one of claims 1 to 5, further comprising a cooling unit that cools the flow path substrate.   The fluidizing device according to claim 6, wherein the cooling means is a Peltier element.   The flow device according to claim 7, wherein the Peltier element is disposed such that a heat absorption surface thereof is in contact with a bottom surface of the flow path substrate.   The fluidizer according to claim 7 or 8, further comprising a second cooling means for cooling the heat radiation surface of the Peltier element. An electroosmotic flow comprising at least one pair of solution tanks and a channel substrate in which a channel communicating with the pair of solution tanks is formed in a concave shape, and the pair of solution tanks or the sample solution introduced into the channel. Alternatively, in a flow device comprising a drive means for driving from one side of the solution tank to the other by electrophoresis, the sample solution is flowed,
The sample solution is partially injected or removed so that a pressure that hinders the driving force of the driving means does not occur due to the difference in the amount of the sample solution in the pair of solution tanks and flow paths generated during the driving. A flow method characterized by the above.   The flow method according to claim 10, wherein the flow path substrate is cooled to suppress evaporation of the sample solution.

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