Electrophoresis device and electrophoresis method

Abstract

This electrophoresis device (100) is provided with: a capillary tube; a pump unit (10) that fills the capillary with an electrophoretic separation medium; an automatic sampling unit (20) that conveys the sample to a position at which the sample is injected into the capillary filled with the electrophoretic separation medium; an irradiation detection means (30) that irradiates the sample, which has been subjected to electrophoresis in the capillary tube, with light and detects the obtained fluorescence; a high-voltage power supply unit (40) that generates a high-voltage power supply applied to both ends of the capillary tube; and a thermostatic bath unit (50) for maintaining the temperature of the capillary within a predetermined range, the pump unit (10) having: an opening / closing mechanism capable of controlling the opening / closing of a flow path formed in the pump unit from the outside; and a pressurizing / depressurizing mechanism for pressurizing / depressurizing the inside of the flow path closed by the opening / closing mechanism, the opening / closing mechanism and the pressurizing / depressurizing mechanism being configured so as to compress bubbles in the flow path when the pressurizing / depressurizing mechanism pressurizes the inside of the flow path, and to depressurize the inside of the flow path when the pressurizing / depressurizing mechanism pressurizes the inside of the flow path. The air bubbles are discharged by re-operating the pressurizing / depressurizing mechanism and the opening / closing mechanism by pressurizing the inside of the flow path and opening the atmosphere using the pressurizing / depressurizing mechanism and the opening / closing mechanism, and by peeling, separating, and moving the air bubbles from the inner wall surface of the flow path using the flow rate generated by the pressure difference between the inside and outside of the flow path and by re-operating the pressurizing / depressurizing mechanism and the opening / closing mechanism.

Description

Technical Field

[0001] The present invention relates to an electrophoresis device and an electrophoresis method. Background Art

[0002] Electrophoresis devices analyze DNA in a sample by applying a high voltage to a flow path filled with a polymer serving as a separation medium, causing electrophoresis. Applying the high voltage while air bubbles remain within the polymer can cause discharge, potentially damaging the device. To address this issue, methods for removing air bubbles from the polymer have been proposed.

[0003] Patent Document 1 discloses a structure for removing bubbles from a polymer block that constitutes a pump unit in an electrophoresis device. Specifically, the structure describes a structure in which the flow of polymer into a buffer container creates a negative pressure, causing polymer in a polymer bottle to flow through a check valve into the capillary tubes within the polymer block, thereby filling the capillary tubes with polymer and removing bubbles.

[0004] Furthermore, Patent Document 2 discloses a structure for removing bubbles to protect the electrophoresis device from discharge caused by residual bubbles. Specifically, the structure describes a structure in which a detachable special electrophoretic medium filling unit or electrophoretic medium container is connected to one end of the capillary tube only when the capillary tube is filled with electrophoretic medium, allowing electrophoresis to be performed with both ends of the capillary tube directly immersed in buffer. Prior art literature Patent Literature

[0005] Patent Document 1: Japanese Patent Application Laid-Open No. 2009-31310 Patent Document 2: Japanese Patent Application Laid-Open No. 2013-140181 Summary of the Invention Technical problem to be solved by the invention

[0006] Patent Document 1 describes a method for using a plunger pump in an electrophoresis device to reduce bubbles within a flow path filled with an electrophoretic separation medium. However, the bubble reduction mechanism in Patent Document 1 requires repeated operation of the plunger and a valve for controlling the pressure within the flow path to reduce bubbles. Furthermore, the bubbles must be expelled along with the polymer serving as the electrophoretic separation medium. This not only requires significant effort on the part of the user, but also results in a significant amount of polymer waste.

[0007] In addition, Patent Document 2 describes the following technical problem: when bubbles exist in the flow path, the flow path is electrically cut off by the bubbles, causing discharge. Depending on the size of the discharge, the electrophoresis device may be damaged. In this document, as a means to solve this technical problem, the following structure is proposed: only when the capillary is filled with electrophoresis medium, a detachable special electrophoresis medium filling unit or an electrophoresis medium container is connected to one end of the capillary, so that electrophoresis can be performed with both ends of the capillary directly immersed in buffer solution. However, this structure has the following problem: it cannot be applied to the prior art disclosed in this document. Figure 1 The structure of the electrophoresis device described in the above claims is that a phoretic medium filling unit and a phoretic medium container are prepared as independent components, and the phoretic medium container and the capillary are indirectly connected via a fixed phoretic medium filling unit having a flow path formed therein.

[0008] The present invention has been made to solve the above-mentioned problems, and an object thereof is to provide an electrophoresis apparatus and an electrophoresis method that can more easily remove bubbles in a flow path inside a phoretic medium-filled unit while suppressing the amount of polymer used to a small amount. Technical solutions to technical problems

[0009] To achieve the above-mentioned object, the electrophoresis device of the present invention includes: a capillary; a pump unit that fills the capillary with an electrophoretic separation medium; an automatic sampling unit that transports a sample to a position where the sample is injected into the capillary filled with the electrophoretic separation medium; an irradiation detection unit that irradiates light to the sample undergoing electrophoresis in the capillary and detects the resulting fluorescence; a high-voltage power supply unit that generates a high-voltage power supply applied to both ends of the capillary; and a constant-temperature bath unit that maintains the temperature of the capillary within a specified range, wherein the pump unit has an opening and closing mechanism. The flow path formed in the pump unit can be controlled to be opened and closed from the outside; and a pressure-increasing and pressure-reducing mechanism is provided, which pressurizes and depressurizes the flow path after being closed by the opening and closing mechanism. The opening and closing mechanism and the pressure-increasing and pressure-reducing mechanism have the following structures: when the pressure-increasing and pressure-reducing mechanism pressurizes the flow path, bubbles in the flow path are compressed; and the pressure-increasing and pressure-reducing structure and the opening and closing mechanism are controlled to pressurize the flow path and open it to the atmosphere, using the flow velocity generated by the pressure difference between the inside and outside of the flow path to cause bubbles to peel off, detach, and move from the inner wall surface of the flow path, and the pressure-increasing and pressure-reducing mechanism and the opening and closing mechanism are moved again to discharge the bubbles. Other aspects of the present invention will be described in the embodiments described below. Effects of the Invention

[0010] According to the present invention, it is possible to provide an electrophoresis apparatus and an electrophoresis method that can more easily remove bubbles in a flow path of a phoretic medium-filled unit while suppressing the amount of polymer used to a small amount. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1 1 is a structural diagram showing an example of an electrophoresis device according to an embodiment. Figure 2 It is an explanatory diagram showing an example of the structure of a plunger pump. Figure 3 It is an explanatory diagram showing the operating range of the plunger of the plunger pump. Figure 4 This is a flowchart showing an example of the operation from activation of the pump unit to start of measurement. Figure 5 This is a structural diagram showing an example of the pump unit according to the first embodiment. Figure 6 This is an operation flowchart showing an example of the operation of the pump unit according to the first embodiment. Figure 7 This is an explanatory diagram showing the movement of bubbles generated in the first embodiment. Figure 8 This is a structural diagram showing an example of a pump unit according to the second embodiment. Figure 9 This is an operation flowchart showing an example of the operation of the pump unit according to the second embodiment. Figure 10 This is an operation flowchart showing an example of the operation of the pump unit according to the third embodiment. Figure 11 This is a structural diagram showing an example of a pump unit according to the fourth embodiment. Figure 12 This is an operation flowchart showing an example of the operation of the pump unit according to the fourth embodiment. DETAILED DESCRIPTION

[0012] First, the details of the problems of conventional examples and the objectives and outline of this embodiment will be described. As described in Patent Document 1, a conventional structure involves flowing polymer from a syringe into a buffer container via a capillary tube within a polymer block and a flow path switching valve, thereby creating a negative pressure on the discharge side of the check valve. This causes polymer from a polymer bottle to flow into the capillary tube within the polymer block, thereby filling the capillary tube within the polymer block with polymer and removing air bubbles within the polymer block. Regarding air bubbles, as described in Patent Document 2, a known problem is that if air bubbles are present within the flow path of an electrophoretic device, the flow path may be electrically disconnected by the bubbles, generating a discharge. Depending on the magnitude of this discharge, the electrophoretic device may be damaged.

[0013] Furthermore, the electrophoresis device described in Patent Document 2 as a prior art, which presupposes that a phoretic medium filling unit and a phoretic medium container are prepared as separate components, and that the phoretic medium container and a capillary tube are indirectly connected via a fixed phoretic medium filling unit having a flow path formed therein, has been adopted by various types of electrophoresis devices to date, and has high versatility and excellent usability. However, if a structure is adopted in Patent Document 1 in which the polymer in the syringe is directed to the buffer container for bubble removal, polymer may be discharged into the buffer container without benefiting the electrophoresis device.

[0014] Therefore, a pump unit structure is being considered that can suppress the discharge amount of the polymer and solve the above-mentioned problem related to bubbles without significantly changing the electrophoresis apparatus.

[0015] That is, the electrophoresis device includes: an opening and closing mechanism, which can control the opening and closing of the flow path formed in the pump unit from the outside; and a pressurization and decompression mechanism, which pressurizes and decompresses the flow path after being closed by the opening and closing mechanism, and the opening and closing mechanism and the pressurization and decompression mechanism are controlled to compress the bubbles in the flow path when the pressurization and decompression mechanism pressurizes the flow path, and by using the pressurization and decompression structure and the opening and closing mechanism, the action part of the pressurization and decompression mechanism in the flow path, that is, the plunger, is moved from near the front end of the chamber to pressurize the flow path, and then the flow path is opened to atmospheric pressure, and the flow velocity generated by the pressure difference between the inside and outside of the flow path is used to cause the bubbles to peel off, detach, and move from the inner wall surface of the flow path, and the pump and the opening and closing mechanism are moved again to discharge the bubbles.

[0016] Here, as a closing mechanism, the pump unit and the capillary can be connected to each other through an opening, and the closing mechanism can be composed of a flow path resistance that prevents the electrophoretic separation medium (for example, a polymer) from flowing into the capillary from the pump unit and a control system that controls the pressure in the flow path of the pump unit. Alternatively, as a closing mechanism, a closing mechanism can be composed of a mechanism having a capillary valve that closes the pump unit from the capillary. In the former case, the number of components can be reduced and the present invention can be implemented by a simple device structure. On the other hand, in the latter case, a capillary valve needs to be added, but a higher closing performance can be obtained thereby, and it can be pressurized to a higher pressure. Therefore, when opened to atmospheric pressure, a faster flow can be obtained by a larger pressure difference.

[0017] Hereinafter, some embodiments of the electrophoresis apparatus and the electrophoresis method of the present invention will be described in detail with reference to the accompanying drawings.

[0018] Figure 1 1 is a structural diagram illustrating an example of an electrophoresis apparatus 100 according to an embodiment. The electrophoresis apparatus 100 includes a pump unit 10 that fills a capillary array 110 with an electrophoretic separation medium; an automatic sampling unit 20 that transports a sample to a position where the sample is injected into the capillary array 110 filled with the electrophoretic separation medium; an irradiation and detection unit 30 that irradiates the sample undergoing electrophoresis within the capillary array 110 with light and detects the resulting fluorescence (reflected light or transmitted light); a high-voltage power supply unit 40 that generates a high-voltage power supply applied to both ends of the capillary array 110; a constant-temperature bath unit 50 that maintains the temperature of the capillary array 110 within a predetermined range; and a control device 60.

[0019] The pump unit 10 includes a plunger pump 120 , an acrylic block 102 , a polymer valve 104 , a buffer valve 105 , a polymer bottle 106 , and a buffer container 107 . The capillary is filled with an electrophoretic separation medium such as a polymer. The buffer container 107 is provided with an anode-side electrode 402 .

[0020] Figure 2 1 is an explanatory diagram showing an example of the structure of the plunger pump 120 . Figure 3 : is an explanatory diagram showing the operating range of the plunger of the plunger pump. Figure 2 、 Figure 3 It is from Figure 1 The plunger pump 120 can be composed of a chamber 116, a plunger 101, a pump drive unit 118, etc. The plunger pump 120 can pull the plunger 101 in the chamber 116 by operating the pump drive unit 118 (see Figure 3 Pulling action state 3A) and pushing action (refer to Figure 3The driving speed of the plunger 101 can be freely adjusted. In addition, the action range of the plunger 101 of the plunger pump 120 is Figure 3 The middle is the range from the pulling action state 3A to the pushing action state 3B.

[0021] return Figure 1 The capillary head 111 at the front end of the capillary array 110 is connected to the acrylic block 102. A capillary sample introduction end-side electrode 401 is provided at the lower end of the capillary array 110. Flow resistance is generated at the capillary connection portion 115, which connects the capillary head 111 to the pump peripheral flow path 103. This prevents the polymer (electrophoretic separation medium) from flowing toward the capillary side below a certain pressure.

[0022] The automatic sampling unit 20 includes a sample container 201 for dispensing a sample, a buffer reservoir 202 for storing a buffer solution containing dissolved electrolytes, and an automatic sampler 203. The automatic sampler 203 transfers a container containing washing water for washing the front end of the capillary or a container containing waste liquid to or from the capillary sample introduction end 204.

[0023] The irradiation detection unit 30 includes a light source 302, such as a laser or LED, that irradiates excitation light toward the capillary detection section 301, and a signal detection mechanism 303 that detects fluorescence from the capillary detection section 301. The high-voltage power supply unit 40 includes a high-voltage power supply 403 that applies a high voltage between a capillary sample introduction end-side electrode 401 and an anode-side electrode 402. The constant-temperature bath unit 50 includes a constant-temperature bath 501 that houses the capillary array 110.

[0024] Figure 4 1 is a flowchart showing an example of the operation from the start of the pump unit 10 to the start of measurement. Figure 4 The operation of the electrophoresis device according to this embodiment will be described. The control device 60 is the main body (subject) of the operation.

[0025] In step S201 , the control device 60 operates the plunger 101 to inject new electrophoresis separation medium into the capillary. In step S202 , the sample container 201 is transported to the capillary sample introduction end 204 by the automatic sampler 203 , and the capillary sample introduction end 204 is immersed in the sample solution in the sample container.

[0026] In step S203 , a voltage is applied to the capillary to electrodynamically inject the sample into the capillary. In step S204 , the buffer container 107 is transported to the capillary sample introduction end 204 by the automatic sampler 203 . In step S205 , a streaming voltage is applied to the capillary to perform electrophoresis.

[0027] Implementation Method 1 Figure 5 1 is a diagram showing the configuration of the pump unit 10 according to the first embodiment. Figure 5 The pump unit 10 and Figure 1 same.

[0028] The plunger 101 is the driving part of the pump, and the driving speed can be freely adjusted for pushing and pulling. There is a pump peripheral flow path 103 inside the acrylic block 102, and the polymer flows in this flow path.

[0029] A polymer valve 104 is provided in the flow path between the inlet for the polymer, which is the electrophoretic separation medium enclosed in the polymer bottle 106, and the insertion portion of the plunger 101, which is the driving portion of the pump. The polymer valve 104 has a check valve structure, and the inflow of polymer is controlled by opening and closing the valve in the pump peripheral flow path 103.

[0030] A buffer valve 105 is provided between the outflow port from which the polymer flows out and the capillary connection portion 115 , which is a connection portion between the capillary array 110 and the pump peripheral flow channel 103 , where a DNA sample is tested by electrophoresis.

[0031] Figure 6 This is an operation flowchart showing an example of the operation of the pump unit according to the first embodiment. Figure 6 , an example of an opening and closing table of the operation of the plunger 101 and two valves (polymer valve 104 and cushion valve 105) is shown. Figure 7 This is an explanatory diagram showing the movement of bubbles generated in the first embodiment.

[0032] The control device 60 (pump unit 10) uses the pulling action of the plunger 101 to cause the polymer to flow (suck) from the inlet into the chamber 116 (step S11). The plunger 101 is then pushed to guide the polymer toward the outlet (step S12). The user confirms whether the flow path has been filled with polymer (step S13). If not (step S13, No), the process returns to step S11. If so (step S13, Yes), the process proceeds to step S14. The control device 60 repeats this pushing and pulling action of the plunger 101 until the entire flow path is filled with polymer.

[0033] After confirming that the entire flow path is filled with polymer, the controller 60 pulls the plunger 101 once. Conventionally, the buffer valve 105 is then opened and the plunger 101 is pushed. At this point, the polymer valve 104, which acts as a check valve, automatically closes, causing the polymer to flow into the buffer container 107 rather than the polymer bottle 106. At this point, bubbles within the polymer are also guided into the buffer container 107 along with the polymer, and are removed from the pump peripheral flow path 103.

[0034] On the other hand, in this embodiment, after confirming that the polymer has filled the entire flow path in step S14, the control device 60 controls the position of the plunger 101 so that it is located near the front end of the chamber 116, which is the area where the plunger 101 moves (see FIG. Figure 7 The flow path is closed by the plunger 101 because the polymer valve 104 is a check valve.

[0035] The controller 60 then controls the amount of plunger 101 pushed in, pushing the plunger 101 so that the flow resistance within the closed pump peripheral flow path 103 reaches a pressure that prevents the polymer from flowing toward the capillary. This increases the pressure within the flow path, compressing any remaining bubbles 117.

[0036] The control device 60 opens the buffer valve 105 once the flow path is pressurized, opening the flow path to atmospheric pressure, thereby causing the polymer to flow. Figure 7 As in the pressurized state 7B, the bubbles 117 trapped in the pump peripheral flow path 103 are peeled off, detached, and moved (step S15). Then, as in the conventional process, the plunger 101 is pulled (step S16), the buffer valve 105 is opened, and the bubbles 117 generated in the polymer are guided to the buffer container 107 along with the polymer. Thus, the bubbles 117 are removed from the pump peripheral flow path 103 (step S17). The control device 60 then pulls the plunger 101 again (step S18), closes the polymer valve 104 and the buffer valve 105 again, and performs the plunger pushing operation (step S19). This allows the capillary array 110 to be guided to a higher filling degree with fewer bubbles.

[0037] Conventional methods require simultaneous removal of a portion of the polymer when removing air bubbles. Furthermore, if air bubbles cannot be removed with a single plunger push-pull operation, one of several possible countermeasures, previously explored as an example, involves performing multiple plunger push-pull operations. However, this approach is not preferred due to the increased number of steps required by the user and the amount of polymer discharged.

[0038] As described above, in this embodiment, the flow path 103 is pressurized while the polymer is not flowing out of the pump peripheral flow path. The flow generated by the subsequent release of the atmosphere is then used to move trapped bubbles and reduce the amount of movement of the plunger 101. This reduces polymer consumption and allows for efficient air bubble expulsion.

[0039] The opening and closing mechanism (e.g., the buffer valve 105) and the pressure-increasing and decompressing mechanism (e.g., the plunger pump 120) of the electric device 100 of this embodiment have the following structure: they are controlled so as to compress the bubbles in the flow path when the pressure-increasing and decompressing mechanism pressurizes the flow path, and the pressure-increasing and decompressing structure and the opening and closing mechanism are used to pressurize the flow path and open it to the atmosphere, and the bubbles are peeled off, detached, and moved from the inner wall surface of the flow path by utilizing the flow velocity generated by the pressure difference between the inside and outside of the flow path, and the pressure-increasing and decompressing mechanism and the opening and closing mechanism are moved again to discharge the bubbles.

[0040] After confirming that the electrophoretic separation medium fills the entire flow path, the control device 60 closes the polymer valve 104 and the buffer valve 105, uses the pressure increase and pressure reduction mechanism to pressurize the flow path to compress the bubbles in the flow path, opens the buffer valve 105 to open the flow path to the atmosphere, and uses the flow rate generated by the pressure difference between the inside and outside of the flow path to cause the bubbles to peel off, detach, and move from the inner wall surface of the flow path.

[0041] As described above, according to this embodiment, by reducing the amount of movement of the plunger 101 for removing bubbles, the outflow of polymer to the outside can be reduced, bubbles can be suppressed, and the waste of expensive polymer can be suppressed, which contributes to cost reduction. In addition, by starting to pressurize near the front end of the chamber 116 (see Figure 7 ), so when the flow path is opened to atmospheric pressure, the flow rate of the polymer flowing in the narrow area near the front end of chamber 116 increases, thereby detaching and discharging bubbles adhering to or stagnating on the wall surface of chamber 116. Furthermore, since this embodiment begins pressurizing near the front end of chamber 116, the volume of polymer within chamber 116 at the start of pressurization is relatively small. Therefore, when the pressure is released to atmospheric pressure, the entire polymer within chamber 116 flows, enabling the expulsion of bubbles regardless of where they are stagnant. In other words, this embodiment achieves its effects regardless of the placement of plunger pump 120.

[0042] Implementation Method 2 In Embodiment 2, as a modification of Embodiment 1, the pump unit 10 includes a capillary valve 112 as a component. An example using this valve will be described as Embodiment 2.

[0043] This embodiment differs from the first embodiment in that it includes a capillary valve 112 and there is no restriction on the driving speed of the plunger 101. However, other parts are the same as those of the first embodiment.

[0044] Figure 8 1 is a structural diagram showing an example of a pump unit 10A according to Embodiment 2. The pump peripheral flow path 103 includes a plunger 101 as a pump driving portion, a polymer valve 104 , a buffer valve 105 , and a capillary valve 112 .

[0045] Figure 8A capillary valve 112 is newly provided in the pump peripheral flow path 103. Other structures are the same as those already described. Figure 1 The structures shown with the same reference numerals attached have the same functions, and thus their descriptions are omitted.

[0046] The capillary valve 112 has a capillary connection portion 115 (see Figure 1 ) is the connection between the capillary head 111, the distal end of the capillary array 110, which performs electrophoresis and testing of DNA samples, and the pump peripheral flow path 103. This valve can be operated to open or close in order to prevent polymers flowing in the pump peripheral flow path 103 from undesirably flowing toward the capillary side.

[0047] Figure 9 This is an operation flowchart showing an example of the operation of the pump unit according to the second embodiment. Figure 9 , an example of an opening and closing table of the operation of the plunger 101 and three valves (polymer valve 104, buffer valve 105, and capillary valve 112) is shown.

[0048] The control device 60 (pump unit 10A) uses the pulling action of the plunger 101 to cause the polymer to flow (suck) from the inlet into the chamber 116 (step S21). The plunger 101 is then pushed to guide the polymer toward the outlet (step S22). The user confirms whether the flow path has been filled with polymer (step S23). If not (step S23, No), the process returns to step S21. If so (step S23, Yes), the process proceeds to step S24. The control device 60 repeats this pushing and pulling action of the plunger 101 until the entire flow path is filled with polymer.

[0049] After confirming that the polymer has filled the entire flow path, the control device 60 controls the position of the plunger 101 so that it is located near the front end of the chamber 116, which is the area where the plunger 101 moves (see Figure 7 (Pre-pressurization state 7A). At this time, the buffer valve 105 and the capillary valve 112 are closed. The polymer valve 104 is a check valve, so the flow path is closed relative to the pushing action of the plunger 101.

[0050] The front end of the plunger 101 is controlled to push inside the closed pump peripheral flow path 103 (inside the chamber 116), thereby pressurizing the flow path and compressing the bubbles remaining in the flow path to the maximum extent possible (step S24).

[0051] After pressurizing the flow path, the control device 60 opens the buffer valve 105 once, opening the flow path to atmospheric pressure, thereby causing the polymer to flow. Figure 7As in the pressurized state 7B, the bubbles trapped in the pump peripheral flow path 103 are peeled off, detached, and moved (step S25). Then, as in the conventional method, the plunger 101 is pulled (step S26), the buffer valve 105 is opened, and the bubbles generated in the polymer are also guided to the buffer container 107 along with the polymer. In this way, the bubbles are removed from the pump peripheral flow path 103 (step S27). The control device 60 then pulls the plunger 101 again (step S28), closes the polymer valve 104 and buffer valve 105 again, opens the capillary valve 112, and performs the plunger pushing operation (step S29). This allows the capillary array 110 to be guided to a higher filling degree with fewer bubbles.

[0052] As described above, by reducing the amount of plunger movement required to remove bubbles, the outflow of polymer to the outside is reduced, bubbles are suppressed, and thus, the waste of expensive polymer is prevented, contributing to cost reduction. Furthermore, compared to Embodiment 1, Embodiment 2 includes a capillary valve 112, thereby increasing the pressurization within the flow path. This increases the velocity of the flow generated after release to the atmosphere, making it easier to remove difficult-to-remove bubbles.

[0053] Implementation Method 3 In the third embodiment, as a modification of the first embodiment, an example is described in which bubbles adhering to and accumulating in the flow channel are peeled off, separated, or moved by pressurizing or depressurizing the flow channel to discharge the bubbles.

[0054] The difference between the third embodiment and the first embodiment lies in the step before the bubbles are peeled off and released. However, the structural components of the other parts are the same.

[0055] Figure 10 This is an operation flowchart showing an example of the operation of the pump unit according to the third embodiment. Figure 10 , an example of an opening and closing table of the operation of the plunger 101 and two valves (polymer valve 104 and cushion valve 105) is shown.

[0056] As in the first embodiment, the control device 60 repeats the push-pull operation of the plunger 101 to fill the entire flow path with the polymer.

[0057] After confirming that the polymer has completely filled the flow channel, the control device 60 closes the buffer valve 105 and pulls the plunger 101 once, controlling and pushing the closed pump peripheral flow channel 103 (step S31). This increases pressure within the flow channel, minimizing the compression of any remaining bubbles. This compression reduces the volume of the bubbles, reducing the contact area between the bubbles and the flow channel walls, making them easier to detach and break away.

[0058] After pressurizing the flow path, the amount of pull-in is controlled, and plunger 101 is pulled so that polymer valve 104 remains closed while maintaining its closed state (step S32). This reduces the pressure in the flow path, causing bubbles remaining in the flow path to expand. As the bubbles expand, their buoyancy increases, making them more likely to detach and escape.

[0059] The user determines whether the bubbles are peeled off or detached from the retention area (step S33). If the bubbles are not peeled off or detached from the retention area (step S33, no), the process returns to step S31. If the bubbles are peeled off or detached from the retention area (step S33, yes), the process proceeds to step S16.

[0060] Repeating this process of pressurizing and depressurizing the flow path causes bubbles trapped in the pump peripheral flow path 103 to separate and escape. Then, as in the conventional method, the plunger 101 is pulled (step S16), and the buffer valve 105 is opened (step S17). This directs the bubbles generated within the polymer into the buffer container 108 along with the polymer, removing the bubbles from the pump peripheral flow path 103. The plunger 101 is pulled again (step S18), and the polymer valve 104 and buffer valve 105 are closed again, followed by a plunger push operation (step S19). This allows the capillary array 110 to be filled with polymer with a higher degree of filling, after reducing the number of bubbles.

[0061] As described above, by reducing the amount of polymer consumed when pushing and pulling the plunger 101 to release bubbles, the outflow of polymer throughout the entire series of operations can be reduced, thereby suppressing waste of expensive polymer and contributing to cost reduction.

[0062] Implementation Method 4 In Embodiment 4, a modification of Embodiment 1 described above and an example of the present invention, a gas phase flow path 113 and an outside air valve 114 (outside air opening and closing mechanism) are provided as components in the pump unit 10. This flow path and an example using this valve will be described as Embodiment 4.

[0063] Figure 11 1 is a structural diagram showing an example of a pump unit 10B according to Embodiment 4. Embodiment 4 includes a gas phase flow path 113 and an external air valve 114, and differs from Embodiment 1 in the method of discharging bubbles.

[0064] The pump peripheral flow path 103 includes a plunger 101 as a pump driving portion, a polymer valve 104 , a cushion valve 105 , a gas phase flow path 113 , and an external air valve 114 . Figure 11 A gas phase flow path 113 and an external air valve 114 are newly provided in the pump peripheral flow path 103. Other structures are the same as those already described. Figure 1The structures shown with the same reference numerals attached have the same functions, and thus their descriptions are omitted.

[0065] The pump peripheral flow path 103 includes a gas phase flow path 113 near the polymer valve 104 . The external air valve 114 is provided in the gas phase flow path 113. The opening and closing state of this valve can be manipulated to prevent the polymer flowing in the pump peripheral flow path from undesirably flowing into the gas phase flow path 113 and to control the pressure in the flow path.

[0066] Figure 12 This is an operation flowchart showing an example of the operation of the pump unit according to the fourth embodiment. Figure 12 , an example of an opening and closing table of the operation of the plunger 101 and three valves (polymer valve 104, cushion valve 105, and external air valve 114) is shown.

[0067] The control device 60 (pump unit 10B) draws (suctions) polymer from the inlet by pulling the plunger 101 (step S41). The control device 60 guides the polymer toward the outlet by pushing the plunger 101 (step S42). The user confirms whether the flow path has been filled with polymer (step S43). If not (step S43, No), the process returns to step S41. If filled (step S43, Yes), the process proceeds to step S44. The control device 60 repeats this push-pull action of the plunger 101 until the entire flow path is filled with polymer.

[0068] After confirming that the entire flow path is filled with the polymer, the control device 60 closes the buffer valve 105, opens the external air valve 114, pulls the plunger 101, and sucks the gas phase into the pump peripheral flow path 103 (step S44).

[0069] During the plunger pulling operation, the external air valve 114 is closed to guide the gas phase into the pump peripheral flow path 103 (step S45).

[0070] The controller 60 opens the buffer valve 105 and performs a plunger push operation. This action discharges the gas phase into the buffer container 107 (step S47). At this point, bubbles remaining in the pump peripheral flow path 103 are trapped when they come into contact with the gas phase, thus discharging even difficult-to-discharge bubbles. This allows the capillary array 110 to be filled with a higher polymer density after reducing the number of bubbles.

[0071] The electrophoresis device of this embodiment has the following structure: the opening and closing structure for external air (for example, the external air valve 14) is controlled to draw a certain amount of gas phase from the gas phase flow path 113 into the flow path, and is controlled to intentionally draw the gas phase into the flow path using the pressurization and decompression mechanism, the opening and closing mechanism and the opening and closing mechanism for external air, and then the pressurization and decompression mechanism and the opening and closing mechanism are used to control the movement of the gas phase to discharge bubbles.

[0072] While some embodiments of the present invention have been described above, the present invention is not limited to these embodiments. For example, configurations obtained by appropriately combining Embodiment 1, Embodiment 2, Embodiment 3, and Embodiment 4 with other elements also fall within the scope of the present invention.

[0073] Finally, the electrophoresis method is further explained. (1) An electrophoresis method using an electrophoresis device having a capillary includes the following steps: a filling step (e.g., steps S11 to S13) of driving a pump until a flow path provided in a pump unit 10 is filled with an electrophoretic separation medium; a closing step (e.g., step S14) of closing the flow path provided in the pump unit (10); and a pressurizing step (e.g., step S15) of compressing bubbles in the flow path closed by the closing step by applying pressure to the flow path near the front end of the chamber 116, which is an operating portion of the pressurizing and depressurizing mechanism in the flow path, using a pressurizing and depressurizing mechanism (e.g., plunger pump 120). a detachment step (e.g., step S14) of detaching the bubbles from the retained area in the flow path pressurized by the pressurization step by utilizing the flow generated by opening the opening and closing mechanism (e.g., buffer valve 105); and a discharge step (e.g., step S17) of controlling the pressure in the flow path using the pressurization and decompression mechanism and the opening and closing mechanism, and discharging the bubbles by reciprocating the piston, i.e., plunger 101, of the pressurization and decompression mechanism within a range controlled to discharge all the electrophoretic separation medium from the position of the detached bubbles to the discharge port, while making a small amount of push-out.

[0074] Thus, by starting pressurization near the front end of chamber 116, the flow rate of the polymer flowing in the narrow region near the front end of chamber 116 increases when the flow path is opened to atmospheric pressure. This allows bubbles adhering to and stagnating on the wall surfaces of chamber 116 to be detached, released, and discharged. Furthermore, since pressurization starts near the front end of chamber 116, the volume of polymer within the chamber at the start of pressurization is relatively small. Consequently, when atmospheric pressure is released, the entire electrophoretic separation medium (e.g., polymer) within the chamber flows, enabling the discharge of bubbles regardless of their location. (2) In the electrophoresis method of (1), the pump unit 10 pressurizes the flow path formed and closed by flow path resistance between the capillary and the pump peripheral flow path 103 and then opens it to the atmosphere, thereby allowing bubbles to escape from the retained area in the flow path, thereby reducing the bubbles in the electrophoretic separation medium. (3) In the electrophoresis method of (1), the opening and closing mechanism and the pressure-increasing and decompression mechanism are controlled to compress the bubbles in the flow path when the pressure-increasing and decompression mechanism pressurizes the flow path, and are controlled to pressurize and decompress the flow path using the pressure-increasing and decompression structure and the opening and closing mechanism. The bubbles in the flow path are repeatedly compressed and expanded, thereby causing the bubbles to peel off, detach, and move from the inner wall surface of the flow path, and the pressure-increasing and decompression mechanism and the opening and closing mechanism are moved again to discharge the bubbles. (4) In the electrophoresis method of (3), the pump unit 10 has a pressurization and decompression mechanism having a plunger 101 that performs a pumping action portion in the pump unit 10. The plunger 101 pressurizes and decompresses the flow path closed by the opening and closing mechanism to compress and expand the bubbles. Label Description

[0075] 3A Pull action state 3B Push action status 7A State before pressurization 7B Pressurized state 10, 10A, 10B pump units (phoretic medium filling units) 20 Automatic sampling unit 30 Irradiation detection unit 40 High voltage power supply unit 50 Constant temperature bath unit 60 Control Device 100 electrophoresis device 101 Plunger 102 acrylic blocks 103 Pump peripheral flow path 104 polymer valve (opening and closing mechanism) 105 Buffer valve (opening and closing mechanism) 106 polymer bottles 107 Buffer Container 110 Capillary array 111 Capillary Head 112 Capillary valve (opening and closing mechanism) 113 Gas Phase Flow Path 114 External air valve (opening and closing mechanism for external air) 115 Capillary connection 116 Chamber 117 Bubbles 118 Pump drive unit 120 plunger pump (pressure increasing and reducing mechanism) 201 sample container 202 Buffer storage 203 Automatic Sampler 204 Capillary sample introduction port 301 Capillary Detection Department 302 light source 303 Signal Detection Agency 401 Capillary sample introduction end electrode 402 anode side electrode 403 High Voltage Power Supply 501 Constant temperature bath.

Claims

1. An electrophoresis device, characterized in that: include: capillary; a pump unit that fills the capillary with an electrophoretic separation medium; an automatic sampling unit, which transports a sample to a position where the sample is injected into the capillary filled with the electrophoretic separation medium; an irradiation detection unit that irradiates the sample electrophoresed in the capillary with light and detects the resulting fluorescence; a high-voltage power supply unit that generates a high-voltage power supply to be applied to both ends of the capillary; and a constant temperature bath unit for maintaining the temperature of the capillary tube within a specified range, The pump unit has: an opening and closing mechanism capable of externally controlling the opening and closing of a flow path formed in the pump unit; and a pressurization and decompression mechanism for pressurizing and decompressing the flow path closed by the opening and closing mechanism; The opening and closing mechanism and the pressurization and decompression mechanism have the following structure: they are controlled to compress the bubbles in the flow path when the pressurization and decompression mechanism pressurizes the flow path, and are controlled to use the pressurization and decompression structure and the opening and closing mechanism to pressurize the flow path and open it to the atmosphere, and the bubbles are peeled off, detached, and moved from the inner wall surface of the flow path by utilizing the flow velocity generated by the pressure difference between the inside and outside of the flow path, and the pressurization and decompression mechanism and the opening and closing mechanism are moved again to discharge the bubbles.

2. The electrophoresis device according to claim 1, wherein The pressurization and depressurization mechanism of the pump unit includes a plunger that performs a pumping operation portion within the pump unit. The plunger compresses and expands bubbles by increasing and decreasing pressure in the flow path closed by the opening and closing mechanism.

3. The electrophoresis device according to claim 2, wherein: The opening and closing mechanism includes, within the flow path, a polymer valve mounted on the flow path into which the electrophoretic separation medium flows, and a buffer valve connected to a flow path toward a discharge port provided in the pump unit.

4. The electrophoresis device according to claim 1, wherein The pump unit and the capillary are connected to each other through an opening. No opening and closing mechanism is provided at the capillary connection portion, and flow path resistance is used to prevent the electrophoresis separation medium from flowing to the capillary side.

5. The electrophoresis device according to claim 3, wherein: The electrophoresis device also has a control device, After confirming that the electrophoretic separation medium has filled the entire flow path, the control device closes the polymer valve and the buffer valve, uses the pressure-increasing and pressure-reducing mechanism to pressurize the flow path to compress the bubbles in the flow path, opens the buffer valve to open the flow path to the atmosphere, and uses the flow rate generated by the pressure difference between the inside and outside of the flow path to cause the bubbles to peel off, detach, and move from the inner wall surface of the flow path.

6. The electrophoresis device according to claim 3, wherein: The opening and closing mechanism further includes a capillary valve that seals between the pump unit and the capillary.

7. The electrophoresis device according to claim 1, wherein: The pump unit also has: a gas phase flow path for sucking a gas phase into the flow path; and The external air opening and closing mechanism controls the pressure in the gas phase flow channel and can be opened and closed from the outside. It has the following structure: the opening and closing mechanism for external air is controlled to draw a certain amount of gas phase from the gas phase flow path into the flow path, and is controlled to use the pressurization and decompression mechanism, the opening and closing mechanism and the opening and closing mechanism for external air to intentionally draw the gas phase into the flow path, and then use the pressurization and decompression mechanism and the opening and closing mechanism to control the movement of the gas phase to discharge bubbles.

8. An electrophoresis method using an electrophoresis device having a capillary tube, characterized in that: include: a filling step of driving the pump until the flow path provided in the pump unit is filled with the electrophoresis separation medium; a closing step of closing the flow path provided in the pump unit; a pressurizing step of compressing bubbles by applying pressure from the vicinity of the front end of the chamber, which is an operating portion of the pressurizing / depressurizing mechanism, in the flow path closed by the closing step; a detachment step of detaching the bubbles from the retained area in the flow path pressurized in the pressurizing step by utilizing the flow generated by the opening of the opening and closing mechanism; as well as The pressure in the flow path is controlled by the pressurization and decompression mechanism and the opening and closing mechanism. The electrophoretic separation medium is completely discharged from the position of the detached bubbles to the discharge port, and the piston, i.e., the plunger, of the pressurization and decompression mechanism is reciprocated within a small range to discharge the bubbles.

9. The electrophoresis method according to claim 8, wherein The pump unit reduces bubbles in the electrophoretic separation medium by pressurizing the closed flow path formed by flow path resistance between the capillary and the pump peripheral flow path and then opening it to the atmosphere to release bubbles from stagnant areas in the flow path.

10. The electrophoresis method according to claim 8, wherein The opening and closing mechanism and the pressurization and decompression mechanism are controlled to compress the bubbles in the flow path when the pressurization and decompression mechanism pressurizes the flow path, and are controlled to use the pressurization and decompression structure and the opening and closing mechanism to pressurize and decompress the flow path. The bubbles in the flow path are repeatedly compressed and expanded, so that the bubbles are peeled off, detached, and moved from the inner wall surface of the flow path, and the pressurization and decompression mechanism and the opening and closing mechanism are moved again to discharge the bubbles.

11. The electrophoresis method according to claim 10, wherein The pressurization and depressurization mechanism of the pump unit includes a plunger that performs a pumping operation portion within the pump unit. The plunger compresses and expands bubbles by increasing and decreasing pressure in the flow path closed by the opening and closing mechanism.

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