JP3136062U - Electrophoresis device with easy bubble removal - Google Patents

Abstract

Bubbles can be removed from a capillary array connection portion.
The capillary array connection portion is connected so that the tip of the capillary array faces obliquely upward, and a flow path structure is formed so that the polymer solution flows upward from the lower side of this portion when bubbles are removed. Also, the bubbles could be easily removed by opening and closing the bubble removal screw. The total cost of the analysis could be reduced by using an inexpensive buffer solution. The bubbles were easily removed by collecting the bubbles in one of the bubble reservoirs by circulating the flow path. By providing a valve for switching between the polymer solution and the buffer solution, a flow channel configuration has been devised in which bubbles in the flow channel are removed by continuing to flow an inexpensive buffer solution.
[Selection] Figure 3

Description

  The present invention relates to an electrophoresis apparatus.

  Capillary electrophoresis is widely used as a technique for separating and analyzing many biological samples including deoxyribonucleic acid (DNA). One of its technical advantages is the heat dissipation characteristics resulting from the surface area of the capillary versus the volume ratio. This characteristic makes it possible to achieve high-speed and high-resolution separation with a high voltage. Furthermore, in the capillary electrophoresis system, a high-throughput electrophoresis apparatus based on the multicapillary system has also been put into practical use, which has a feature that it is easy to perform multi-analysis using a large number of capillaries simultaneously.

  Here, the capillary is a thin tube having an inner diameter of several tens to several hundreds of microns, and the main material is quartz.

  Since the outer side of quartz is coated with polyimide having a thickness of about several tens of microns, the mechanical strength is improved. During electrophoresis, the capillary is filled with a component (polymer solution) that serves as a sample separation medium.

  In some cases, non-flowable cross-linked polymers have been used as sample separation media. However, in recent years, non-cross-linkable flowable polymer solutions with excellent productivity and performance stability have become mainstream. Yes. The electrophoresis apparatus using the fluid polymer solution as described above is provided with an injection pump for filling the capillary with the polymer solution. For example, 3730 manufactured by Applied Biosystems, Inc. injects a fluid polymer solution serving as a sample separation medium into the capillary array by an injection pump using a sapphire plunger.

  In the capillary electrophoresis apparatus, the length and number of capillaries can be changed in accordance with the type of analysis and analysis conditions required by the user.

  In order to meet the needs of users, devices capable of mounting various types of capillary arrays with a single device have been put into practical use.

  For example, in the electrophoresis apparatus disclosed in Japanese Patent Application Laid-Open No. 2004-144479, 48 or 96 capillary arrays can be mounted, and several types of capillaries can be used for each. In addition, since the capillary array has a structure that can be replaced by a user as a consumable item, the ease of replacement of the capillary array is an important point in evaluating the operability of the apparatus.

  One of the important points when evaluating the ease of replacement of the capillary array is the removal of bubbles that have entered the flow path when the capillary array is attached. In the electrophoresis device, a high voltage of several kilograms to several tens of kilovolts is applied between both ends of the electrophoresis path, so in particular, air intrusion into the electrophoresis path may be discharged inside the acrylic flow path block. May cause problems. Usually, the connection part of the capillary array becomes a part of the electrophoresis path. Therefore, when attaching the capillary array, it is necessary to reliably remove bubbles from the flow path part and the connection part.

  The electrophoresis apparatus disclosed in Japanese Patent Application Laid-Open No. 2004-144479 has a structure in which a capillary array is fixed at two locations, a sample introduction end and a polymer solution supply end, and is held by an optical detection unit that emits a laser. Yes. The polymer solution supply end is connected to the infusion pump of the apparatus.

  Specifically, the tip of the polymer solution supply end is inserted horizontally into an acrylic flow path block and is sealed and connected by a sleeve and a push screw. The acrylic channel block is provided with a polymer container for storing the polymer solution and an injection pump for injecting the polymer solution into the capillary array at a high pressure. The injection pump is connected to the capillary array through a flow path in an acrylic block.

  Since an electrode on the anode side is inserted into the buffer solution, the flow path leading from the capillary array to the buffer solution on the polymer solution supply end side becomes an electrophoresis path, and a high voltage is applied during electrophoresis. . In the apparatus, when the user attaches the capillary array, the air bubbles must be removed from the piping serving as the electrophoresis path.

  Specifically, the removal of the bubbles will be described. The polymer solution sucked by the injection pump is pressurized by the injection pump and then reaches the capillary array through the flow path. By supplying the polymer solution into the acrylic channel block in which the capillary array is inserted, bubbles are discharged together with the polymer solution through the buffer solution tube for the electrophoresis path using the flow of the polymer solution. Was. The flow path diameter of the buffer solution tube is as small as about 1 mm, but is sized so that bubbles can be easily removed. The capillary array connection portion must be a relatively wide space in order to accommodate the diameter of the bundle of capillary arrays. Further, in this portion, the polymer solution flows from the infusion pump (upper left) to the polymer solution outlet (lower right). In the flow of the polymer solution, it is difficult to remove the bubbles because the bubbles are in a direction opposite to the direction in which bubbles rise. In some cases, a large amount of expensive polymer solution is consumed in order to remove the bubbles because the bubbles remaining in the capillary array connection portion are not easily removed.

JP 2004-144479 A

  In an apparatus in which a capillary array to which the present invention belongs is attached to the apparatus and the bubbles in the electrophoresis path including the capillary array connection portion need to be removed, the bubbles can be easily removed in order to improve the operability of the apparatus. Ingenuity is required. Also, the flow path configuration must be simple and the maintenance of the components must be easy.

  The present invention relates to facilitating removal of bubbles from an electrophoresis path.

  The present invention relates to a flow path configuration of an injection pump for supplying a polymer solution to a capillary array in a capillary electrophoresis apparatus. The capillary array connection part is a part where the polymer solution supply end of the capillary array is connected to the flow path system of the injection pump and is used as an electrophoresis path.

  The buffer solution is a buffer solution that is connected to the flow path of the injection pump mechanism and serves as an anode during electrophoresis.

  In the present invention, at the portion where the capillary array is connected to the infusion pump, the tip of the capillary head of the capillary array is prevented from adhering bubbles by making it conical, and the tip of the capillary head is tilted downward. By inserting it into the acrylic flow path block in the upward direction, a flow of bubbles is created to make it easier for the bubbles to rise.

  Preferably, further, the inner surface of the acrylic flow path block into which the capillary array is inserted has a corrugated surface (for example, a rounded tip of the crest with a thin screw), and further the surface of the inner surface is roughened to further reduce the bubbles. It is characterized in that the surface tension is reduced and bubbles are easily separated from the inner wall of the inner surface. Furthermore, by providing a funnel-shaped cylinder for collecting bubbles when the bubbles rise above the capillary head, there is an effect of enlarging the bubbles by collecting and integrating small bubbles in the funnel-shaped cylinder. Large bubbles collected in the funnel-shaped cylinder are easily sucked with a bubble suction syringe by opening and closing a bubble removal screw. Furthermore, by making the piping position for injecting the polymer solution a flow path system that flows upward from the lower side of the capillary head, it is possible to create a flow of bubbles near the tip of the capillary head, and to create more bubbles It floats in a funnel-shaped bubble reservoir.

  Furthermore, a flow for circulating the polymer solution or the buffer solution near the capillary head by the difference in the injection pressure from the injection pump by utilizing the difference between the flow path diameter of the injection pump and the circulation channel diameter by providing a circulation channel. The flow path system was made. Furthermore, the bubbles near the tip of the capillary head can be easily lifted and the bubbles can be easily guided to the bubble reservoir. The generated bubbles are collected in a bubble reservoir below the bubble removal screw in the middle, and the bubbles can be easily removed by opening and closing the bubble removal screw.

  Furthermore, by switching the polymer solution sucked from the pump for injection with a switching valve, the expensive polymer solution is switched to an inexpensive buffer solution, and the bubbles are removed by continuing to flow the buffer solution until the bubbles can be removed. To do.

  Furthermore, it is possible to supply a polymer solution or an inexpensive buffer solution to the capillary head by using a circulation channel and using a manual syringe instead of an injection pump.

  Furthermore, it is characterized in that the bubble that has entered by being injected into the polymer solution supply end with a manual syringe can be lifted, and the bubble can be easily removed from the bubble removal screw.

  However, when air bubbles are removed with the buffer solution, it is necessary to replace it with the polymer solution. A buffer solution having a lighter density than the polymer solution can be easily removed by opening a bubble removal screw.

  In the electrophoresis apparatus equipped with the polymer injection mechanism of the present invention, when the capillary array is attached, bubbles can be easily removed from the portion where the capillary array is connected by using a bubble removal screw. Further, the capillary array can be easily attached and the preparation time before measurement can be shortened. Furthermore, the amount of polymer used for removing bubbles can be reduced and the economic efficiency is improved (since the polymer solution is usually very expensive).

  FIG. 1 shows a configuration diagram of a capillary electrophoresis apparatus according to the present invention. The capillary electrophoresis apparatus includes a capillary array 119, an excitation optical system including a laser light source 124, a light receiving optical system 117 for detecting fluorescence, and a high voltage applying unit 125 for performing electrophoresis. The capillary array 119 has a structure in which the capillaries 120 (capillaries with an inner diameter of several tens to several hundreds of μm) are arranged in a plane, and a sample (fluorescent sample) labeled with a phosphor from a direction parallel to the arrangement surface of the capillaries 120 is provided. Laser light is irradiated to the fluorescent samples in all the capillaries 120 by irradiating the filled capillaries with laser light and condensing the laser light by the lens action of the capillaries. The fluorescent sample irradiated with the laser light emits fluorescence and emits light in a direction substantially perpendicular to the irradiation direction of the laser light. This is a device for measuring by detecting fluorescence from a fluorescent sample with a light receiving optical system 117. An injection pump 116 for injecting a highly viscous polymer solution 101 (hereinafter referred to as a polymer solution) serving as an electrophoretic separation medium is provided. The injection pump 116 performs an operation of sucking the polymer solution 101 from the bottle 102 containing the polymer solution 101 (hereinafter referred to as a polymer container) and injecting it into the capillary array 119 by operating the injection pump 116. It is also used when air bubbles in the acrylic flow path block 111 are removed.

  The outline of the apparatus configuration will be described below. One end (sample injection end) 121 of the capillary array 119 is immersed in the buffer solution 123 in the buffer container 122, and the other end is connected to the acrylic flow path block 111. In addition to the capillary array 119, the acrylic flow path block 111 includes an injection pump 116, a manual syringe 108 that can inject a buffer solution or a polymer solution, a bubble suction syringe 110, a check valve 106, and a check valve. A valve 107 and a bubble removal screw 109 are included. One is a polymer container 102 containing a polymer solution 101 to be injected into a capillary array via a switching valve 105 directly connected to the check valve 106, and the other one of the switching valves 105 is a buffer solution 103 dedicated to removing air bubbles. Is placed, and each component forms a flow path through a tube for piping. The acrylic flow path block 111, the polymer container 102, and the buffer container 104 are connected to a check valve 106 via a switching valve 105.

  The check valve 106 prevents back flow into the polymer container 102 and the buffer container 104 when the polymer is injected from the injection pump 116 into the capillaries of the capillary array 119.

  An electric buffer valve 113 is disposed between the buffer container 126 and the acrylic flow path block 111. When the polymer solution 101 is injected into the capillaries 120 of the capillary array 119, the electric buffer valve 113 is closed. Thus, the polymer solution 101 is filled in the flow path between the capillary array 119 and the buffer container 122. Further, during electrophoresis, the electric buffer valve 113 is opened and the flow path is opened to allow the capillary array 119 and the buffer container 126 to communicate with each other.

  FIG. 2 shows an external view of the capillary array. The following will be described with reference to FIGS. Each capillary 203 constituting the capillary array has an outer diameter of about 0.1 to 0.7 mm, an inner diameter of about 0.02 to 0.5 mm, and the outer coat is coated with a polyimide resin. The capillaries themselves are quartz pipes, and a plurality of capillaries (96 in this example) are arranged to constitute a capillary array. The capillary array is a load header 202 for taking a sample into a capillary by an electrical action from a sample container containing a fluorescently labeled DNA sample or the like, and a detection unit (window) for arranging and fixing the capillaries 203 in order of sample numbers of the load header 202 Unit) 206 and a capillary head 205 in which a plurality of capillaries 203 are bundled and bonded. A hollow electrode 201 for applying a migration voltage to the capillary is provided at the sample injection end protruding from the load header 202. The detection unit (window unit) 206 includes an opening 204 for irradiating the aligned capillary array with a laser beam from the side and an opening 207 for taking out light emitted from the capillary.

  The capillary head 205 of the capillary array is connected to the acrylic flow path block 111. After the sleeve is attached to the circular capillary head 205 in which the capillaries are bundled together, the push screw is tightened to deform the sleeve. By filling the gap, the acrylic channel block 111 can be attached.

  In the capillary array shown in FIG. 2, the capillary 203 protrudes several millimeters from the tip of the hollow electrode 201 integrated with the load header 202. The DNA sample placed in the sample container with the sample injection end fluorescently labeled from the tip of the capillary 203 is immersed in the buffer container 122 connected to the acrylic flow path block 111 and the load header 202. A high voltage of several kV is applied to the hollow electrode 201 from the high voltage applying unit 125, and the sample in the sample container is injected into each capillary of the capillary array 119. Thereafter, the sample injection end of the capillary array 119 is immersed in the buffer container 122, and the samples injected into the respective capillaries 120 are separated by electrophoresis.

  The laser light emitted from the laser light source 124 is guided to the detection unit (window unit) 206 by the excitation optical system and irradiated. Fluorescence, which is signal light emitted from a sample migrating in the capillary due to excitation light irradiation, is detected by the light receiving optical system 117 via the detection optical system.

  The apparatus configuration shown here shows one specific example, and does not impose conditions or restrictions on the present invention.

  The capillary array is a replaceable part, and one having a different number or length of capillaries is selected depending on the purpose of use by the user. The capillary array is also replaced when it is deteriorated due to contamination or the like. The replacement of the capillary array is usually performed by the user, but it is necessary to remove the air that has entered the acrylic flow path block 111 when the capillary array is attached. This is because the flow path of the acrylic flow path block 111 is a portion to which a high voltage is applied using the flow path between the capillary array 119 and the buffer container 126 as an electrophoresis path, and air remains as bubbles. Measurement may be hindered due to discharge or the like. When the apparatus is used for the first time, it is necessary to clean and dry the parts constituting the flow path of the acrylic flow path block 111. After washing the parts, it is necessary to fill the polymer solution 101 in all the flow paths including the capillary array connection portion in which the capillary array 119 is attached to the acrylic flow path block 111. At that time, the polymer solution 101 is injected using the injection pump 116.

  When the capillary array is exchanged, the bubbles mixed in the acrylic flow path block 111 are discharged by pouring the polymer solution 101 from the injection pump 116 after the capillary array exchange. If there is a portion where bubbles remain partially, the polymer solution 101 is further poured and the bubbles are pushed out by the polymer solution. As a result, a large amount of valuable polymer solution 101 may be consumed.

  In the configuration as in the embodiment shown in FIG. 1, the bubbles are discharged as follows. First, the capillary array 119 is connected and the electric buffer valve 113 is opened. The polymer solution 101 is sucked from the injection pump 116 and the bubbles that have entered the flow path are discharged from the buffer container side 126 together with the polymer solution 101. Usually, the diameter of the flow path in the acrylic flow path block 111 is made as small as about 1 mm so that bubbles can be easily removed, and the flow path is made to flow uniformly in the acrylic flow path block.

  On the other hand, the portion where the capillary array 119 is connected cannot be easily reduced because a space for accommodating the capillary head 112 is required. In addition, since the polymer solution 101 flowing through the conventional capillary array connection portion is a flow path that flows from the upper side to the lower side, air bubbles are pushed downward in the polymer solution 101. Bubbles were stuck in the migration path. As a result, in the acrylic channel block 111, it is one of the portions where bubbles are most likely to remain. As a result, if there are portions where bubbles are difficult to escape, a large amount of polymer solution may be used to remove bubbles.

  Therefore, a flow path configuration as shown in FIG. 3 was devised as the present invention.

  The point of the configuration of FIG. 3 is that the capillary array 119 is connected so as to face obliquely upward, and the flow path configuration is such that the polymer solution 102 is sent from the lower side to the capillary array connection portion. Here, the infusion pump 116 plays the same role as the manual syringe 108 of FIG. With this arrangement, bubbles in the capillary array connection portion are pushed up from below by the polymer solution 101 and discharged into the upper flow path 301. That is, the flow path arrangement utilizing the property that the above-mentioned bubbles float upward in the polymer solution 101 was adopted. Bubbles can be easily discharged from the capillary array connection part, resulting in improved usability of the apparatus and a reduction in the amount of polymer solution required to remove bubbles, improving the economics of the apparatus. .

  Specific contents will be described with reference to FIGS. 1 and 3.

  In the portion where the capillary array 119 is connected to the injection pump 116, the tip of the capillary head 112 of the capillary array 119 is conical to prevent the attachment of bubbles, and further, the tip of the capillary head 112 is inclined downward. It is made easier to create a flow of bubbles by inserting it into the acrylic flow path block 111 upward.

  Furthermore, the inner surface of the acrylic flow path block 111 into which the capillary array 119 is inserted has a corrugated surface (for example, a rounded tip at the top of the crest with a thin screw), and further the surface of the inner surface is roughened so that the air bubbles are adhered. The feature is that bubbles are easily separated from the inner wall of the inner surface. Further, by providing a funnel-shaped tube 302 for collecting air bubbles when air bubbles rise above the capillary head 112, the effect of enlarging the bubbles by collecting and integrating small bubbles in the funnel-shaped tube 302 is effective. is there. Large bubbles collected in the funnel-shaped tube 302 are easily sucked by the bubble removing syringe 110 by opening and closing the bubble removing screw 109. Furthermore, the flow of bubbles near the tip of the capillary head 112 can be created by making the piping position for injecting the polymer solution 101 a flow path system that flows upward from the lower side of the capillary head 112. It is characterized in that air bubbles are floated in the funnel-shaped bubble reservoir. Further, by providing the circulation channel 303, the difference between the channel diameter of the injection pump 116 and the circulation channel diameter makes it possible to use the difference in the injection pressure from the injection pump 116 to cause the polymer solution 101 near the capillary head 112 or the buffer. By creating a flow that circulates the solution 103, bubbles near the tip of the capillary head 112 can be easily lifted and the bubbles can be easily introduced into the bubble reservoir. The generated bubbles are collected in a bubble reservoir below the bubble removal screw 109 in the middle, and the bubbles are easily removed by opening the bubble removal screw 109.

  Further, by switching the polymer solution 101 sucked from the injection pump 116 with the switching valve 105, the expensive polymer solution 101 is switched to the cheap buffer solution 103, and the buffer solution 103 is kept flowing until the bubbles can be removed. It is characterized by removing.

  Further, the polymer solution or the inexpensive buffer solution can be directly injected into the capillary head 112 by using the circulation channel and using the manual syringe 108 instead of the injection pump 116. Furthermore, by using the manual syringe 108, the invading air bubbles float up in the air bubble reservoir, and the air bubbles can be easily removed from the air bubble removal screw 109.

  However, when bubbles are removed with the buffer solution 103, it is necessary to replace them with the polymer solution 101.

  The buffer solution 103 having a lighter density than the polymer solution 101 can be easily removed by opening the bubble removal screw 109.

The block diagram of a capillary electrophoresis apparatus. The external view of a capillary array. The flow-path block diagram of the acrylic flow-path block regarding this invention.

Explanation of symbols

DESCRIPTION OF SYMBOLS 101 Polymer solution 102 Polymer container 103,115,123 Buffer solution 104,122,126 Buffer container 105 Switching valve 106,107 Check valve 108 Manual syringe 109 Bubble extraction screw 110 Bubble suction syringe 111 Flow path block 112 made of acrylic, 205 Capillary head 113 Electric buffer valve 114 Pipe tube 116 Injection pump 117 Light receiving optical system 118 Detection unit 119 Capillary array 120, 203 Capillary 121 Sample injection end 124 Laser light source 125 High voltage application unit 201 Hollow electrode (sample injection end)
202 Load header 204 Opening 206 for irradiating laser light Detection unit (window unit)
207 Opening 301 for extracting emitted light The upper channel (bubble reservoir)
302 funnel 303 circulation channel

Claims (5)

In a capillary electrophoresis apparatus equipped with an injection pump for injecting a polymer solution as a sample separation medium into a capillary array,
Capillary array tip is made of a curved part at the tip of the capillary head. By inserting the capillary array into the acrylic channel block obliquely and downwardly, air bubbles can be easily detached from the capillary head surface. Electrophoresis device. The electrophoresis apparatus according to claim 1,
1. An electrophoretic device, characterized in that corrugated surface irregularities are provided on an inner surface of an acrylic flow path block so as to facilitate separation of bubbles. The electrophoresis apparatus according to claim 1,
By providing a switching valve in the infusion pump, the polymer solution and the buffer solution are easily switched, and the bubbles are removed by continuing to flow the buffer solution instead of continuing to flow the polymer solution and removing bubbles. Electrophoresis device. The electrophoresis apparatus according to claim 1,
An electrophoresis apparatus, wherein bubbles in the acrylic flow path block can be easily sucked with a bubble suction syringe by opening and closing a bubble release screw. The electrophoresis apparatus according to claim 1,
A metal for transmitting vibration to the vicinity of the capillary head tip is disposed in the acrylic flow path block, and by applying vibration to the vibration transmitting metal, bubbles closely attached to the vicinity of the capillary head tip are easily detached. Electrophoresis device characterized by floating.

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