CAPILLARY ELECTROPHORESIS DEVICE

Abstract

The present disclosure provides a capillary electrophoresis device including a light source, a first irradiation fiber configured to guide light from the light source, a detector configured to detect light from a capillary, and a first detection fiber configured to guide light to the detector to make an optical light detection system less susceptible to an external environment. A capillary cartridge including a capillary, a second irradiation fiber, and a second detection fiber is attached to the capillary electrophoresis device by connecting the first irradiation fiber and the second irradiation fiber, and connecting the first detection fiber and the second detection fiber. In the capillary cartridge, the second irradiation fiber and the second detection fiber are fixed such that their optical axes intersect each other in the lumen of the capillary.

Description

Technical area

[0001] The present disclosure relates to a capillary electrophoresis device, in particular to a device that optically detects an analyte in a capillary. State of the art

[0002] In analysis using capillary electrophoresis, a sample to be analyzed is injected into a capillary filled with a separation medium, and a voltage is applied to both ends to perform separation based on the difference in the mobility of the analysis target. There are a variety of means for measuring the separated sample, including a method for detecting fluorescence emitted by the sample, a method for detecting light absorption by the sample, and the like.

[0003] For example, there is a method in which DNA labeled with a fluorescent dye is subjected to electrophoresis in a polymer-filled capillary to separate each chain length. A detection site provided on the capillary is irradiated with excitation light, and the generated fluorescence is detected. The DNA molecules in the sample move through the capillary and pass the detection site at different times depending on the chain length. As a result, the chain length distribution of the DNA molecules in the sample is recorded as a fluorescence intensity waveform.

[0004] In a capillary electrophoresis device, the inner diameter of a capillary for separating a sample is generally several tens of µm. To increase detection sensitivity, it is desirable to irradiate the capillary inner diameter through which the sample passes with fluorescence excitation light as losslessly as possible. Therefore, it is desirable that the excitation light be condensed to an extent equal to or smaller than the inner diameter of the capillary and applied to the inner diameter of the capillary. At this time, the positions of the capillary and the condensed excitation light must be adjusted with a positional accuracy equal to or greater than the size of the capillary inner diameter.In particular, when a configuration is adopted in which a plurality of capillaries are arranged in a line and excitation light is incident from their side surfaces to excite all capillaries at once, the excitation light must pass through all capillary inner diameters as losslessly as possible. Therefore, the demand for positioning accuracy becomes greater, and the positioning error must be approximately 10 µm or less.

[0005] In a case where there are a variety of fluorescent dyes to be labeled on the measurement target, the generated fluorescence is introduced into a spectroscopic optical system and measured by an imaging element after wavelength separation. As an example, spectroscopy is performed through a grating. In this case, when the position of the light emission point in the capillary changes in the wavelength separation direction, the fluorescence wavelength appears to be shifted on a detector. Since the type of fluorescent dye that emitted light is distinguished from the spectral shape of the measured light, the discrimination accuracy of the fluorescent dye decreases as the fluorescence wavelength shifts.For example, a decrease in dye discrimination accuracy may lead to erroneous detection of a DNA strand that does not originally exist in the DNA analysis, which is undesirable.

[0006] From the above, the positional relationship between the excitation light irradiation optical system, the capillary, and the detection optical system must be precisely adjusted with an error of about several tens of μm or less. In addition, it is undesirable for the detection performance of the device to change, and it is desirable that the positional relationship be maintained even if the device is used for a long period of time or the device is moved. On the other hand, in capillary electrophoresis, the capillary is a consumable item and deteriorates when the measurement is performed a certain number of times, so replacement is required. Therefore, it is necessary to maintain the above positional accuracy even if the capillary is replaced. From the perspective of user convenience, it is desirable that capillary replacement can be performed easily. Citation listPatent literature PTL 1: JP 8-304339 A PTL 2: JP 2004-532384 A PTL 3: US 2021 / 0003530 A1 Nicht-Patentliteratur

[0007] NPL 1: H. Zhai et al., „A simple and compact fluorescence detection system for capillary electrophoresis and its application to food analysis“, Electrophoresis, 36, 2509 (2015). Zusammenfassung der ErfindungTechnisches Problem

[0008] A capillary used for electrophoresis, an optical system that irradiates the capillary with light, and an optical system that detects light are typically integrated and fixed in an electrophoresis device. However, the position of each component and the optical element within the component may change due to vibration or shock transmitted from the outside, such as when the device is moved. In addition, even when no vibration or shock is applied, a change in the position of the optical element may occur due to expansion or contraction of each element due to an external environment, especially a temperature change.

[0009] Replacing the capillary tube may also cause a change in position. The position of the capillary tube may change from that before replacement due to a tolerance in the outer diameter of the capillary tube to be installed, an error in the fixing position generated in a capillary fixing mechanism, and the like. When there are multiple capillaries, the capillary assembly is often delivered to the user in a state where the capillaries are fixed to a fastener, but the position of the capillary tube assembly may change due to an error in the assembly of the fastener and a main body.

[0010] A method for reducing the occurrence and influence of the positional error as described above by mounting an optical fiber system on an electrophoresis channel has been proposed. For example, PTL 1 discloses a method in which an optical fiber for performing excitation and detection is installed on a channel chip, and a capillary, which is a consumable, is attached to the channel chip so that the relative positional relationship of the optical detection system does not change even if the capillary is replaced. However, this method has a problem in that the separation performance of electrophoresis is deteriorated when an event such as the generation of a gap between channels at a junction between a capillary and a channel chip or the shift of the central axes of the capillary and the channel chip occurs.Therefore, it is desirable that the electrophoresed sample be detected at a detection point provided in a portion of the capillary. Furthermore, in the method of PTL 1, when a plurality of capillaries are mounted, it is necessary to assemble a plurality of disclosed structures, resulting in a large device size. Furthermore, the excitation light must be branched by the number of detection fibers, resulting in a problem that the excitation light power is reduced and the detection performance is deteriorated.

[0011] PTL 2 describes a method for reducing the alignment accuracy required at the time of capillary replacement by providing a cartridge in which optical components, such as an optical fiber and a lens, are mounted on a capillary. In the configuration disclosed in PTL 2, a detection fiber assembly including a light source, such as an LED, and a lens is connected to a capillary cartridge containing an irradiation fiber. In this configuration, the relative positional relationship between the capillary and the irradiation fiber hardly changes. However, the positional adjustment accuracy of the capillary and the detection fiber assembly, and the light source and the irradiation fiber at the time of cartridge replacement depends on the accuracy of the detachable mechanical fastening mechanism.Given that the capillary cartridge is a consumable, the mechanical fastening mechanism must be cost-effective. This mechanical fastening mechanism must be robust against external vibrations and shocks and must always maintain the positional relationship of each component. Patent 2 does not disclose that the described fastening mechanism meets the above requirement.

[0012] PTL 1 describes a method for performing detection by mounting a capillary and irradiation and detection fibers on a plate having a groove formed from polydimethylsiloxane. Since the capillary and the optical element are mounted on the same base, this method hardly causes positional shift of the optical element due to vibration and shock. However, a method for capillary replacement is not described, nor is a method for adjusting the relative positional relationship between the capillary and the optical system at the time of capillary replacement. In addition, when the number of capillaries is multiple, a problem similar to that of PTL 1 may occur.

[0013] In capillary electrophoresis, it is necessary to adjust the temperature of the capillary tube uniformly. For example, when analyzing DNA by capillary electrophoresis, the capillary temperature is maintained at approximately 60°C, so that electrophoresis is performed in a state where DNA is denatured. To maintain the separation efficiency of electrophoresis and return the same measurement result every time for the same sample, it is necessary for the entire capillary to have a uniform temperature and low temperature fluctuation. This temperature adjustment mechanism must also achieve both temperature adjustment performance and ease of capillary replacement.

[0014] Regarding capillary temperature adjustment, PTL 1 and PTL 2 each describe a capillary temperature adjustment method, but do not mention that the disclosed structure is particularly advantageous for temperature adjustment. NPL 1 does not describe capillary temperature adjustment. Solution to the problem

[0015] An example of a capillary electrophoresis device according to the present disclosure is an electrophoresis device that includes: a light source; a first irradiation fiber configured to guide light from the light source; a detector configured to detect light from a capillary; and a first detection fiber configured to guide light to the detector, in which a capillary cartridge including the capillary, a second irradiation fiber, and a second detection fiber is attached to the electrophoresis device by connecting the first irradiation fiber and the second irradiation fiber and connecting the first detection fiber and the second detection fiber, and in the capillary cartridge, the second irradiation fiber and the second detection fiber are fixed such that optical axes of the second irradiation fiber and the second detection fiber intersect each other in an inner cavity of the capillary. Advantageous effects of the invention

[0016] According to the capillary electrophoresis apparatus of the present disclosure, it is possible to make the light detection optical system in the capillary electrophoresis apparatus less susceptible to changes in the external environment, shocks, and vibrations.

[0017] Since the main body and the optical system of the capillary cartridge can be connected by the optical fiber connection, the capillary cartridge can be easily attached and removed.

[0018] In the configuration of the present disclosure, a detection window of the capillary can be isolated from the external environment. Therefore, the temperature of the entire capillary, including the measurement window, can be precisely adjusted.

[0019] The configuration of the present disclosure can be expanded even when the number of capillaries is multiple. By applying the excitation light from the side surface of the capillary array, the reduction in excitation light power due to the division of the excitation light can be prevented even when the number of capillaries is large. Short description of the drawings Fig. 1] Fig. 1 is a configuration diagram of an electrophoresis apparatus 100 according to a first embodiment of the present disclosure. [ Fig. 2] Fig. 2 is a diagram illustrating a measurement flow by the electrophoresis apparatus 100 according to the first embodiment of the present disclosure. [ Fig. 3] Fig. 3 illustrates a method for fixing capillaries 111 and a cartridge-side irradiation fiber 114 at a detection site 116. [ Fig. 4] Fig. 4 illustrates a method for fixing cartridge-side detection fibers 115 at the detection site 116. [ Fig. 5] Fig. 5 illustrates a method for fixing the capillaries 111, the cartridge-side irradiation fiber 114, and the side detection fibers 115 at the detection site 116. [ Fig. 6] Fig. 6 is a schematic view illustrating a connecting portion of a capillary cartridge 110. [ Fig. 7] Fig. 7 is a schematic view illustrating a method for adjusting the temperature of the capillaries 111. [ Fig. 8] Fig. 8 is a configuration diagram of an electrophoresis apparatus 800 according to a second embodiment of the present disclosure. [ Fig. 9] Fig. 9 is a structural diagram of the electrophoresis apparatus 800 and a capillary cartridge 801 according to the second embodiment of the present disclosure. [ Fig. 10] Fig. 10 is a schematic view illustrating a method for fixing the detection site 116. [ Fig. 11] Fig. 11 is a diagram illustrating a structure of a detection site 116 according to a third embodiment of the present disclosure. [ Fig. 21 Fig. 12 is a diagram illustrating an arrangement of optical fibers in a case of performing both fluorescence measurement and absorption measurement. [ Fig. 13] Fig. 13 is a diagram illustrating a structure of a fixing substrate 1301 according to a fourth embodiment of the present disclosure. [ Fig. 14] Fig. 14 is a diagram illustrating an arrangement of capillaries 1401, an irradiation fiber 1402, and detection fibers 1403 with respect to the fixing substrate 1301. [ Fig. 15] Fig. Figure 15 is a diagram illustrating one cause of intercapillary crosstalk. [ Fig. 16] Fig. 16 is a diagram illustrating an effect of reducing intercapillary crosstalk by the fixing substrate 1301. Description of embodiments<Erste Ausführungsform>

[0020] Fig. Figure 1 is a configuration diagram of an electrophoresis apparatus 100 according to a first embodiment of the present disclosure. This embodiment illustrates an example of a case where fluorescence detection is adopted as a detection method. DNA is considered as a sample to be measured, but the sample is not limited thereto. The electrophoresis apparatus 100 includes an irradiation optical system 101 that generates fluorescence excitation light for fluorescence measurement, and an optical detection system 102 that detects fluorescence.In addition, the electrophoresis device 100 includes, as devices for performing electrophoresis, a high-voltage power supply 103, a polymer container 104 holding a polymer as a separation medium, a pump unit 105 for filling the polymer, a temperature adjusting device 106 for adjusting the temperature of the capillary, a buffer container 107 electrically connected to both ends of the capillary and holding a buffer that applies the voltage of the high-voltage power supply to the capillary, and an autosampler unit 108. These components are controlled by a control device 109. A capillary cartridge 110, which is a consumable, is connected to the electrophoresis device 100. The capillary cartridge includes capillaries 111 therein. The high-voltage power supply 103 applies a voltage to both ends of the capillaries 111 via the buffer.

[0021] The optical irradiation system 101 in the electrophoresis device 100 includes a body-side irradiation fiber 112 for guiding generated excitation light, and the optical detection system 102 includes body-side detection fibers 113 for guiding fluorescence. On the other hand, the capillary cartridge 110 includes a cartridge-side irradiation fiber 114 for guiding excitation light to the capillaries and cartridge-side detection fibers 115 for guiding fluorescence generated in the capillaries. The relative positions of the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fibers 115 are adjusted and fixed at a detection point 116. The body-side irradiation fiber 112 and the cartridge-side irradiation fiber 114 and the body-side detection fibers 113 and the cartridge-side detection fibers 115 are connected by a fiber connector 117.

[0022] The pump unit 105 includes a channel block 118, a syringe 119, a check valve 120, and a valve 121. The check valve 120 is installed so that the fluid flows only in the direction from the polymer container 104 to the channel block 118. The autosampler unit 108 includes a buffer tray 122, a cleaning water tray 123, a waste liquid tray 124, a sample tray 125, and a stage 126 that controls the positions of these trays. An electrode 127 is provided near the sample injection ends of the capillaries 111. The ends of the capillaries 111 and the high-voltage power supply 103 are electrically connected when the buffer tray 122 and the sample tray 125 are installed near the injection ends.

[0023] The following describes a mechanism in which fluorescence detection is performed by the electrophoresis device 100 of the present disclosure. The excitation light generated in the optical irradiation system 101 is introduced into the body-side irradiation fiber 112 and is introduced into the cartridge-side irradiation fiber 114 through the fiber connector 117. The excitation light reaches the detection point 116 through the cartridge-side irradiation fiber 114 and is applied to the inner diameters of the capillaries 111. The fluorescence generated at the excitation light irradiation points of the capillaries 111 is collected by the cartridge-side detection fibers 115 and delivered to the body-side detection fibers 113 through the fiber connector 117. Thereafter, the fluorescence is detected by the optical detection system 102 and converted into electrical signals. This signal is recorded by the control device 109.

[0024] The following describes a mechanism for analyzing a sample by electrophoresis in the electrophoresis device 100 of the present disclosure. As an example, the electrophoresis analysis is performed in the order of polymer injection into the capillaries, pre-electrophoresis, sample injection, and sample electrophoresis. The operating steps of the device in the electrophoresis analysis are shown in Fig. 2 illustrates.

[0025] When the measurement is started (S201), the capillaries 111 are first filled with the polymer by the pump unit 105. First, the waste liquid tray 124 is installed at the ends of the capillaries 111 (S202), and the valve 121 is closed (S203). When the valve 121 is closed, the syringe 119 is placed under negative pressure, and the polymer from the polymer container 104 is filled into the syringe 119 (S204). Note that the inner diameters of the capillaries 111 are several tens of µm and sufficiently smaller than the channel diameter of the channel block 118, and the buffer does not flow from the capillaries 111 to the syringe 119 due to the large resistance. Next, the syringe 119 is pressurized, and the capillaries 111 are filled with the polymer (S205). Thereafter, the valve 121 is opened and the ends of the capillaries 111 are electrically connected to the high voltage power supply 103 (S206).

[0026] After the capillaries are filled with the polymer, pre-electrophoresis is performed. The cleaning water tray 123 is moved to the sample introduction ends of the capillaries 111 to clean the tip portions (S207). Next, the buffer tray 122 is placed at the sample introduction ends of the capillaries 111 (S208), a high voltage is applied for approximately several minutes by the high-voltage power supply 103, and pre-electrophoresis is performed (S209). Impurity ions of the polymer filled in the capillaries are removed by pre-electrophoresis before sample injection.

[0027] After pre-electrophoresis, the sample is injected into the capillaries 111. First, the cleaning water tray 123 moves to the sample introduction ends of the capillaries 111 to clean the tip portions (S210). Then, the sample tray 125 is placed at the sample introduction ends of the capillaries 111 (S211), and the sample is electrically injected into the capillaries 111 by applying a short-term voltage of approximately several seconds to both ends of the capillaries 111 using a high-voltage power supply (S212). This step injects the sample only into small areas of the ends of the capillaries 111. Next, the cleaning water tray 123 moves back to the sample introduction ends of the capillaries 111 to clean the tip portions and removes the excess sample attached to the outer walls of the capillaries 111 (S213). Then, the buffer tray 122 is installed at the sample introduction ends of the capillaries 111 (S214).

[0028] After sample injection, the injected sample is separated by electrophoresis. A voltage is applied across the capillaries 111 by the high-voltage power supply 103, and the injected sample is migrated (S215). During electrophoresis, the temperature adjustment device 106 maintains the capillaries 111 at a constant temperature. A phosphor is applied to the sample, and fluorescence measurement is performed by the method described above when the sample passes the detection site 116. The movement speed of each component in the sample varies depending on the amount of charge and the molecular size, creating a difference in the time it takes to reach the detection site 116. Therefore, the time waveform of the fluorescence signal fluorescing at the detection site 116 provides information about the component of each sample.After the acquisition of the signal waveform is completed, the voltage application is stopped and the measurement is terminated (S216).

[0029] Fig. 3 and Fig. 4 illustrates details of a structural example of the detection site 116. As an example, the detection site 116 can be formed by fixing the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fiber 115 on the substrate having the V-shaped grooves. Fig. 3(a) illustrates a structure of a fixing substrate 301 for fixing the capillaries 111 and the cartridge-side irradiation fiber 114. The fixing substrate 301 includes capillary fixing grooves 302 and a fiber fixing groove 303.

[0030] Fig. 3(b) is an enlarged view of the vicinity of an intersection of the capillary fixing grooves 302 and the fiber fixing groove 303. The fiber fixing groove 303 has a lens fixing groove 304 immediately before an intersection with the capillary fixing grooves 302. A through-hole 305 is provided at an intersection portion between the fiber fixing groove 303 and the capillary fixing grooves 302. The through-hole 305 is provided so that the laser light emitted from the cartridge-side irradiation fiber 114 attached to the fiber fixing groove 303 is not blocked by the wall surface of the capillary fixing grooves 302. In this example, the through-hole 305 penetrates the substrate, but may be a recess that does not penetrate the substrate. The substrate having the shape of Fig. 3(a) can be formed, for example, by anisotropic etching of silicon.

[0031] Fig. 3(c) is a diagram in a case where the capillaries 111, the cartridge-side irradiation fiber 114, and a ball lens 306 are installed on the fixing substrate 301. The capillaries 111 are generally coated with a coating such as polyimide, and this coating interferes with the optical measurement around the through-hole 305 and is thus removed. The excitation light passed through the cartridge-side irradiation fiber 114 is collimated by the ball lens 306. The collimated excitation light passes through the four capillaries 111 and excites the phosphor within the capillaries 111. In the case of the optical configuration in the form of Fig. 3(c), it is known that the outer diameters, inner diameters, and spacing of the capillaries 111 can be adjusted so that the excitation light propagates sequentially through each capillary due to the lensing effect of the capillaries.

[0032] Generally, the outer diameters of the capillaries 111, the cartridge-side irradiation fiber 114, and the ball lens 306 differ from each other. By adjusting the depth of each of the V-shaped grooves for fixing each of the capillaries 111, the cartridge-side irradiation fiber 114, and the ball lens 306 so that the central axis of each element coincides with the substrate front surface, it is possible to perform axial alignment of each element. Here, the ball lens 306 is used to collimate the excitation light emitted from the cartridge-side irradiation fiber 114. Alternatively, collimation can be performed by another method, such as a GRIN lens or a lens fiber. When the number of capillaries is one, there is an option not to perform collimation.

[0033] Fig. 4 illustrates a method for further attaching the cartridge-side detection fibers 115 to the Fig. 3 illustrates the fixing substrate 301, to which the capillaries 111 and the cartridge-side irradiation fiber 114 are fixed. A holding substrate 401 ( Fig. 4(b)) having a structure similar to that of the fixing substrate 301 is attached to the fixing substrate 301 ( Fig. 4(a)) to which the respective components have been installed. Like the fixing substrate 301, the holding substrate 401 has the capillary fixing grooves 302 and the fiber fixing groove 303. On the other hand, the structure of the holding substrate 401 differs from that of the fixing substrate 301 in that the detection fiber array mounting hole 402 is provided. The fixing substrate 301 and the holding substrate 401 are fixed such that the grooves face each other ( Fig. 4(c)).

[0034] As an example, the capillary fixing grooves 302 and the fiber fixing groove 303 of the holding substrate 401 are grooves in which the central axes thereof are located on the substrate front side when the capillaries 111 and the cartridge-side irradiation fiber 114 are installed similarly to the structure of the fixing substrate 301. In this structure, the capillaries 111 and the cartridge-side irradiation fiber 114 are fixed by being sandwiched between the fixing substrate 301 and the holding substrate 401. In the example of Fig. 4, since the ball lens 306 is in contact only with the fixing substrate 301, it is necessary to fix it with an adhesive or the like. It is possible to adopt a structure in which the ball lens 306 is fixed to the cartridge-side irradiation fiber 114 in advance by bonding with a transparent adhesive, or the ball lens 306 is installed in a position sandwiched by the substrates by separating the distance between the ball lens 306 and the capillaries 111.

[0035] As an example, the cartridge-side detection fibers 115 are fixed to V-shaped grooves formed in another substrate, thereby forming a detection fiber array 403. The detection fiber array 403 is fixed in a state where it is inserted into the detection fiber array mounting hole 402 ( Fig. 4(d)). For example, an array fixing member 404 may be pressed and fixed to the support substrate 401 with an adhesive or the like, and the detection fiber array 403 may be fixed to the array fixing member 404 with an adhesive or the like.

[0036] The alignment of the fiber array 403 and the capillaries 111 can be performed by a variety of means. As an example, the size of the fiber array 403 and the size of the fiber array mounting hole 402 are adjusted to match each other, and when the fiber array 403 is fitted, the cartridge-side detection fibers 115 can be adjusted to be fixed so that they face the excitation light irradiation locations on the capillaries 111. Alternatively, the detection fiber array mounting holes 402 are holes corresponding to the number of capillaries formed at the position of the central axis of each capillary ( Fig. 4(e)), and the cartridge-side detection fibers 115 can be aligned by inserting them into these holes.

[0037] The alignment of the fiber array 403 can be performed using some observation means. For example, the capillaries 111 and the fiber array 403 can be observed through the through-hole 305 with a camera, and the fiber array 403 can be fixed in a state where the capillaries and the fiber array are aligned. Alternatively, water, an aqueous solution of a fluorescent dye, or the like is injected into the capillaries 111, and a Raman signal, a fluorescence signal, or the like output from the cartridge-side detection fibers 115 in a state of excitation light irradiation is monitored. The position of the fiber array 403 can be adjusted and fixed so that these signals are maximized.

[0038] In Fig. 3 and Fig. Figure 4 illustrates an example where the number of capillaries is four, but any number of capillaries can be used. If the number of capillaries is large, two cartridge-side detection fibers 115 can be provided to balance the power of the excitation light applied to each capillary, and the excitation light can be applied from both side surfaces of the capillary array.

[0039] When the number of capillaries is 2 or less, the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fibers 115 can be installed on the same substrate. Fig. 5(a) illustrates a structural example of a fixing substrate 501 in a case where the number of capillaries is two. In this example, the fixing substrate 501 includes capillary fixing grooves 502, an irradiation fiber fixing groove 503, a ball lens fixing hole 504, and detection fiber fixing grooves 505. These grooves are formed, for example, by dry etching a silicon substrate to form grooves with a quadrangular cross-section. When the grooves are quadrangular, the depths or widths of the grooves are adjusted so that the center axes of the capillaries 111, the cartridge-side irradiation fiber 114, the ball lens 306, and the cartridge-side detection fibers 115 are on the same plane.

[0040] Fig. 5(b) is a diagram in which the capillaries 111, the cartridge-side irradiation fiber 114, the ball lens 306, and the cartridge-side detection fibers 115 are installed on the fixing substrate 501. Each element is aligned by a groove and then fixed by an adhesive or the like. As in the example of Fig. 3, the coating of the capillaries 111 is removed at the detection positions.

[0041] The structure of the detection site 116 is not limited to the configuration described above, and other configurations can be adopted as long as the relative positions of the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fiber 115 can be fixed. Alignment through the structure and the groove on the substrate is not always necessary. For example, a method of adjusting the positions of the capillaries or fibers with a jig or the like on the substrate and fixing the capillaries or fibers with an adhesive can be used.

[0042] The cartridge-side irradiation fiber 114 and the cartridge-side detection fiber 115 are not necessarily arranged so that their central axes face the detection points on the capillaries 111. For the cartridge-side irradiation fiber 114 and the cartridge-side detection fiber 115 whose central axes do not face the detection points on the capillaries 111, light emitted from the cartridge-side irradiation fiber 114 can be guided to the detection points on the capillaries 111 by a reflecting mirror provided on the fixing substrate 501, and fluorescence emitted from the detection points on the capillaries 111 can be incident on the cartridge-side detection fiber 115.

[0043] The requirement regarding the optical fiber arrangement is similar even if an element other than the reflecting mirror is provided on the fixing substrate 501. Assume that a light beam is emitted from the optical fiber end face in the direction of the central axis of the optical fiber, and an optical element on the fixing substrate 501 exerts an optical effect such as reflection, refraction, or diffraction on the light beam. At this time, the trajectory of the light beam is defined as the optical axis of the optical fiber. To detect the fluorescence emitted by the substances in the capillaries, the optical axes of the cartridge-side irradiation fiber 114 and the cartridge-side detection fiber 115 may intersect in the lumen of the capillaries 111.It should be noted that the optical axes in the lumens of the capillaries 111 do not have to intersect exactly and an error is allowed if the optical axes lie within a range in which fluorescence is incident on the cartridge-side detection fibers 115.

[0044] Fig. Figure 6 is a diagram for describing the connection in a case where the capillary cartridge 110 is installed in the electrophoresis apparatus 100. The capillary assembly must be connected to the pump unit 105 to inject the polymer therein and electrically connect to the electrodes in the buffer tank 107. As an example, the capillaries 111 are bundled at a connecting portion and connected to the pump unit 105 using a fitting 601 or the like.

[0045] When an electrode 127 is included in the capillary cartridge 110, the high-voltage power supply 103 and the electrode 127 are connected by an electrical connector 602. The temperature adjustment device 106 is connected via a temperature adjustment connector 603. When temperature adjustment is performed by a heater or the like included in the capillary cartridge 110, the temperature adjustment connector 603 is an electrical connector. When temperature adjustment is performed by flowing a fluid such as air, the temperature adjustment connector 603 is a channel connection connector.

[0046] As described above, the body-side irradiation fiber 112 and the cartridge-side irradiation fiber 114, and the body-side detection fiber 113 and the cartridge-side detection fiber 115 are connected by the fiber connector 117. A commonly used SC connector, FC connector, LC connector, or the like can be used as the fiber connector 117. If the number of fibers to be connected is large, a multi-fiber connector such as an MPO connector can be used. Alternatively, a specially designed fiber connector can be used.

[0047] By adopting a structure in which the optical system of the electrophoresis device 100 and the capillary cartridge 110 are connected by connecting the body-side irradiation fiber 112 and the cartridge-side irradiation fiber 114, and connecting the body-side detection fiber 113 and the cartridge-side detection fiber 115, it is possible to achieve both ease of replacement of the capillary cartridge 110 and resistance to vibrations and external environmental changes. The user can attach and detach the fiber cartridge 110 with respect to the optical system only by attaching and detaching the fiber connector 117. Since the light transmission is performed by optical fibers, it is robust against the influence of vibrations and external environmental changes.

[0048] For example, if a capillary 10 cm in front of the light source is irradiated with excitation light with a positioning accuracy of ±10 µm by beam propagation in a free space, the angular variation of the beam must be only about ±0.01 or less. To achieve this accuracy and stability, it is necessary to form the support structure of the optical system with a rigid material that is not deformed by vibration or shock and has low thermal expansion. As a result, the device becomes large and heavy. On the other hand, if transmission is performed by optical fibers, the rigidity of a section from the light source to the fiber and a section from the fiber to the capillary can be kept high. Since the distance between these two components can be adjusted to about several mm or less, the tolerance for beam angle variation due to deformation of the support structure also increases.

[0049] The effects described above relate to a general optical fiber system. Specifically, in the structure of the present disclosure, the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fiber 115 are fixed to the substrate, and a connection point to the outside is separately provided, so that it is easy to achieve both easy attachment and detachment and fixing accuracy in the connection between the electrophoresis device 100 and the capillary cartridge 110. In the case of the manner in which the fiber array is attached to and detached from the capillary array as in PTL 2, the fixing structure of the fiber array must achieve both easy attachment and detachment and fixing accuracy, and there is a problem that the attachment and detachment mechanism is complicated and the cost is high.On the other hand, in the structure of the present disclosure, both the easy attachment to and detachment from the external connection and the fixing accuracy are ensured by the fiber connector, which is a component that has been widely used in general, and it is sufficient to meet the requirement of the fixing accuracy between the capillary and the fiber, so that it is possible to avoid complications and cost increase of the attachment and detachment mechanism.

[0050] Fig. 7 illustrates a method for adjusting the temperature of the capillaries 111 in the capillary cartridge 110 of the present disclosure. Fig. Figure 7(a) illustrates an example in which the capillaries 111 are temperature-adjusted by a heating element, such as a film heater 701. The capillaries 111 and the detection site 116 are arranged to be in contact with the film heater 701. The film heater 701 receives a power supply from the electrophoresis device 100 via the temperature-adjusting connector 603. The film heater 701 may be provided with a temperature sensor for feedback control.

[0051] Fig. 7(b) illustrates an example in which the temperature of the capillaries 111 is adjusted by supplying a fluid, such as temperature-adjusted air or inert liquid, to the cartridge. In this example, the temperature-adjusting connector 603 includes a fluid supply port 702 and a fluid discharge port 703. The temperature-adjusted fluid is supplied from the fluid supply port 702. A partition wall 704 is provided inside the cartridge, and the fluid flows without remaining inside, adjusting the temperature of the capillaries 111. The fluid then returns to the electrophoresis device 100 from the fluid discharge port 703.

[0052] In a case where a liquid is used as the temperature adjustment fluid, since the refractive index of the liquid is different from that of air, the liquid enters the detection point 116, so the optical adjustment state may change. In this case, a structure may be adopted in which the detection point 116 is sealed and the liquid does not enter the interior. Alternatively, a structure may be adopted in which the liquid enters the detection point 116, and an optical design may be made assuming that the measurement is performed in a state where the light passage point is filled with the liquid.

[0053] According to the configuration of the present disclosure, most of the capillaries, including the detection site 116, can be integrally temperature-controlled. When separating a sample by electrophoresis, the mobility of the sample changes depending on the temperature of the separation medium, and thus, it is desirable to minimize the temperature distribution of the capillaries and the temperature fluctuation over time to obtain a stable measurement result. It is desirable that the temperature of the capillaries be kept constant not only when the device is in a specific environment, but also when the air temperature around the device changes.

[0054] However, the conventional capillary electrophoresis device has a problem that the spatial distribution and temporal fluctuation of temperature are likely to occur at the detection point 116 compared to other locations on the capillaries. To perform fluorescence measurement at the detection point 116, it is necessary to irradiate the capillaries 111 with excitation light and guide the generated fluorescence to the detector. To realize fluorescence measurement by propagating light in a free space, it is necessary to provide an opening through which excitation light and fluorescence pass, and a temperature adjustment mechanism cannot be provided in this opening portion. In addition, since the capillary assembly must be attached to the optical measuring mechanism within the device, heat conduction occurs through the attachment portion, and the temperature changes.

[0055] For example, regarding the spatial distribution and temporal variation of temperature due to the influence of the opening, it is possible to take measures such as providing a transparent window with high thermal insulation properties in the opening or providing an individual temperature adjustment mechanism in the opening. However, installing a transparent window may cause deterioration in optical performance due to reflection by the window or the like, and there is still uneven heat conduction due to a difference between a material and structure of the window and a surrounding material and structure.One countermeasure to provide an individual temperature adjustment mechanism in the opening section is to perform heating and cooling by considering the heat conduction state near the opening, but the structure and control for maintaining the temperature uniformity with other sections can be complicated. Since there is no change in the fact that the opening section frequently exchanges heat with the outside compared to other sections, a problem remains that the capillary temperature is likely to change due to changes in the temperature of the external environment.

[0056] On the other hand, in the structure of the present disclosure, since the excitation light and fluorescence are exchanged via the optical fiber, the measurement window portion can be almost completely isolated from the outside. When the temperature of the entire capillaries 111 including the detection site 116 is controlled by the Fig. 7 or the like, it is possible to uniformly and integrally adjust the temperature of all locations except the sample injection ends of the capillaries and the connecting portions with the pump unit 105, which are structurally forced to contact the outside. Furthermore, it is not necessary to provide a special temperature adjustment mechanism, and stable temperature adjustment can be easily performed.

[0057] Furthermore, since the positions of the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fiber 115 are fixed within the detection site 116, the entire capillaries 111 including the detection site 116 do not need to be firmly fixed to the cartridge 110 or the electrophoresis device 100. Therefore, it is possible to install a heater or a heat-insulating material around the entire capillaries 111 including the detection site 116, which fixes the detection site 116 so that it does not contact the casing of the capillary cartridge 110 as much as possible to reduce heat transfer with the peripheral portion.

[0058] As a result, it is possible to control the entire capillaries 111 to a uniform temperature without providing an individual heat insulation mechanism or a temperature control mechanism in the measurement window section. It is also possible to reduce the heat transfer path with the outside and separate the sample more stably by electrophoresis, even when the temperature outside the device changes. <Erste Ausführungsform: Zusammenfassung>

[0059] The electrophoresis apparatus 100 according to the first embodiment includes the optical irradiation system 101, the body-side irradiation fiber 112, the optical detection system 102, and the body-side detection fibers 113. The capillary cartridge 110 includes the cartridge-side irradiation fiber 114 and the cartridge-side detection fibers 115. The relative positions of the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fibers 115 are adjusted and fixed at the detection site 116. The body-side irradiation fiber 112 and the cartridge-side irradiation fiber 114, and the body-side detection fibers 113 and the cartridge-side detection fibers 115 are connected by the fiber connector 117. With the above configuration, it is possible to achieve both robustness against vibrations and external environmental changes and ease of capillary replacement.In addition, by integrally adjusting the temperature of the entire capillaries 111 including the detection point 116, the temperature distribution of the capillaries can be made uniform, and the temperature fluctuation can be reduced. <Zweite Ausführungsform>

[0060] Fig. 8 is a configuration diagram of an electrophoresis apparatus 800 according to a second embodiment of the present disclosure. The components of the electrophoresis apparatus 800 according to the second embodiment are similar to those of the electrophoresis apparatus 100 according to the first embodiment. However, the second embodiment differs from the first embodiment in that consumables such as a polymer, a buffer, and a cleaning liquid, and a channel structure for flowing them, are integrated into the capillary cartridge 801. As in the first embodiment, the capillary cartridge includes capillaries 111, a cartridge-side irradiation fiber 114, cartridge-side detection fibers 115, and a detection site 116 therein. A sample injection-side channel 802 is provided at the sample injection ends of the capillaries 111. A solution tank 803 and a waste liquid tank 804 are connected to the sample-side injection channel 802.An electrode 127 is installed in the sample injection-side channel 802. The solution tank stores a buffer, a cleaning liquid, and the like. The opposite ends of the capillaries 111 are connected to a polymer injection channel 805. A polymer tank 104 and a buffer tank 107 are connected to the polymer injection channel 805. With this configuration, the maintenance work performed by the user is concentrated on replacing the capillary cartridge 801, and the time and effort for maintenance can be reduced compared to the configuration of the first embodiment.

[0061] In the second embodiment, a process similar to the process of the first embodiment ( Fig. 2) by flowing each liquid in the channel. The polymer injection channel 805 has a structure similar to that of the pump unit 105 and performs polymer injection into the capillaries 111 (S202 to S206). In the present embodiment, a drive unit 806, which drives a mechanism corresponding to the syringe 119, is provided on the electrophoresis device 800 side.

[0062] The cleaning (S207, S210, S213) of the capillaries and the electrical connection (S208, S214) between the capillary tips and the electrode by buffer injection are performed by supplying a cleaning liquid and a buffer from the solution tank to the sample injection-side channel 802. Each solution can be stored in a syringe, and the solution can be supplied by pressing the syringe, or a mechanism for supplying the solution can be included separately. As in the structure for injecting a polymer, a liquid supply unit 807 for supplying power for supplying liquid is provided on the electrophoresis device 800 side. The waste liquid is discarded to the waste liquid tank 804.

[0063] Sample injection (S212) is performed by externally injecting the sample into the sample injection-side channel 802. The sample is held in a sample cartridge 808, and the sample is injected into the sample injection-side channel 802 by a sample cartridge control unit 809. Note that the sample cartridge 808 may simply temporarily hold the sample introduced by the user and supply the sample to the sample injection-side channel 802 at the time of sample injection (S212), or may perform pretreatment such as cleaning the sample or mixing with a reagent in addition to supplying. In a case where the sample cartridge 808 also performs preprocessing, the sample cartridge control unit 809 generally controls the liquid supply, mixing, and the like of various necessary reagents.For example, PTL 3 discloses an apparatus that performs continuously from sample pretreatment to analysis by capillary electrophoresis and its structure.

[0064] In the electrophoresis device 800 according to the second embodiment, consumables are contained in the capillary cartridge 801. Therefore, it is not possible to individually replace each consumable, such as a capillary, a polymer, and a buffer, according to a consumption state. On the other hand, the simplicity of maintenance of the device, including the replacement of consumables, is emphasized. Such a configuration is particularly suitable for use by a user who is unfamiliar with the handling of the device. Therefore, it is expected that the replacement of the capillary cartridge 801 can be easily performed without requiring any special operation.

[0065] Fig. 9 illustrates an example of a structure in which the capillary cartridge 801 is installed in the electrophoresis apparatus 800 according to the second embodiment. Fig. 9(a) illustrates the electrophoresis device 800, the capillary cartridge 801, the sample cartridge 808, and the control device 109. In this structure, the capillary cartridge 801 is connected to be inserted into a capillary cartridge insertion section 901 provided in the electrophoresis device 800. The sample cartridge 808 is connected to be inserted into a sample cartridge insertion section 902 provided in the electrophoresis device 800. Although only the screen is illustrated, the control device 109 can be Fig. 9(a) may be a tablet PC, a notebook PC, a desktop PC, or the like, or may be integrally integrated into the electrophoresis apparatus 800.

[0066] Fig. Figure 9(b) illustrates an example of the structure of the capillary cartridge 801. The function and operation of each component are as described above. In this example, the temperature adjustment of the capillaries is performed by a heater 903. Fig. 9(b), the heater 903 is installed below the capillaries 111. Alternatively, the heater 903 may be arranged to sandwich the capillaries 111 to improve temperature setting accuracy, or a heat-insulating material may be installed on the upper surfaces of the capillaries 111.

[0067] In the example of Fig. 9(b), electrical and optical connections are made through a connector 904. An electrical connector and an optical connector are included in the connector 904. When the temperature of the capillaries is adjusted by circulating the fluid, a connector is provided for connecting the channel. The supply of external force from the drive unit 806 and the liquid supply unit 807 is performed, for example, by applying a force to a liquid supply mechanism 905 from above the cartridge through a mechanical mechanism provided in the electrophoresis device 800. As an example, the liquid supply mechanism 905 has a syringe-like structure and performs liquid supply by being moved up and down by the electrophoresis device 800. A valve 906 is also opened and closed by a mechanical force from the electrophoresis device 800.It should be noted that these mechanical forces are not limited to vertical movement, and rotational force or the like may be applied. It is not excluded that a component that generates a mechanical force, such as a motor or solenoid, is installed within the capillary cartridge 801.

[0068] With the Fig. With the structure illustrated in Figure 9, the user can easily attach and detach the sample cartridge 801 to and from the electrophoresis device 800. When the sample cartridge 801 is inserted deeply into the capillary cartridge insertion portion 901 of the electrophoresis device 800, the sample cartridge 801 and the electrophoresis device 800 are electrically and optically connected by the connector 904. A mechanism for applying mechanical force to the liquid supply mechanism 905, the valve 906, and the like is accessed from the upper portion of the sample cartridge 801. The mechanical mechanism is installed so that it does not interfere when the sample cartridge 801 is attached or detached, or is moved to a position where the mechanical mechanism does not interfere when the sample cartridge 801 is attached or detached.The sample cartridge 801 is a consumable item and must be replaced with a new one after a certain number of uses. Therefore, one advantage is that the sample cartridge 801 can be easily attached and detached. The optical connection using the optical fiber and the optical connector makes the optical unit robust against vibrations and changes in the external environment, as described above in 47.

[0069] When attached to the electrophoresis device 800, the sample cartridge 801 is mechanically fixed to the electrophoresis device 800 to prevent falling off. The fixing mechanism is provided in a housing portion of the sample cartridge 801. On the other hand, a commercially available optical connector generally includes a fixing mechanism for fixing to an adapter. The fixing mechanism of the sample cartridge 801 and the fixing mechanism of the optical connector can be fixed simultaneously when attached to the electrophoresis device 800. Alternatively, the fixing mechanism may not be included in the optical connector, and fixing may be performed only by the fixing mechanism of the sample cartridge 801.

[0070] As described above, the optical system of the electrophoresis device 800 of the present disclosure is robust against vibrations and external environmental changes by using the optical fiber and the optical connector. However, if a strong impact is applied to the device, the position of the optical component may change, and the component may be damaged.

[0071] As protection against a strong impact, there is a method of attaching a damping material to an object to be protected. In the configuration where the light beam propagates in free space, the relative positions of the excitation light irradiation optical system, the capillaries, and the optical detection system must be fixed. Therefore, for example, it is conceivable that the excitation light irradiation optical system, the capillaries, and the optical detection system are mounted on the same structural support, and the structural support described above is protected by the damping material. In this case, the protection target includes a light source, a detection system, a structural support, and the like, and has a corresponding weight. It is necessary to install an interference mechanism that supports this weight and has sufficient damping performance.

[0072] On the other hand, in the structure of the present disclosure, the optical irradiation system 101, the optical detection system 102, and the detection site 116 are connected by an optical fiber. Since the optical fiber has a flexible property, the optical irradiation system 101, the optical detection system 102, and the detection site 116 can be individually protected with the cushioning material. Specifically, the detection site 116 includes the capillaries 111 from which the coating has been removed, the cartridge-side irradiation fiber 114, and the cartridge-side detection fibers 115. The positional accuracy between these three components is important for measurement performance, and thus, protection against impact is particularly required.

[0073] As in Fig. As illustrated in FIG. 10, the detection site 116 can be fixed to a structural support 1001, such as the inner wall of the capillary cartridge 801, via a damping material 1002. The detection site 116 is lightweight and includes only lightweight elements such as the capillaries 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fibers 115. Therefore, sufficient damping performance can be achieved with a simple configuration in which a soft member such as rubber is simply used as the damping material 1002 and the structural support 1001, the damping material 1002, and the detection site 116 are bonded and fixed.

[0074] In the configuration disclosed in PTL 2, a detection fiber array is connected to a capillary array contained in a capillary cartridge. In this way, even if the connecting mechanism including the fiber is connected to the capillary within the cartridge, it is possible to protect the cartridge and the connecting mechanism from impact in an integrated state. However, the configuration of the present disclosure can reduce the weight of the detection site 116 compared to the above structure. This is advantageous for protection against vibration and impact. <Zweite Ausführungsform: Zusammenfassung>

[0075] The electrophoresis device 800 according to the second embodiment has components similar to those of the electrophoresis device 100 according to the first embodiment and performs similar operations, but differs in that consumables such as polymers and buffers are installed within the capillary cartridge 801. In addition to an optical connector, electrical and fluid connectors are installed in the capillary cartridge 801. When the capillary cartridge 801 is inserted into the electrophoresis device 800, the two are connected by these connectors. A mechanical force is supplied from the electrophoresis device 800. The detection site 116 is fixed to the structural support 1001 via the damping material 1002. <Dritte Ausführungsform>

[0076] In the first and second embodiments, the detection of the sample in the capillary is performed by fluorescence measurement. The configuration of the present disclosure also works in a method other than fluorescence detection. As an example, in a third embodiment, a case where a sample is detected by light absorption measurement will be described. The structure and operation of an electrophoresis device in the third embodiment are the same as those in the first and second embodiments and are therefore omitted. The third embodiment differs from the first and second embodiments in the method of photodetection, and therefore the structure of the detection site 116 differs.

[0077] Fig. Figure 11 illustrates details of the structure of the detection point 116 in the third embodiment. In light absorption measurement, it is necessary to irradiate the capillary 111 with light emitted from the cartridge-side irradiation fiber 114, cause the light to pass through the capillary 111, and then collect the light with the cartridge-side detection fiber 115. In the present embodiment, this is realized by a structure in which the cartridge-side irradiation fiber 114 and the cartridge-side detection fiber 115 face each other, with the capillary 111 interposed therebetween.

[0078] Fig. Figure 11(a) illustrates a structure of the fixing substrate 1101 for fixing the capillary 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fiber 115 to the substrate, and adopts the configuration described above. The fixing substrate 1101 is provided with a capillary fixing groove 1102, an irradiation fiber fixing groove 1103, and a detection fiber fixing groove 1104. Fig. 11 (b) is a diagram in which the capillary 111, the cartridge-side irradiation fiber 114, and the cartridge-side detection fiber 115 are fixed on the fixing substrate 1101.

[0079] It should be noted that in the present embodiment, no optical element other than the optical fiber is used. Alternatively, an optical element other than the optical fiber may be installed on the fixing substrate 1101 as needed. If a plurality of capillaries are provided, a plurality of Fig. 11 illustrated structures may be arranged in parallel or a structure similar to the Fig. 3 and Fig. 4 and the irradiation fiber array and the detection fiber array may be arranged to sandwich the capillary array.

[0080] It is also possible to achieve both the light absorption measurement configuration and the fluorescence measurement configuration. For example, as in Fig. 12, an absorbance measurement irradiation fiber 1201 and an absorbance measurement detection fiber 1202 may be installed for the capillary 111, and a fluorescence measurement irradiation fiber 1203 and a fluorescence measurement detection fiber 1204 may be arranged at an angle of 45 degrees with respect to the absorbance measurement irradiation fiber 1201 and the absorbance measurement detection fiber 1202. <Dritte Ausführungsform: Zusammenfassung>

[0081] In the detection site 116 according to the third embodiment, the cartridge-side irradiation fiber 114 and the cartridge-side detection fiber 115 are fixed to face each other via the capillary 111. The light emitted from the cartridge irradiation fiber 114 passes through the capillary 111 and then is incident on the cartridge-side detection fiber 115, thereby performing absorption measurement. <Vierte Ausführungsform>

[0082] In a fourth embodiment, with respect to a mounting substrate for mounting capillaries, an irradiation fiber, and a detection fiber, a method of forming a through hole by forming grooves on both a substrate front surface and a substrate back surface and a structure of a substrate manufactured by this method are described.

[0083] Fig. 13 illustrates a structure of a fixing substrate 1301 according to a fourth embodiment. Fig. 13(a) illustrates the front surface of the fixing substrate 1301 and Fig. 13(b) illustrates the back surface of the fixing substrate 1301. Similar to the fixing substrate 301 according to the first embodiment, the fixing substrate 1301 fixes four capillaries, one irradiation fiber, and four detection fibers. Capillary fixing grooves 1302 for positioning four capillaries and an irradiation fiber fixing groove 1303 for positioning one irradiation fiber are provided on the front surface of the fixing substrate 1301. A through-hole forming groove 1304 formed in a direction orthogonal to the capillary fixing groove 1302 is provided on the back surface. Through-holes 1305 are formed at an intersection of the capillary fixing grooves 1302 and the through-hole forming groove 1304. The four detection fibers are positioned by inserting them into the through-holes 1305.

[0084] A method for positioning the capillaries, the irradiation fiber and the detection fibers is similar to the method for positioning the capillaries, the irradiation fiber and the detection fibers on the substrate described in Fig. 4(e) of the first embodiment. However, the Fig. 4(e) is different in that the capillary fixing grooves are formed and then the through-holes are formed by another means, whereas the substrate of the present embodiment is different in that the fixing grooves and the through-holes are formed simultaneously by forming the grooves on both the front surface and the back surface.

[0085] A method for forming the through-hole will be described in detail below. The capillary fixing grooves 1302 and the through-hole forming groove 1304 are each formed to a depth that does not penetrate the substrate (groove depth < substrate thickness). On the other hand, the sum of the depth of the capillary fixing groove 1302 and the depth of the through-hole forming groove 1304 is set to be greater than the thickness of the substrate. At this time, the bottoms of the capillary fixing grooves 1302 and the through-hole forming groove 1304 overlap, and the overlapped portions become the through-holes 1305 connected to the grooves and penetrating the substrate. By adopting this method, groove formation and through-hole formation can be performed simultaneously at positions along the grooves.

[0086] In the manufacture of the mounting substrate 1301, as an example, the substrate material may be silicon, and grooves with a V-shaped cross section may be formed by anisotropic etching of silicon. First, thermal oxide films are formed on the front and back surfaces of a 100-level silicon substrate. After that, a resist is applied, exposed, and developed, and then etched with hydrofluoric acid to remove the oxide film at the portion where the V-grooves are formed. That is, the oxide film in the portions corresponding to the capillary fixing grooves 1302, the irradiation fiber fixing groove 1303, and the through-hole forming groove 1304 is removed. After that, the substrate is anisotropically etched with an alkali solution, such as an aqueous potassium hydroxide solution, to form V-grooves. The capillary fixing grooves 1302, the irradiation fiber fixing groove 1303 and the through-hole forming groove 1304 are formed by etching.As etching progresses and the bottoms of capillary fixing grooves 1302 and the bottom of through-hole formation groove 1304 intersect, through-holes 1305 are formed. The sizes of the through-holes can be adjusted by adjusting the width of through-hole formation groove 1304 and the etching time. After the V-shaped grooves are formed, the remaining oxide film is removed. The material of the mask used in anisotropic etching, the solution used in etching, and the like may differ from those described above.

[0087] In the case of a configuration where laser light is emitted from the side surface of the capillary array, where the capillaries are arranged in a row to excite the phosphor in the capillary, the distances from the substrate front surface of the irradiation fiber and the capillaries must be aligned with an accuracy of about 10 µm or less. In the case of anisotropic etching of silicon, the widths and angles of the V-grooves can be controlled with high precision, and the heights of the capillaries and the irradiated fiber from the substrate front surface can be aligned with high precision.

[0088] It should be noted that the substrate material of the fourth embodiment is not necessarily silicon, and the method for forming the fixed grooves is not necessarily anisotropic etching. It is sufficient that grooves for fixing the capillaries, the irradiation fiber, and the detection fiber can be formed with sufficient accuracy, and it is sufficient that grooves with a depth sufficient to form through holes in the substrate can be formed by etching from the front surface and the back surface.

[0089] Fig. 14(a) is a structural diagram when capillaries 1401, an irradiation fiber 1402, and detection fibers 1403 are attached to the fixing substrate 1301. Fig. 14(b) is a cross-sectional view of the structure of Fig. 14(a) along a plane perpendicular to the capillary fixing grooves 1302 at the position of the through holes.

[0090] The capillary 1401 is fixed by the capillary fixing groove 1302 such that the center axis of the capillary 1401 is located at a position a certain distance from the front surface of the fixing substrate 1301. For example, the capillary 1401 can be fixed with an adhesive or the like in a state where it is pressed against the capillary fixing groove 1302 by a capillary holding substrate (not shown).

[0091] In Fig. In FIG. 14(a), the irradiation fiber 1402 is fixed to the fixing substrate 1301 in a state where it is inserted into a position adjustment component 1404. The position adjustment component 1404 is a cylindrical component, and a hole into which the irradiation fiber 1402 is inserted is provided in the center. A portion into which a lens for collimating the excitation light emitted from the irradiation fiber 1402 is inserted is provided at the end on the capillary side of the position adjustment component 1404. Adjustment of the optical axis of the irradiation fiber 1402 and the lens is performed by inserting a lens into this portion. As an example, a conical hole may be provided at the end of the position adjustment component 1404, and the end of the ball lens may be fixed with an adhesive or the like by fitting the ball lens into the conical hole.The distance between the irradiation fiber 1402 and the lens can be adjusted while confirming a spot shape of light formed by the lens while light is emitted from the irradiation fiber 1402. When the number of capillaries 1401 is one, the lens is not necessarily used. The lens is not necessarily fixed by the position adjustment member 1404, and a method of providing a recess for attaching the lens to the fixing substrate 1301 can be adopted.

[0092] The detection fibers 1403 are inserted into the through holes 1305 and fixed thereto. At this time, the excitation light 1405 and the detection fibers 1403 are arranged orthogonally to each other, as shown in Fig. 14(b) illustrates this.

[0093] The fixing substrate of the fourth embodiment also has the effect of reducing crosstalk between capillaries when the number of capillaries is multiple. When the number of capillaries is multiple, fluorescence emitted from a particular capillary may enter an optical fiber for detecting different capillaries. In such a case, fluorescence from a particular capillary is mistakenly detected as light emitted from another capillary (crosstalk). When crosstalk exists, fluorescence caused by component a of sample A analyzed in a particular capillary is mistakenly identified as a signal from a capillary analyzing a different sample B, which may lead to an erroneous analysis result that component a is contained in sample B.

[0094] The crosstalk between the capillaries can be seen, for example, in the Fig. 15(a) illustrated way. In Fig. 15(a), a fluorescent light beam (arrow) generated in the left capillary is reflected by the front surface of the right capillary and enters the detection fiber, which detects a fluorescent light beam from the right capillary. The occurrence of crosstalk in the above path is suppressed by the structure of the fixing substrate of the fourth embodiment. In the fixing substrate of the fourth embodiment, a region other than the portion where the groove for fixing the capillary is formed is a wall separating the capillaries. As shown in Fig. 15(b), the Fig. 15(a) is blocked by this wall.

[0095] Fig. Figure 16 is a simulation result illustrating the crosstalk suppression effect of the fixing substrate. In this simulation, four capillaries with an inner diameter of 50 µm and an outer diameter of 343 µm are arranged at a pitch of 1 mm, and fluorescence is detected through optical fibers with a core diameter of 200 µm and an NA of 0.5. In this simulation, a region with a length of 50 µm in an inner diameter section of one of the four capillaries is caused to emit light, and the ratio of fluorescence incident on the detection fibers for the other capillaries, i.e., the crosstalk ratio, is calculated.

[0096] Fig. Figure 16(a) illustrates a crosstalk value in a case where there is no fixing substrate. The horizontal axis of the graph represents a light-emitting fiber, and each bar graph represents crosstalk observed in a fiber other than the fiber detecting the light-emitting capillary. When there is no fixing substrate, crosstalk of approximately 0.08% is observed in an optical fiber detecting a capillary adjacent to a light-emitting capillary. On the other hand, Fig. 16(b) Crosstalk in a case where a fixing substrate is present. In a case where the fixing substrate is present, it can be seen that the observed crosstalk is about 0.002%, and the crosstalk ratio is reduced to about 1 / 40. <Vierte Ausführungsform: Zusammenfassung>

[0097] The fixing substrate 1301 according to the fourth embodiment has the capillary fixing grooves 1302 and the irradiation fiber fixing groove 1303 on the substrate front surface, and has the through-hole forming groove 1304 on the back surface. The through-holes 1305 are formed at an intersection of the capillary fixing grooves 1302 and the through-hole forming groove 1304. A region where the capillary fixing grooves 1302 are not formed serves as a wall separating adjacent capillaries and reduces crosstalk between the capillaries. <modifikationen>

[0098] The present disclosure is not limited to the embodiments described above and includes various modifications. For example, the embodiments described above have been described in detail to facilitate understanding of the present disclosure and are not necessarily limited to those having all of the described configurations. Furthermore, a part of the configuration of one embodiment may be replaced with the configuration of another embodiment, and further, the configuration of one embodiment may be added to the configuration of another embodiment. In addition, it is possible to add, delete, and replace other configurations for a part of the configuration of each embodiment. List of reference symbols 100 electrophoresis device 101 optical irradiation system 102 optical detection system 103 High-voltage power supply 104 polymer containers 105 Pump unit 106 Temperature adjustment device 107 buffer tanks 108 Autosampler unit 109 Control device 110 Capillary cartridge 111 Capillary 112 body-side irradiation fibers 113 body-side detection fiber 114 cartridge-side irradiation fiber 115 cartridge-side detection fiber 116 Detection point 117 fiber connectors 118 Canal Block 119 syringe 120 check valve 121 Valve 122 buffer shell 123 Cleaning water bowl 124 Waste liquid tray 125 sample tray 126 level 127 Electrode 301 Fixing substrate 302 Capillary fixing groove 303 Fiber fixing groove 304 lens fixing groove 305 through hole 306 spherical lens 401 Holding substrate 402 detection fiber array mounting hole 403 Detection fiber array 404 Array Fastener 501 Fixing substrate 502 Capillary fixing groove 503 Irradiation fiber fixing groove 504 Ball lens fixing hole 505 Detection fiber fixing groove 601 Fitting 602 electrical connector 603 Temperature adjustment connector 701 foil heater 702 Fluid supply connection 703 Fluid discharge connection 704 Partition wall 800 Electrophoresis device 801 Capillary Cartridge 802 sample injection side channel 803 Solution tank 804 Waste liquid tank 805 Polymer injection channel 806 drive unit 807 Fluid supply unit 808 Sample Cartridge 809 Sample Cartridge Control Unit 901 Capillary cartridge insertion section 902 Sample cartridge insertion section 903 Heating device 904 connectors 905 Fluid supply mechanism 906 valve 1001 structural supports 1002 Damping material 1101 Fixing substrate 1102 Capillary fixing groove 1103 Irradiation fiber fixing groove 1104 Detection fiber fixing groove 1201 Absorption measurement irradiation fiber 1202 absorption measurement detection fiber 1203 Fluorescence measurement irradiation fiber 1204 Fluorescence measurement detection fiber 1301 Fixing substrate 1302 Capillary fixing groove 1303 Irradiation fiber fixing groove 1304 Through-hole forming groove 1305 through hole 1401 capillary 1402 irradiation fiber 1403 Detection fiber 1404 Position adjustment component 1405 Excitation light QUOTES CONTAINED IN THE DESCRIPTION

[0000] This list of documents submitted by the applicant was generated automatically and is included solely for the convenience of the reader. This list is not part of the German patent or utility model application. The DPMA assumes no liability for any errors or omissions. Cited patent literature

[0000] JP 2004-532384 A

[0006] US 2021 / 0003530 A1

[0006] Cited non-patent literature

[0000] H. Zhai et al., "A simple and compact fluorescence detection system for capillary electrophoresis and its application to food analysis", Electrophoresis, 36, 2509 (2015

[0007] < / modifikationen>

Claims

[1] A capillary electrophoresis device comprising: a light source; a first irradiation fiber configured to guide light from the light source; a detector configured to detect light; and a first detection fiber configured to guide light to the detector, wherein a capillary cartridge including a capillary, a second irradiation fiber, and a second detection fiber is attached to the electrophoresis device by connecting the first irradiation fiber and the second irradiation fiber and connecting the first detection fiber and the second detection fiber, and in the capillary cartridge, the second irradiation fiber and the second detection fiber are fixed such that optical axes of the second irradiation fiber and the second detection fiber intersect each other in an inner cavity of the capillary. [2] The capillary electrophoresis apparatus according to claim 1, wherein a sample detection method is fluorescence measurement. [3] The capillary electrophoresis apparatus according to claim 1, wherein a sample detection method is light absorption measurement. [4] Capillary electrophoresis device according to claim 2, wherein the capillary cartridge contains a large number of capillaries, Excitation light is incident through the second irradiation fiber onto the capillaries arranged in a line on a substrate, the light being incident on a side surface of an array configured by the capillaries arranged in a line, and Fluorescence from each of the capillaries collected by a plurality of the second detection fibers is passed through the first detection fiber to a detector. [5] A capillary electrophoresis apparatus according to claim 2 or 4, wherein in the capillary cartridge, a substrate to which the capillary and the second irradiation fiber are fixed and a substrate to which the second detection fiber is fixed are fixed so as to be substantially perpendicular to each other. [6] Capillary electrophoresis device according to claim 2 or 4, wherein the capillary, the second irradiation fiber and the second detection fiber are attached to the same substrate, so that the second detection fiber is substantially perpendicular to a plane formed by the capillary and the second irradiation fiber, and the alignment of the capillary, the second irradiation fiber and the second detection fiber is achieved by grooves and a through hole formed on the substrate. [7] Capillary electrophoresis device according to claim 2 or 4, wherein in the capillary cartridge, the capillary, the second irradiation fiber and the second detection fiber are mounted on a substrate so that they are all in the same plane, and the adjustment of the positions between the capillary, the second irradiation fiber and the second detection fiber is carried out by grooves formed on the substrate. [8] The capillary electrophoresis apparatus according to claim 1, wherein the capillary is temperature-adjusted integrally with the second irradiation fiber, the second detection fiber, and a fixing member for fixing the second irradiation fiber, the second detection fiber, and the capillary. [9] A capillary electrophoresis device according to claim 1, wherein the capillary is installed in a housing and is connected to an electrophoresis device via an optical connector installed on an outer wall of the housing. [10] The capillary electrophoresis apparatus according to claim 9, wherein a portion fixed such that the optical axes of the second irradiation fiber and the second detection fiber intersect each other in the lumen of the capillary is fixed to the casing via a buffer structure. [11] Capillary cartridge comprising a capillary, a first irradiation fiber and a first detection fiber, wherein the first irradiation fiber and the first detection fiber are fixed such that optical axes of the first irradiation fiber and the first detection fiber intersect each other in a lumen of the capillary, and the capillary cartridge is attached to an electrophoresis device that includes a light source, a second irradiation fiber configured to guide light from the light source, a detector configured to detect light, and a second detection fiber configured to guide light to the detector by connecting the first irradiation fiber and the second irradiation fiber and connecting the first detection fiber and the second detection fiber. [12] A capillary cartridge, wherein, when the capillary cartridge is installed in the electrophoresis device, the connection of the first irradiation fiber and the second irradiation fiber and the connection of the first detection fiber and the second detection fiber are achieved by inserting the capillary cartridge into a location for insertion into the electrophoresis device. [13] The capillary electrophoresis device according to claim 6, wherein the substrate has on a front surface a groove for fixing the capillary and a groove for fixing the second irradiation fiber, and has on a rear surface a through-hole forming groove orthogonal to the groove for fixing the capillary, and the second detection fiber is installed in a through-hole formed at an intersection of the groove for fixing the capillary and the through-hole forming groove, [14] The capillary electrophoresis device according to claim 6, wherein the substrate has a structure for blocking light between the through-hole. [15] The capillary cartridge according to claim 12, wherein a capillary, the second irradiation fiber, and the second detection fiber are fixed to a substrate; the substrate has, on a front surface, a groove for fixing the capillary and a groove for fixing the second irradiation fiber, and has, on a rear surface, a through-hole forming groove orthogonal to the groove for fixing the capillary, and the second detection fiber is installed in a through-hole formed at an intersection of the groove for fixing the capillary and the through-hole forming groove. [16] A mounting substrate manufacturing method for manufacturing a mounting substrate in a capillary cartridge connected to an electrophoresis device including a light source, the first irradiation fiber configured to guide light from the light source, a detector configured to detect light, and the first detection fiber configured to guide light to the detector 74, the mounting substrate for mounting a capillary, a second irradiation fiber connected to a first irradiation fiber, and a second detection fiber connected to a first detection fiber, the method comprising forming, on a front surface of the mounting substrate, a groove for mounting the capillary and a groove for mounting the second irradiation fiber, forming, on a rear surface, a groove orthogonal to the groove for mounting the capillary,and forming a through-hole for attaching the second detection fiber by crossing the groove for attaching the capillary and the orthogonal groove to each other. [17] The mounting substrate manufacturing method according to claim 16, wherein a material of the mounting substrate is silicon, and the groove for mounting the capillary, the groove for forming the irradiation fiber, and the groove orthogonal to the groove for mounting the capillary are formed by anisotropic etching.

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