JP2006047150A - Cataphoretic device - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a cataphoretic device for separating a plurality of samples by simultaneous cataphoresis, measuring the absorbance or fluorescent intensity of each sample simultaneously, and saving space and reducing the number of components. <P>SOLUTION: A microchip 10 for cataphoresis comprises a first quartz glass substrate 1 and a second one 7 that are bonded each other. An optical waveguide 3 and a plurality of cataphoresis grooves 2, vertically crossing the optical waveguide 3, are formed on the first quartz glass substrate. When guiding light from a light source for excitation to the cataphoresis grooves 2 through the optical waveguide 3, fluorescence generated from the inside of the cataphoresis grooves 2 is detected by a photodiode 14 on a platform substrate 11. <P>COPYRIGHT: (C)2006,JPO&NCIPI

Description

  The present invention relates to an electrophoresis apparatus for separating and analyzing proteins and DNAs, which includes a quartz microchip and an optical detector on a platform substrate, in which two quartz glass substrates are bonded and an analysis channel is formed inside. Is.

  In recent years, regarding electrophoresis devices, quartz microchips for electrophoresis formed by joining two quartz glass plates have been proposed and commercialized as those that can be expected to speed up analysis and downsize the device ( For example, see Patent Document 1.) FIG. 8 shows an example of a conventional quartz microchip for electrophoresis.

  Two quartz glass substrates 51 and 52 are prepared, and an electrophoresis groove 53 is formed in one quartz glass substrate 51. On the other quartz glass substrate 52, an anode side electrode tank 54 including a through hole for injecting an electrophoretic liquid and inserting a high voltage application electrode at positions corresponding to both ends of the electrophoresis groove 53, a cathode side electrode Each tank 55 is formed. By joining both the quartz glass substrates 51 and 52, a quartz microchip 60 for electrophoresis (see FIG. 9) is obtained. Measurement is performed by attaching this to the electrophoresis apparatus.

  The electrophoresis apparatus shown in FIG. 9 includes the electrophoresis quartz microchip 60, the ultraviolet excitation light source 61, the photodetector 62, the anode side electrode 63, and the cathode side electrode 64 obtained in FIG.

  There are mainly two measurement methods. One of them is the following method. First, the electrophoresis groove 53 is filled with the electrophoresis solution and sample molecules. Next, a high voltage is applied to the anode-side electrode 63 and the cathode-side electrode 64 inserted into both ends of the electrophoresis groove 53 via the anode-side electrode tank 54 and the cathode-side electrode tank 55, respectively. Thereby, the sample molecules are electrophoresed in the electrophoresis groove 53. The ultraviolet light is transmitted from the ultraviolet excitation light source 61 to the sample molecules migrating in the electrophoresis groove 53, and the ultraviolet absorbance is detected by the photodetector 62.

  Another measurement method is as follows. First, the electrophoresis groove 53 is filled with the electrophoresis solution and sample molecules that are fluorescently labeled. Next, a high voltage is applied to the anode side electrode 63 and the cathode side electrode 64 inserted at both ends of the electrophoresis groove 53. Thereby, the sample molecules are electrophoresed in the electrophoresis groove 53. Excitation light is irradiated from the ultraviolet excitation light source 61 to the fluorescently labeled sample molecules that migrate in the electrophoresis groove 53, and the fluorescence intensity is detected by the photodetector 62.

Regardless of which method is used, the absorbance or fluorescence intensity is measured by the photodetector 62 to analyze proteins and DNA.
JP 2000-310613 A

  However, the electrophoretic apparatus using such a microchip is greatly improved in handleability as compared with the conventional capillary electrophoretic apparatus. However, since only one electrophoretic groove is provided, a plurality of electrophoretic apparatuses are provided. In order to perform the separation analysis of the specimens, it is necessary to separate and analyze the specimens one by one in sequence, which increases the processing time as a whole and increases the analysis cost.

  On the other hand, in order to process a plurality of specimens in parallel, it is necessary to prepare a plurality of electrophoresis apparatuses. This increases the cost of the apparatus and makes the preparation and the like troublesome.

  Therefore, in order to reduce both analysis cost and time in an electrophoresis apparatus, it is important to analyze many samples simultaneously, that is, parallel processing of detection is an important issue, and downsizing the apparatus body Therefore, the reduction of the number of components such as the quartz microchip for electrophoresis and the light source for ultraviolet excitation for realizing the price reduction is a big issue.

  The present invention has been made on the basis of the above-mentioned problems. It is possible to simultaneously separate a plurality of samples by electrophoresis, perform absorbance measurement or fluorescence intensity measurement on each sample, and perform a plurality of electrical measurements. An object of the present invention is to provide an electrophoresis apparatus capable of saving space and reducing the number of parts compared to preparing an electrophoresis apparatus.

  In order to achieve the above-described object, the present invention provides a plurality of analysis channels through which a fluid containing a sample to be analyzed can flow, an optical waveguide provided so as to intersect all of the plurality of analysis channels, An excitation light source for supplying light to the optical waveguide, and fluorescence emitted in the plurality of analysis channels when the light from the excitation light source is guided into the plurality of analysis channels through the optical waveguide And an electrophoretic device comprising a plurality of photodetectors for detecting the above.

  According to the present invention, by simultaneously exciting a plurality of samples through one optical waveguide intersecting a plurality of electrophoresis grooves in optical measurement, simultaneous and parallel processing (inspection) of a plurality of specimens becomes possible. Analysis costs and time can be reduced.

  Further, since one excitation light source and one apparatus can simultaneously process a plurality of specimens, it is possible to reduce the apparatus space and reduce the apparatus cost.

  Hereinafter, the electrophoresis apparatus according to the present invention will be described.

[Configuration of electrophoresis apparatus]
FIG. 1 is a configuration diagram of an embodiment of an electrophoresis apparatus according to the present invention. The quartz microchip 10 for electrophoresis is composed of a first quartz glass substrate 1 and a second quartz glass substrate 7. A plurality of electrophoresis grooves 2 having a size of 50 μm in width, 50 μm in depth, and 5 cm in length are formed on the surface of the first quartz glass substrate 1. The optical waveguide 3 is formed so as to be orthogonal. An optical fiber 4 is connected to the input end face of the optical waveguide 3. A plurality of recesses are provided on the back surface of the first quartz glass substrate 1.

  A part of the plurality of recesses constitutes a plurality of photodiode recesses 5 serving as spaces for accommodating photodiodes 14 described later, and the remaining recesses are positioning protrusions provided on a platform substrate 11 described later. A plurality of positioning recesses 6 used for positioning the electrophoresis quartz microchip 10 are configured by fitting with the portion 13. Therefore, the photodiode recess 5 is provided corresponding to the intersection of the plurality of electrophoresis grooves 2 and the optical waveguide 3 provided in the first quartz glass substrate 1. The positioning recess 6 is provided at a position that does not hinder fluorescence analysis.

  Here, the interval between the plurality of electrophoresis grooves 2 is preferably 60 μm to 120 μm in order to suppress loss of light passing through the electrophoresis grooves 2. If the interval is longer than this, the loss increases and the sample in many grooves cannot be excited. Further, although it is possible to couple an excitation light source directly to the input end face of the optical waveguide 3, it is necessary to finely adjust the position of the electrophoresis quartz microchip 10 in the apparatus. It is preferable that an optical fiber 4 for guiding light to the electrophoresis quartz microchip 10 is connected to the electrophoresis quartz microchip 10 in advance.

  In the present embodiment, the optical fiber 4 is connected to only one of the two end faces of the optical waveguide 3, but the present invention is not limited to this, and the light is applied to both of the two end faces. A fiber may be connected, or an optical fiber may not be connected.

  In the second quartz glass substrate 7, injection of electrophoresis solution and insertion of high voltage application electrodes are performed at positions corresponding to both ends of the plurality of electrophoresis grooves 2 provided in the first quartz glass substrate 1. An anode side electrode tank 8 and a cathode side electrode tank 9 made of through holes are formed. The first quartz glass substrate 1 and the second quartz glass substrate 7 are bonded to form a quartz microchip 10 for electrophoresis.

  The electrophoresis quartz microchip 10 obtained as described above is mounted on the platform substrate 11. The platform substrate 11 is provided with a chip mounting recess 12 on the upper surface for mounting the electrophoresis quartz microchip 10. The upper surface of the chip mounting recess 12 is fitted with the positioning recess 6 in a position corresponding to the positioning recess 6 provided on the electrophoresis quartz microchip 10 to be mounted. A plurality of positioning convex portions 13 for fixing the microchip 10 are provided. A plurality of photodiodes 14 are further mounted on the upper surface of the chip mounting recess 12 at positions corresponding to the intersections of the plurality of electrophoresis grooves 2 and the optical waveguide 3 of the electrophoresis microchip 10. At this time, the number of electrophoresis grooves 2 provided in the electrophoresis quartz microchip 10 is the same as the number of photodiodes 14 mounted on the platform substrate 11.

  On the platform substrate 11 on which the plurality of photodiodes 14 are mounted, the electrophoresis quartz microchip 10 in which the optical waveguide 3 and the electrophoresis groove 2 are formed with the platform-side convex portion 13 as an alignment mark is hybrid-mounted, and the anode The electrophoresis apparatus according to this embodiment is configured by inserting electrodes into the side electrode tank 8 and the cathode side electrode tank 9, respectively.

  Then, a separate sample is put in each of the plurality of electrophoresis grooves 2, the electrophoresis is performed, and the fluorescence generated by the irradiation of the common excitation light through the optical waveguide 3 is converted into a photodiode corresponding to each electrophoresis groove 2. By detecting 14, a plurality of electrophoretic measurements can be performed at the same time, and protein and DNA can be analyzed by electrophoresis similar to the conventional one while saving space and cost.

  FIG. 2 is a side sectional view of the configuration diagram of the electrophoresis apparatus in the present embodiment. In FIG. 2, the electrophoresis groove 2, the anode-side electrode tank 8, and the photodiode 14 are illustrated as having three each.

[Preparation of quartz microchip for electrophoresis]
Next, FIGS. 3 to 6 show process diagrams for producing the electrophoresis quartz microchip 10 according to the present embodiment. 3 to 6, the description will be made assuming that the electrophoresis quartz microchip is provided with three electrophoresis grooves.

  3 to 6, each process is described using two cross-sectional views. 3, 4, and 6, one of the two cross-sectional views is a vertical cross-sectional view (the left-side cross-sectional view in each step) along the long axis of the optical waveguide, and B in the right-side cross-sectional view. This corresponds to a cross-sectional view taken along the plane B- '. The other is a vertical cross-sectional view (a cross-sectional view on the right side in each step) along the electrophoresis groove, which corresponds to a cross-sectional view taken along the line A-A ′ of the left-side cross-sectional view.

  In FIG. 5, one of the two cross-sectional views is a vertical cross-sectional view (cross-sectional view on the left side in each step) passing through all of the plurality of anode-side electrode tanks, and CC ′ in the right-side cross-sectional view. This corresponds to a cross-sectional view of the cut surface. The other is a vertical cross-sectional view (a cross-sectional view on the right side in each step) passing through a pair of the anode-side electrode tank and the cathode-side electrode tank, and corresponds to a cross-sectional view taken along the A-A ′ section of the left-side cross-sectional view.

  First, as shown in FIG. 3A, a quartz glass substrate 20 having an outer diameter of 4 inches and a thickness of 500 μm is prepared. Then, as shown in FIG. 3B, photolithography and reactive ion etching are performed on the back surface of the quartz glass substrate 20 to form a plurality of photodiode recesses 5 and a plurality of positioning recesses 6.

Next, as shown in FIG. 3C, a SiO ₂ —TiO ₂ glass or a SiO ₂ —GeO ₂ glass was formed on the surface of the quartz glass substrate 20 by an electron beam evaporation method, a plasma CVD method, or a sputtering method. Then, in order to make this glass film have a desired refractive index, the core glass film 21 is formed by performing heat treatment within a range of 1000 ° C. to 1200 ° C. At that time, the refractive index value of the obtained core glass film 21 needs to be smaller than the refractive index value of the quartz glass substrate 20.

  Next, as shown in FIG. 3D, the core glass film 21 is processed by photolithography and reactive ion etching so as to form an optical waveguide 23 serving as a core having a desired pattern.

Next, as shown in FIG. 3 (e), a SiO ₂ clad glass is formed on the optical waveguide 3 by using a plasma CVD method, and then the SiO ₂ glass film has a desired refractive index 1100. The clad layer 22 is formed by performing a heat treatment in an oxygen atmosphere at a temperature. At that time, the value of the refractive index of the obtained cladding layer 22 needs to be larger than the value of the refractive index of the optical waveguide 3 and is preferably equal to that of the quartz glass substrate 20.

  Next, as shown in FIG. 4A, a metal film 23 is formed on the clad layer 22 by sputtering, and thereafter, photolithography and reactive ion etching are performed as shown in FIG. 4B. By doing so, the metal film 23 is processed, and the electrophoresis groove processing mask 24 is formed.

  Next, as shown in FIG. 4C, the glass layer is etched by performing reactive ion etching to form a plurality of electrophoresis grooves 2 having a depth of 50 μm and a width of 50 μm.

Next, the electrophoretic groove processing mask 24 is removed by performing photolithography and reactive ion etching, and further, the optical fiber 4 is irradiated to the input end face of the optical waveguide 3 by fusion-bonding by CO ₂ laser light. Then, as shown in FIG. 4D, the first quartz glass substrate 1 is obtained.

  Next, as shown in FIG. 5A, an anhydrous synthetic quartz substrate 30 having an outer diameter of 4 inches and a thickness of 1000 μm is prepared. And as shown in FIG.5 (b), it penetrates for the injection | pouring of an electrophoretic liquid, and the application of a high voltage to the position corresponding to the both ends of the several electrophoresis groove | channel 2 provided in the 1st quartz substrate by machining. By forming the anode side electrode tank 8 and the cathode side electrode tank 9 provided with holes, the second quartz glass substrate 7 is obtained.

  Next, the first quartz glass substrate 1 on which the electrophoresis groove 2 and the optical waveguide 3 are formed, and the second quartz glass substrate 7 on which the anode side electrode tank 8 and the cathode side electrode tank 9 are formed. The surface is activated by scientific treatment. Then, as shown in FIG. 6, the first quartz glass substrate 1 and the second quartz glass substrate 7 that have been subjected to the surface treatment are overlapped and heated to about 500 ° C. in an electric furnace to thereby obtain two quartz glass substrates. Then, it is cut into a desired size by a dicing apparatus, and a quartz microchip 10 for electrophoresis as shown in FIG. 1 can be obtained.

[Configuration of platform board]
Next, FIG. 7 shows a process chart for creating a platform substrate according to the present embodiment.

  First, as shown in FIG. 7A, a metal plate 40 that is at least one size larger than the electrophoresis microchip is prepared. Then, as shown in FIG. 7B, the surface of the metal plate 40 is cut to form the chip mounting recess 12.

  Next, as shown in FIG. 7C, when the electrophoresis quartz microchip 10 is mounted on the chip mounting recess 12 on the upper surface of the chip mounting recess 12, the electrophoresis quartz microchip 10. A plurality of positioning protrusions 13 for fixing the electrophoresis quartz microchip 10 by fitting with the positioning recesses 6 are installed at positions corresponding to the positioning recesses 6 provided in To do. In addition, when performing the cutting process for forming the chip mounting portion 12, the cutting process may be performed so that the positioning 13 is formed in advance.

  Next, as shown in FIG. 7D, when the electrophoresis quartz microchip 10 is mounted on the chip mounting recess 12 on the upper surface of the chip mounting recess 12, the electrophoresis quartz microchip 10. A plurality of photodiodes 14 are installed at positions corresponding to the intersections of the plurality of electrophoresis grooves 2 and the optical waveguide 3 provided in FIG. Thereby, the platform substrate 11 as shown in FIG. 1 can be obtained.

  In addition, although the platform substrate 11 in this embodiment is produced by cutting a metal plate, the platform substrate according to the present invention is a metal processing other than cutting, for example, a method such as cold forging or casting. It may be created in Further, a material other than metal may be used. For example, when a platform substrate is formed using a resin, a method such as injection molding may be used.

  Moreover, the manufacturing process of the electrophoresis microchip in this embodiment is only one of the preferred embodiments of the manufacturing process of the electrophoresis microchip according to the present invention, and as a result, the effect according to the present invention can be obtained. Any microchip for electrophoresis may be obtained by any other process.

  The microchip for electrophoresis in the present embodiment uses quartz glass as a material. However, the present invention is not limited to this, and any light-transmitting material may be used. For example, acrylic resin, polycarbonate, It may be PET or the like.

  In addition, the size of the electrophoresis groove 2 in the present embodiment is only one preferred example, and may be an electrophoresis microchip having electrophoresis grooves of different sizes.

  In addition, a plurality of positioning recesses 6 and positioning protrusions 13 are provided in the present embodiment, but the present invention is not limited to this, and each of the positioning recesses 6 and positioning recesses 6 is provided. The thing which does not have the convex part 13, or the thing in which the number of the positioning recessed parts 6 is larger than the number of the positioning convex parts 13 may be sufficient.

  Furthermore, the number of photodiode recesses 5 in the present embodiment is the same as the number of photodiodes 14 mounted on the platform substrate 11 so that one photodiode 14 can be accommodated in one photodiode recess 5. However, the present invention is not limited to this, and a plurality of photodiodes 14 may be accommodated in one photodiode recess 5.

  Furthermore, the photodiode 14 may be embedded in the platform substrate 11 so that the upper surface thereof is flush with the upper surface of the chip mounting recess 12. In this case, the quartz microchip 10 for electrophoresis is used. In this case, the photodiode recess 5 is not necessary.

It is a perspective view which shows the structure of the electrophoresis apparatus in one Embodiment of this invention. It is sectional drawing which shows the structure of the electrophoresis apparatus in embodiment of this invention. It is the 1st manufacturing process figure of the 1st quartz glass substrate which constitutes the quartz microchip for electrophoresis in the embodiment of the present invention. It is a continuation of the manufacturing process figure of the 1st quartz glass substrate which comprises the quartz microchip for electrophoresis in embodiment of this invention. It is a manufacturing-process figure of the 2nd quartz glass substrate which comprises the quartz microchip for electrophoresis in embodiment of this invention. It is a manufacturing-process figure of the quartz microchip for electrophoresis using the 1st quartz glass substrate and the 2nd quartz glass substrate in embodiment of this invention. It is a manufacturing-process figure of the platform board | substrate which mounts the quartz microchip for electrophoresis in embodiment of this invention. It is a perspective view which shows the structure of the quartz microchip for electrophoresis in the conventional electrophoresis apparatus. It is a perspective view which shows the structure of the conventional electrophoresis apparatus.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 1st quartz glass substrate 2, 53 Electrophoretic groove 3 Optical waveguide 4 Optical fiber 5 Recessed portion for photodiode 6 Recessed portion for positioning 7 Second quartz glass substrate 8, 54 Anode side electrode tank 9, 55 Cathode side electrode tank 10 , 60 Quartz microchip for electrophoresis 11 Platform substrate 12 Recess for chip mounting 13 Protrusion for positioning 14 Photodiode 20, 51, 52 Quartz glass substrate 21 Core glass film 22 Cladding layer 23 Metal film 24 Electrophoresis groove processing mask 30 Anhydrous synthetic quartz substrate 61 Light source for UV excitation 62 Photo detector 63 Anode side electrode 64 Cathode side electrode

Claims (6)

A plurality of analysis channels through which a fluid containing the sample to be analyzed can flow;
An optical waveguide provided to intersect all of the plurality of analysis channels;
An excitation light source for supplying light to the optical waveguide;
A plurality of photodetectors for detecting fluorescence emitted in the plurality of analysis channels when the light from the excitation light source is guided into the plurality of analysis channels through the optical waveguide. An electrophoresis apparatus characterized by the above. The plurality of analysis channels and the optical waveguide are provided on an electrophoresis microchip made of a light-transmitting material,
The electrophoresis apparatus according to claim 1, wherein the plurality of photodetectors are mounted on a platform substrate that supports the electrophoresis microchip. The electrophoresis microchip is obtained by bonding substrates made of first and second light-transmitting materials,
The analysis channel is formed by covering a groove formed on the surface of the substrate made of the first light transmissive material on which the optical waveguide is formed with the substrate made of the second light transmissive material. The electrophoresis apparatus according to claim 1 or 2, wherein   4. The electrophoretic device according to claim 1, wherein the substrate made of the first and second light transmissive materials is a quartz glass substrate.   5. The optical waveguide provided on the substrate made of the first light-transmitting material has an optical fiber connected to an input end face or both of the input end face and the output end face. 6. Electrophoresis device.   The electrophoretic device according to claim 1, wherein an interval between the plurality of grooves formed in the substrate made of the first light transmitting material is 60 μm to 120 μm.

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