JP2008157814A - Substrate for analysis, and analyzer - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To realize a substrate for analysis that enables measurements reduced in background light and is capable of measuring with high sensitivity, only for the emission from a sample which is held by a sample-holding part. <P>SOLUTION: The analyzing substrate 1 enabling the optical analysis of a sample has a cavity provided to a substrate 2, and the flow channel-shaped cell 5 formed by a cover layer 3 formed so as to cover the cavity. Furthermore, the analyzing substrate 1 has an antireflection layer 4 provided on the surface on the detection side of emission produced, when the sample held to the flow channel-shaped cell 5 is irradiated with a light. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to an analysis substrate and an analysis apparatus for performing optical analysis of a sample.

  In recent years, techniques for analyzing biological samples such as DNA, RNA, and proteins have been remarkably developed. Examples of methods for detecting these biological samples include a fluorescence method, an absorption method, a chemiluminescence method, and a radioisotope method. Of these, the fluorescence method is mainly used by many researchers. This is because the fluorescence method has a high detection sensitivity and is easy to handle.

  In the fluorescence method, in order to perform measurement with higher detection sensitivity, it is important to reduce the amount of background light generated due to various causes that may be mixed in the fluorescence signal emitted from the sample.

  For example, in Non-Patent Document 1, a pinhole and a microlens are provided on the surface of the chip, and these spatially separate the optical path between the transmitted light of the excitation light and the fluorescence from the sample. A method for reducing the contamination of background light into a fluorescent signal is disclosed.

FIG. 8 shows a state of fluorescence detection using the chip 200 described in Non-Patent Document 1. The chip 200 includes a microchannel 201 through which a sample passes, and further includes a chromium layer 202a / b on the surface of the chip 200. The pinhole that is the opening of the chromium layer 202a on the excitation light incident side is provided with a microlens 203a for transmitting the excitation light into the chip 200, and is the opening of the chromium layer 202b on the fluorescence detection side. The pinhole is provided with a microlens 203b for transmitting fluorescence generated from the sample. The excitation light 204 is incident on the surface of the chip 200 as parallel light from an angle of 45 degrees. Of the excitation light 204, the light component that has passed through the excitation light incident side lens 203 a is condensed at one point on the microchannel 201. As a result, the sample flowing through the microchannel 201 emits fluorescence 206. Of these, only the light passing through the microlens 203b on the fluorescence detection side is detected by the light emission detector. On the other hand, the transmitted light 207 transmitted through the microchannel 201 does not pass through the chromium layer 202b on the fluorescence detection side of the chip. Therefore, the fluorescence 206 and the transmitted light 207 of the sample are spatially separated. As a result, it is possible to reduce the background light generated by the transmitted light of the excitation light from being mixed into the fluorescent signal emitted by the sample.
"Performance of an Integrated Microoptical System for Fluorescence Detection in Microfluidic Systems", Jean-Christophe Roulet et al, Anal. Chem. 2002, 74, 3400-3407 JP 11-30703 A (published February 2, 1999) JP 2003-66003 A (published on March 5, 2003)

  However, in the method described in Non-Patent Document 1, the light beam of the excitation light 204 is irradiated in a wider range than the range occupied by the microlens 203a in the chromium layer 202a on the excitation light incident side. As a result, a part of the excitation light 204 is irradiated outside the range occupied by the microlens 203a, causing the following problems.

  More specifically, a part of the excitation light 204 irradiated outside the range occupied by the microlens 203a is reflected by the chromium layer 202a on the excitation light incident side, and then is reflected several times inside the measurement apparatus. The reflected light then becomes stray light in the measuring apparatus and is likely to cause background light. In addition to the reflected light, the background light is diffusely reflected inside the chip 200, passes through the pinholes that are openings of the chromium layers 202a and b on the fluorescence detection side, jumps out of the chip, and is reflected again. Repeated reflected light is also included. The background light generated by the reflection described above is incident on the light emission detector and may reduce the detection sensitivity.

  The present invention has been made to solve the above problem, and it is possible to reduce the amount of background light reaching the light emission detector and to provide an analysis substrate and an analysis apparatus that enable light emission detection with higher detection sensitivity. The purpose is to provide.

  In order to solve the above problems, the analysis substrate of the present invention has a sample holding portion formed by a depression provided on the surface of the base substrate and a cover layer formed so as to cover the depression. An analysis substrate capable of optical analysis of a sample held in a sample holding portion is characterized in that an antireflection means is provided on the surface of the analysis substrate.

  According to said structure, the reflection preventing means is provided in the surface of the board | substrate for analysis. Therefore, reflection of light can be suppressed on the surface of the analysis substrate, and when the sample is optically analyzed, the amount of background light reflected on the surface of the analysis substrate and incident on the device that detects luminescence is reduced. Can do.

  Therefore, according to the above configuration, it is possible to perform measurement with less background light, and to realize an analytical substrate capable of measuring only light emitted from the sample held in the sample holding portion in the analytical substrate with high sensitivity. Can do.

  In the analysis substrate of the present invention, a reflection layer may be provided between the base substrate and the antireflection means.

  According to the above configuration, since the reflective layer is provided, for example, even if a part of scattered light generated at the end of the lens for collecting incident light passes through the analysis substrate, it is reflected. Thus, it is possible to reduce the transmission to the light emission detection side. Therefore, by providing a reflection layer in addition to the antireflection means, it is possible to reduce the amount of background light and more reliably measure only light emission.

  In addition, for example, even when the reflecting means is a film and the thickness thereof is thin, by using the reflecting layer in combination, the amount of background light can be reduced and sensitive measurement can be performed. An analysis substrate can be realized.

  In the analysis substrate of the present invention, the antireflection means may be a black carbon compound layer or a non-light emitting pigment-containing layer.

  According to said structure, the reflection preventing means which has the outstanding reflection preventing function with a high light absorption function can be obtained.

  In the analysis substrate of the present invention, the antireflection means may be a fine concavo-convex structure or a fine porous structure formed on its surface.

  According to the above structure, the antireflection means can be formed without increasing the thickness of the analysis substrate.

  Further, the analysis substrate of the present invention may be configured such that an opening is provided in the antireflection means located on the sample holder.

  According to said structure, the light emission from the sample hold | maintained at the sample holding part is analyzed by the opening provided in the antireflection means located on the said sample holding part, without being attenuated by the antireflection means. It can be taken out of the substrate. Thereby, it is possible to realize light emission detection with better detection sensitivity.

  In the analysis substrate of the present invention, the antireflection means may be an antireflection film composed of a multilayer film of inorganic oxide.

  According to the above configuration, the light emission from the sample can be taken out of the analysis substrate without providing the opening as described above in the antireflection means, and the reflection of light on the analysis substrate surface can be suppressed. Is possible.

  In the analysis substrate of the present invention, the reflection layer may be provided with a guide means for guiding incident light for optical analysis.

  According to the above configuration, when the incident light is guided to the guide means on the analysis substrate using, for example, an optical pickup, the incident light can be accurately guided to the position of the sample holder.

  As a result, even when light is incident on a position other than the position of the sample holder, it is possible to accurately guide the incident light to the position of the sample holder and detect light emission from the sample.

  Moreover, in order to solve the above-mentioned problems, the analyzer of the present invention sandwiches any one of the above-described analysis substrates between the analysis substrate and the light irradiation means for making incident light incident on the analysis substrate. It is characterized by comprising photodetection means located on the opposite side of the light irradiation means and disposed outside the optical path of the incident light.

  According to the above configuration, since the light detection means provided in the analyzer is arranged outside the optical path of the incident light, transmitted light that has passed through the analysis substrate can be prevented from entering the light detection means. . Therefore, the background noise due to the transmitted light that has passed through the analysis substrate can be kept low, and the light emission from the sample can be measured with high sensitivity.

  Further, the analyzer according to the present invention causes the incident light to enter the analysis substrate having the guide means, and detects the reflected light from the analysis substrate and the optical pickup along the guide means. Driving means for moving the optical pickup in a specific direction, and control means for controlling the optical pickup so that the incident light follows the guide means and enters based on a detection signal output from the optical pickup. It is characterized by that.

  According to said structure, the said optical pick-up reads the guide means on the said board | substrate for an analysis, and can focus an incident light on the exact position on the board | substrate for an analysis. Moreover, the reflected light from the reflective layer that is generated when the light is incident can further follow the guide means.

  As a result, the sample in the sample holder can be accurately irradiated with excitation light, and an optical change from the sample can be detected with good detection sensitivity and further automatically and accurately.

  As described above, the analysis substrate according to the present invention includes antireflection means on its surface on the side that detects light emission that occurs when the sample held by the sample holder is irradiated with light.

  According to said structure, the reflection preventing means is provided in the surface of the board | substrate for analysis. Therefore, reflection of light can be suppressed on the surface of the analysis substrate, and when the sample is optically analyzed, the amount of background light reflected on the surface of the analysis substrate and incident on the device that detects luminescence is reduced. Can do.

  Therefore, according to the above configuration, it is possible to perform measurement with less background light, and to realize an analytical substrate capable of measuring only light emitted from the sample held in the sample holding portion in the analytical substrate with high sensitivity. Can do.

[Embodiment 1]
An embodiment of the present invention will be described below based on FIG. 1 and FIG. In the present embodiment, a flow path cell is used as an example of a sample holder that holds a sample provided on an analysis substrate. The sample holder for holding the sample is not particularly limited, and examples thereof include a channel cell used for electrophoresis and chromatography, a microcell used for DNA hybridization, and the like.

  FIG. 1 is a perspective view of an analysis substrate 1 in the present embodiment. As shown in FIG. 1, the analysis substrate 1 includes a substrate 2 (basic substrate) having a groove-like depression having a rectangular vertical cross-sectional shape, a cover layer 3 that covers the surface of the substrate 2, and a cover layer 3. And an antireflection layer 4 formed. Further, the analysis substrate 1 includes a channel cell 5 capable of holding a sample solution (sample) formed by the recess between the substrate 2 and the cover layer 3, and further on the channel cell 5. And an opening 6 provided in a part of the antireflection layer 4.

  The substrate 2 is formed of a light transmissive material such as plastic or glass. It may be made of other materials. The substrate 2 has a groove-like depression whose surface has a rectangular vertical cross-section with respect to the extending direction of the channel-like cell 5, and the groove-like depression has a depth depending on the thickness of the substrate. It is formed in a size within a range of several μm to several hundred μm and a width of several tens μm to several mm.

  The cover layer 3 is a layer that covers the surface of the substrate 2 having the groove-like depression. The cover layer 3 is formed of a light-transmitting material such as plastic or glass, as with the substrate 2. For example, a film-like plastic having a thickness of 200 μm or less may be used. The cover layer 3 and the substrate 2 can be bonded to each other by a method using an adhesive such as a UV coin resin, a bonding method using thermocompression bonding, or the like. Any method may be used as long as it is an adhesion method that does not leak out of the road.

  The antireflection layer 4 is formed on the cover layer 3, that is, so as to cover the surface of the analysis substrate 1. Here, the antireflection layer 4 is formed to absorb background light accompanying stray light generated in the analyzer. The material of the antireflection layer 4 may be any material that does not emit light and has a high absorbance. In particular, a black carbon compound or a non-emitting pigment is preferable because of its high durability. In addition, as a method for producing the antireflection layer 4, these antireflection agents, a solvent capable of dispersing or dissolving the antireflection agent, and a polymer capable of being dissolved in the solvent were mixed. A method of applying the solution on the cover layer 3 and drying it may be used. As a coating method of the antireflection agent, a method other than the above method may be used.

  The channel-like cell 5 is formed by covering a groove-like depression provided on the surface of the substrate 2 with the cover layer 3. Thereby, the channel-like cell 5 has the same size as the groove-like depression, that is, the size defined in the range of several μm to several hundred μm in depth and several tens of μm to several mm in width.

  In addition, an opening 6 is formed in a part of the antireflection layer 4 located on the channel cell 5, and the antireflection layer is not formed. As will be described in detail later, the antireflection layer 4 of the analysis substrate 1 is formed by forming the opening 6 to emit light from the sample in the channel cell 5 excited by incident light. It can permeate | transmit toward the light emission detector arrange | positioned at the surface side.

  The analysis substrate 1 according to the present embodiment is characterized by the antireflection layer 4, and this effect will be described with reference to FIG.

  FIG. 2 is a schematic diagram of the analyzer according to the present embodiment using the analysis substrate 1, and the analysis substrate 1 shown in FIG. 1 is cut along the extension direction of the flow channel cell 5. It is sectional drawing of the part enclosed with the dotted line shown by. In FIG. 2, incident light 11, a lens 12 for condensing the incident light 11 on the flow path cell 5 of the analysis substrate 1, and light emission emitted from the sample in the flow path cell 5 by the collected incident light 11. 13, an optical filter 15 installed outside the optical path of the transmitted light 14 transmitted through the analysis substrate 1, and a light emission detector 16 arranged on the back side of the optical filter 15 are illustrated. The analysis apparatus according to the present embodiment includes at least an incident optical system device (light irradiation means) (not shown) and a light emission detector (light detection means) 16 that is each device of a light emission detection system. Is irradiated with light, and the light emission from the sample held in the flow path cell 5 is analyzed.

  The dotted line in the outermost frame in FIG. 2 includes the incident optical system, each device of the light emission detection system, or the component of each device that constitutes the analyzer of the present embodiment for analyzing the sample of the analysis substrate 1. A container 18 is shown. In addition, a conventionally well-known apparatus can be used for the incident optical system, and description and illustration are omitted.

  The optical filter 15 is an optical component having a property that the transmittance of the light of the wavelength included in the incident light 11 is low and the transmittance of the light of the wavelength included in the light emission 13 from the sample is high. A conventionally known light emission detector 16 can be used.

  According to the above structure, since the light emission detector 16 is disposed outside the optical path of the transmitted light 14, the background light due to the transmitted light 14 can be reduced, and the light emission 13 of the sample can be detected with high sensitivity. Can do.

  On the other hand, since various stray lights other than the light emission 13 emitted from the sample are mixed in the light emission detector 16 as background light, the light emission detection sensitivity may be lowered. For example, although the transmitted light 14 does not directly enter the light emission detector installed outside the optical path, a part of the transmitted light 14 is reflected by the inside of the container 18 including the entire analysis device, so that part of the transmitted light 14 is stray light. There is a possibility that the light enters the light emission detector 16 as 17a. The stray light 17 a incident on the light emission detector 16 is finally reflected on the surface of the analysis substrate 1 and incident on the light emission detector 16 as shown in the figure. However, since the antireflection layer 4 is formed on the surface of the analysis substrate 1 in the present embodiment on the side of the light emission detector 16, the stray light 17a is not reflected on the surface of the analysis substrate 1, and the light emission detector 16 is used. Is not reached (indicated by x in the figure).

  In this way, the luminescence detector 16 is arranged so as to face the analysis substrate surface in order to capture the luminescence 13 emitted from the sample, but by preventing reflection of stray light by the antireflection layer 4 on the substrate surface, It is possible to prevent stray light from reaching the light emission detector 16. As a result, it is possible to reduce the amount of stray light mixed in the light emission detector 16.

  The cause of stray light is not only the transmitted light 14 of the incident light 11. For example, when the lens 12 that converges the incident light 11 is irradiated with light, part of the light is scattered at the end of the lens. This scattered light is not used for excitation of the sample held in the flow channel cell 5, and a part of it is reflected several times in the container 18 including the whole analyzer, and the emission detector as stray light 17b. There is a possibility of heading to 16.

  However, since the antireflection layer 4 is formed on the surface of the analysis substrate 1 according to the present embodiment as described above, the stray light 17b does not enter the light emission detector 16 (in FIG. 2, × Luminescence measurement with good detection sensitivity is possible.

  Furthermore, as a generation source of stray light, various components such as an optical component related to incidence of incident light may be the generation source in addition to the generation source. When the light emission detector 16 is arranged so as to face the substrate surface side of the analysis substrate 1 as described above, the stray light reflected by the substrate surface is highly likely to enter the light emission detector 16. It is effective to form the antireflection layer 4 on the surface of the analysis substrate 1 as in the embodiment. The antireflection layer 4 can prevent stray light from traveling to the light emission detector 16.

  As described above, the analysis substrate 1 in the present embodiment can reduce stray light generated in the analysis apparatus and detect light emission with high sensitivity.

  In the description using FIG. 2, the incident light 11 is described as being collected by the lens 12 and incident on the channel cell 5, but the incident method of the incident light 11 is not limited to this. For example, the present application is effective even when parallel light is incident on the channel cell 5.

  In the above description, as an example of the antireflection means according to the present invention, the example in which the detection sensitivity is improved by forming the antireflection layer 4 on the analysis substrate 1 has been described. However, the present invention is not limited to this, and a well-known antireflection structure (for example, a fine concavo-convex structure or a fine porous structure) is formed on the surface of the analysis substrate to reduce background light. I do not care. Here, the size of the irregularities of the fine structure is preferably 100 to 300 nm and the pitch is preferably 100 to 1000 nm, but these values are not limited.

  In the above example, the antireflection layer 4 is configured to cover almost the entire surface of the cover layer 3, but it is not always necessary to cover the entire surface, and the substrate closer to the light emission detector 16 than the opening 6 in FIG. It is good also as a structure which covers only the surface 300. FIG. In this case, although the antireflection function is slightly lower than the case where the entire surface of the substrate is covered, since the portion on the substrate where the reflected light is most likely to enter the light emission detector 16 is covered near the light emission detector 16, the reflection prevention for the covered area The function is very large.

Further, on the analysis substrate 1 in the present embodiment, an opening 6 for taking out the light emission 13 from the sample held in the flow channel cell 5 in the direction of the light emission detector 16 is formed. Need not be formed. For example, when a well-known antireflection film of a lens is used instead of the antireflection layer 4, the opening 6 is not necessary. This is because the antireflection film generally has a structure composed of a multilayer film of inorganic oxide, and has a property of transmitting light but suppressing reflection. Such a structure of the antireflection film is disclosed in Patent Document 1, for example. According to Patent Document 1, the antireflection film is formed by alternately laminating a high refractive index film and a low refractive index film in multiple layers. As an example, an antireflection film having a multilayer structure composed of ZrO ₂ (high refractive material) forming the high refractive index film and SiO ₂ (low refractive material) forming the low refractive index film is disclosed. When this antireflection film is used, the light emission 13 from the sample in the flow path cell 5 passes through the antireflection film, passes through the light emission detector 16, and reaches the light emission detector 16. On the other hand, since the stray light 17 is not reflected on the surface of the analysis substrate, it does not reach the light emission detector 16. Therefore, when the antireflection film is provided on the substrate surface in this way, the opening 6 is not necessary.

Moreover, in this Embodiment, although the vertical cross-sectional shape of the channel-shaped cell was made into the rectangle, not only that but another shape may be sufficient.
[Embodiment 2]
Other embodiments of the present invention will be described below with reference to FIGS. 3 and 4. Also in the following embodiments, as in the first embodiment, a description will be given using a flow channel cell as an example of a cell holding a sample provided on an analysis substrate.

  3A is a perspective view of the analysis substrate 21 in the present embodiment, and FIG. 3B is a cross-sectional view of the flow path cell 25 held by the analysis substrate 21 along the extension direction ( FIG. 3A shows a portion indicated by a dotted line.

  As shown in FIG. 3A, the analysis substrate 21 covers a substrate (basic substrate) 22 having a groove-like depression whose vertical cross-sectional shape is rectangular, and a surface of the substrate 22 having the groove-like depression. The cover layer 23 includes a reflection layer 28 formed on the cover layer 23 and an antireflection layer 24 formed thereon. In addition, the analysis substrate 21 includes a flow channel cell 25 capable of holding a sample formed by a groove-shaped recess between the substrate 22 and the cover layer 23, and an antireflection layer 24 on the flow channel cell 25. The opening part 26 provided in a part of is provided.

  The difference between the analysis substrate 21 in the present embodiment and the analysis substrate 1 shown in the first embodiment is that a reflection layer 28 is formed between the cover layer 23 and the antireflection layer 24 on the analysis substrate 21. It is a point. The reflective layer 28 formed on the cover layer 23 of the analysis substrate 21 is made of a metal thin film such as aluminum or silver, and may have a thickness of several to several hundred nm. Further, instead of the reflective layer made of these metals, a reflective layer made of a dielectric multilayer film may be used. The configuration other than the reflective layer 28 is the same as the configuration of the first embodiment shown in FIG. 1. The substrate 22 is the substrate 2, the cover layer 23 is the cover layer 3, the antireflection layer 24 is the antireflection layer 4, Since the flow cell 25 corresponds to the flow cell 25 and the opening 26 corresponds to the opening 6, the description thereof is omitted.

  The effect of the reflective layer 28 newly added in the present embodiment will be described below with reference to FIG.

  FIG. 4 is a schematic diagram of an analysis apparatus using the analysis substrate 21. Similar to the analysis apparatus shown in the first embodiment, the incident light 31 and the flow path cell of the analysis substrate 21 are shown. A lens 32 that condenses light 25, light emission 33 emitted from the sample in the channel cell 25 by the collected incident light 11, and optics installed outside the optical path of the transmitted light 34 that has passed through the analysis substrate 21. A filter 35 and a light emission detector 36 disposed on the back side of the optical filter 35 are shown. In addition, the dotted line of the outermost frame in the drawing indicates the container 38 that includes the components of each device of the incident optical system and the light emission detection system that constitute the analyzer. Further, the analysis substrate 21 shown in FIG. 3 is cut along the extension direction of the flow path cell 25 in the analysis substrate 21 as shown in FIG. It is sectional drawing of the part enclosed by the dotted line part. The analysis apparatus according to the present embodiment includes at least an incident optical system device (light irradiation means) (not shown) and a light emission detector (light detection means) 36 which are each device of a light emission detection system. Is irradiated with light, and the light emission from the sample held in the channel cell 25 is analyzed.

  As in the first embodiment, the optical filter 35 has an optical component having a property of low transmittance of light having a wavelength included in the incident light 31 and high transmittance of light having a wavelength included in the light emission 33 emitted from the sample. It is.

  In the analysis apparatus according to the present embodiment, according to the above structure, as described in the first embodiment, since the emission detector 36 is disposed outside the optical path of the transmitted light 34, the background due to the transmitted light 34 is reduced. Light can be reduced, and light emission 33 emitted from the sample can be detected with high sensitivity. Further, the antireflection layer 24 on the substrate surface suppresses reflection of stray light on the substrate surface, so that stray light can be prevented from traveling toward the light emission detector 36. As a result, the amount of stray light mixed in the light emission detector 36 can be reduced.

  The reflective layer 28 provided on the analysis substrate 21 plays a role as described below. As described with reference to FIG. 2 in the first embodiment, the incident light 31 is scattered at the end of the lens 32 that collects the incident light 31. A part of this becomes scattered light 37 that goes directly in the direction in which the light emission detector 36 is installed. In order to measure the emission detection with high sensitivity, it is necessary to prevent the scattered light 37 from being transmitted toward the emission detector 36. Here, by forming the reflective layer 28 made of a metal thin film or the like on the cover layer 23, even if the reflective layer 28 is thin (several to several hundred nm), the scattered light 37 is efficiently reflected. Can be made. Therefore, when the antireflection layer 24 is very thin and the scattered light 37 is transmitted in the direction of the light emission detector 36, the transmission of the scattered light 37 can be reduced by using the reflective layer 28 together. Can do.

  As described above, the analysis substrate 21 in the present embodiment can reduce the stray light generated in the analysis apparatus and the light transmitted in the direction of the light emission detector 36, and can detect light emission with high sensitivity.

[Embodiment 3]
Still another embodiment of the present invention will be described below with reference to FIGS. In the following embodiments, a case where a circular analysis disk 41 is used as an analysis substrate and a sample is analyzed using electrophoresis will be described. In this embodiment, a case where fluorescence from a sample to be electrophoresed is detected will be described.

  FIG. 5A is a front view showing an analysis disk 41 that can be measured by electrophoresis, and is also a schematic diagram of a circular analysis disk. As shown in FIG. 5A, the analysis disk 41 includes a center hole 42 for fixing the analysis disk 41 to the center of the turntable of the analyzer, and an analysis chip 43 for analyzing the sample by electrophoresis. And a guide groove 44 and an address information recording unit 45. In this embodiment, the analysis disk 41 is described as including four analysis chips 43, but the number is not limited. The cross-sectional structure (layer structure) of the analysis disk 41 will be described later.

  Each analysis chip 43 has liquid reservoirs / injection ports 46 a to 46 d and an electrophoresis path (flow channel cell) 47. In FIG. 5A, the grayed out part is a part where the antireflection layer 48 is formed. An opening 49 is formed in a part of the antireflection layer 48 on the migration path 47. Since the antireflection layer 48 is the same as the antireflection layer 4 of the first embodiment, and the opening 49 is the same as the opening 6 of the first embodiment, description thereof is omitted. To do.

  In the migration path 47 of the analysis chip 43, a first migration path 47a having a long length and a second migration path 47b having a short length are formed in a cross shape. The structure of the analysis chip 43 is a structure that is generally used in analysis by electrophoresis. For example, Patent Document 2 discloses an analysis example together with an analysis example.

  The first migration path 47a extends in the radial direction of the analysis disk 41, and the second migration path 47b extends perpendicular to the first migration path 47a. One liquid reservoir / injection port 46 a to 46 d is arranged so as to communicate with the four ends of the migration path 47. That is, a liquid reservoir / injection port 46a is disposed at one end of the second migration path 47b, and a liquid reservoir / injection port 46c is disposed at the other end, and the liquid reservoir / injection port 46b is disposed at one end of the first migration path 47a. A liquid reservoir / injection port 46d is arranged in the section.

  FIG. 5B is a cross-sectional view of the analysis disk 41 taken along the dotted line shown in FIG. 5A, that is, a plane perpendicular to the extending direction of the first migration path 47a at the site where the opening 49 is formed. FIG. It is also a diagram showing the layer structure of the analysis disk 41. The analysis disk 41 includes a substrate (basic substrate) 51 having a groove-like depression whose vertical cross-sectional shape is rectangular, a reflection layer 52 formed on a portion of the substrate 51 other than the depression, and an insulation covering the reflection layer 52. And a protective layer 53 having properties. Further, the guide groove 44 formed on the protective layer 53, the cover layer 54 covering the surface of the substrate 51 having the groove-like depression, the antireflection layer 48 formed on the cover layer 54, the substrate 51, An electrophoresis path 47 capable of holding the sample formed by the depression between the cover layer 54 and the cover layer 54 is provided. An arrow in the figure indicates incident light 102 incident on the analysis disk 41 from an optical pickup 101 (illustrated later in FIG. 7) used for a CD, DVD or the like serving as a light source in the present embodiment.

  If fluorescence detection is performed using the analysis disk 41 having the above-described structure, the disk surface is covered with the antireflection layer 48. Therefore, the light emission detection using the analysis substrate of the first and second embodiments is the same. In addition, since stray light generated in the analyzer can be reduced, highly sensitive fluorescence detection is possible.

  When performing electrophoretic measurement using a metal thin film for the reflective layer 52, the metal thin film is exposed to the electrophoretic path 47 without the insulating protective layer 53, so that a false electrode can be formed in electrophoretic measurement. Therefore, by covering with an insulating protective layer 53, generation of false electrodes can be prevented. The reason is as follows. When the reflective layer 52 is a conductive material, a conductor exists along the migration path 47. Even if a voltage for electrophoresis is applied thereto, an electric field is not applied in the flow path due to this conductor, or a false electrode is formed in the flow path. That is, good electrophoresis becomes difficult.

  Therefore, by covering the conductive reflective layer 52 with the protective layer 53 made of an insulating material (dielectric material), the electric field in the flow path becomes uniform and electrophoresis becomes possible. However, as described above, when a metal thin film is used, it is difficult to apply an electric field for electrophoresis. Therefore, it is necessary to take a distance between the flow path and the metal thin film as much as possible. This distance is determined by the balance with the electric field strength. When a dielectric multilayer film is used as the reflective layer 52 instead of the metal film, it is not necessary to provide a distance between the reflective layer 52 and the migration path 47, and the protective layer 53 can be omitted.

  Below, the detail of the electrophoresis method using the disc 41 for analysis is demonstrated.

  In the analysis process (electrophoresis process) in the analysis disk 41 described above, a gel solution as a buffer is injected from the liquid reservoir / injection port 46d and filled in the electrophoresis paths 47a and 47b. Next, a gel solution as a buffer is also injected into 46b and 46c. Thereafter, a sample solution in which a sample as an analysis sample is dissolved is injected into the liquid reservoir / injection port 46a.

  For example, a DNA fragment is used as the sample. This DNA fragment needs to be fluorescently labeled by a method such as introduction of a fluorescent dye at the end or mixing with an intercalator type fluorescent dye. For example, a method of fluorescent staining when DNA passes through the flow path by adding an intercalator type fluorescent dye to the gel solution that is the buffer described above may be used.

  Next, the electrodes supplied from the power supply of the analyzer are arranged in the liquid reservoir / injection ports 46a to 46d, and these electrodes are connected as follows. First, the electrode arranged at the liquid reservoir / injection port 46c is connected to a + voltage power source (several tens to several hundred volts), and the electrodes arranged at the liquid reservoir / injection ports 46a, 46b, 46d are connected to the ground. As a result, a positive voltage (several tens to several hundred volts) is applied to the liquid reservoir / injection port 46c, zero volt is applied to the liquid reservoir / injection port 46a, and the sample (DNA fragment: negatively) injected into the migration path 47. Charge) migrates in the second migration path 47b from the liquid reservoir / injection port 46a (-electrode) toward the liquid reservoir / injection port 46c (+ electrode).

  Next, after the migrated sample reaches the crossing point of the cross in the migration path 47, the electrode arranged at the liquid reservoir / injection port 46d is connected to a + voltage power source (several tens to several kilovolts). On the other hand, the electrodes disposed in the liquid reservoir / injection ports 46a and 46c are connected to a + voltage power source (several volts to several kilovolts) that outputs a voltage lower than the voltage of the electrode disposed in the liquid reservoir / injection port 46d. The electrode disposed in the liquid reservoir / injection port 46b is connected to the ground. As a result, a positive voltage (several tens to several kilovolts) is applied to the electrode of the liquid reservoir / injection port 46d, zero volt is applied to the electrode of the liquid reservoir / injection port 46b, and the sample injected into the migration path 47 is the first sample. In the one migration path 47a, the electrophoresis proceeds from the intersection of the ten letters toward the liquid reservoir / injection port 46d. At this time, the mobility of the sample (DNA) varies depending on the molecular weight due to the molecular sieving effect of the filled gel, and can be divided into fragments of each molecular weight fragment. Since the principle of electrophoretic analysis in such a microcapillary is well known as disclosed in the above-mentioned Patent Document 2, detailed description thereof is omitted.

  The analysis disk 41 includes the analysis chip 43, a guide groove 44 for guiding the incident light 102 from the optical pickup 101, and an address information recording unit 45 in which address information is recorded. When the analysis disk 41 rotates, the optical pickup 101 accesses the incident light 102 to the desired radial position of the analysis disk 41 while reading the radial position included in the address information of the analysis disk 41. Therefore, fluorescence from the sample can be detected at a desired position in one analysis chip 43, that is, at the opening 49 on the migration path 47.

  In addition, a total of four analysis chips 43 are formed, and each is arranged at an interval of 90 degrees. Furthermore, between the adjacent analysis chips 43, a guide groove 44 and an address information recording unit 45 are formed side by side in the circumferential direction. Since the incident light accesses the analysis chips 43 arranged in the circumferential direction while reading the number of the analysis chip 43 included in the address information of the address information recording unit 45, the desired analysis among the four analysis chips 43 is performed. Analysis of the sample (fluorescence detection) can be performed on the chip for use. At this time, when the incident light 102 scans one track (guide groove 44) in the circumferential direction, these are repeated in the order of tracking area of the guide groove 44 → address information recording unit 45 → tracking area of the guide groove 44 → migration path 47. Will be scanned.

  FIG. 6 is a longitudinal sectional view of the analysis disk 41 showing a state in which the incident light 102 passes through the opening 49 and moves to the tracking region of the guide groove 44 in tracking of the incident light 102 using the analysis disk 41. is there.

  As shown in the figure, the incident light 102 is condensed on the guide groove 44 through the optical path shown in the figure. Further, the incident light 102 is reflected by the reflective layer 52 and returns to the lens 71 as reflected light 63. In this process, the incident light 102 is diffracted in the guide groove 44 and the distribution of the diffracted light is reflected in the reflected light 63, so that the incident light 102 is caused to follow the guide groove 44 by controlling the optical pickup 101. Can do. Therefore, if the guide groove 44 is formed so as to cross the desired detection position of the migration path 47 (but not present in the migration path 47 itself), the incident light is on the extension of the guide groove 44. The fluorescence of the sample can be detected (analyzed) at a desired position of the migration path 47 across the desired position. Further, as described above, since the tracking control is maintained in a state immediately before the incident light 102 passes through the migration path 47 during the period in which the incident light 102 passes through the migration path 47, a guide groove is formed in the migration path 47 itself. Even if it does not exist, stable tracking is continuously performed from the guide groove 44a before passing through the migration path 47 to the guide groove 44b on the extension thereof. In the above example, the guide groove 44 has been described as an example of the guide means. However, the present invention is not limited to this, and for example, a wobble pit may be used as the guide means.

  FIG. 7 is a block diagram showing the configuration of the main parts of the incident light control device 70 and the optical pickup (light irradiation means) 101 in the analyzer. The optical pickup 101 includes a lens 71 and an actuator 101 a that drives the lens 71. Furthermore, an optical system 101b composed of optical components such as a semiconductor laser, a photodetector, and a beam splitter is provided. The incident light control device 70 includes an address reproduction circuit 72, a controller 73 (control means), and a servo circuit 74 (control means).

  In the control device 70, the incident light 102 emitted from the optical pickup 101 is collected by the lens 71 and is incident on the analysis disk 41. The incident light 102 is applied to the guide groove 44 or the address information recording unit 45 of the analysis disk 41. The reflected light from the guide groove 44 or the address information recording unit 45 returns to the optical pickup 101 again, and the optical pickup 101 outputs a focus error signal / track error signal 75.

  The focus error signal / track error signal 75 is fed back to the actuator 101 a via the servo circuit 74. Thereby, the servo circuit 74 controls the actuator 101a so that the incident light is guided as the new incident light 102 to the guide groove 44 or the address information recording unit. The light quantity detection signal 76 is input to the address reproduction circuit 72, and the address reproduction circuit 72 detects the address information 77 from the light quantity detection signal 76 and sends this address information 77 to the controller 73. The controller 73 outputs a control signal 78 to the servo circuit 74 while confirming the address information 77, and performs a process of allowing the incident light 102 to access the desired guide groove 44. Accordingly, the incident light 102 can be moved to a desired position on the analysis substrate 41 and can be accurately incident on the migration path 47.

  Note that the tracking state in the immediately preceding guide groove 44 is maintained at the position of the opening 49 of the guide groove 44. For this purpose, it is sufficient that the time crossing the opening 49 is sufficiently shorter than the response time of the servo (proportional to the reciprocal of the servo band). Alternatively, the servo operation may be temporarily held when crossing the opening 49 (hold operation). Accordingly, even when the incident light 102 crosses the migration path 47, tracking is not disturbed, and the sample can be irradiated with the incident light 102 accurately. Then, the incident light 102 can reach the guide groove 44 again and return to the tracking operation.

  As described above, also in the present embodiment, since the surface of the analysis disk 41 is covered with the antireflection layer 48, as described in the first and second embodiments, it occurs in the analyzer. Stray light can be reduced, and sensitive fluorescence detection is possible.

  Further, if the analysis disk 41 is used, it is possible to measure a sample using electrophoresis.

  Furthermore, the analyzer in the present embodiment includes an optical pickup 101 that reads the guide groove 44 formed in the reflective layer 52, and can cause the incident light 102 to follow the guide groove 44. Therefore, the incident light 102 can accurately irradiate the migration path 47 on the extension of the guide groove 44. Thereby, the analyzer can detect an optical change from the sample with high detection sensitivity and further automatically and accurately.

  In the above-described embodiment, the case of fluorescence detection in electrophoresis has been described. However, the present invention is not limited to this, and can be applied to fluorescence detection such as chromatography. Further, the analysis substrate is not limited to the disk shape (analysis disk 41), but may be other shapes. In the above description, an example of fluorescence detection from a sample has been described. However, the present invention can also be applied to an apparatus for detecting a sample that emits light when excited by an external light source, such as phosphorescence.

  In the above embodiment, the details of the electrode wiring method for performing electrophoresis, the device for supplying power, the circuit for detecting fluorescence are not described, but these methods and devices are conventionally known. Can be used. These methods and apparatuses are the same as, for example, the methods and apparatuses described in Japanese Patent Application Laid-Open No. 2006-58250.

  The present invention is not limited to the above-described embodiments, and various modifications are possible within the scope shown in the claims, and embodiments obtained by appropriately combining technical means disclosed in different embodiments. Is also included in the technical scope of the present invention.

  The analysis substrate and the analysis apparatus of the present invention can be applied to the field of analyzing a very small amount of sample. For example, it can be used in the medical device field, the measuring device field, and the like.

It is a perspective view of the board | substrate for analysis in Embodiment 1 of this invention. It is the schematic of the analyzer which used the board | substrate for analysis in Embodiment 1 of this invention. (A) is a perspective view of the analysis substrate in Embodiment 2 of the present invention, and (b) is a cross-sectional view along the extension direction of the flow path cell held by the analysis substrate of FIG. is there. It is the schematic of the analyzer which used the board | substrate for analysis in Embodiment 2 of this invention. (A) is a front view showing an analysis disk that can be measured by electrophoresis, and (b) is in the extending direction of the first electrophoresis path at the site where the opening of the analysis disk of FIG. It is sectional drawing of a perpendicular | vertical surface. FIG. 6 is an explanatory diagram showing a state in which incident light passes through a flow path and moves to a tracking region of a guide groove using a longitudinal section of the analysis disk in the analysis apparatus using the analysis disk shown in FIG. 5. It is a block diagram which shows the structure of the principal part of the incident light control apparatus with which the incident light irradiation apparatus of the analyzer in Embodiment 3 of this invention is provided, and an optical pick-up. It is sectional drawing of the conventional analysis chip | tip.

Explanation of symbols

1,21 Analysis substrate 2, 22, 51 Substrate (basic substrate)
3, 23, 54 Cover layers 4, 24, 48 Antireflection layer (antireflection means)
5, 25 Channel-like cells 6, 26, 49 Apertures 11, 31, 102, 207 Incident light 12, 32, 71 Lens 13, 33 Light emission 14, 34 Transmitted light 15, 35 Optical filter 16, 36 Light emission detector 17a 17b Stray light 18, 38 Container 28, 52 Reflective layer 37 Scattered light 41 Analysis disk (analysis substrate)
42 Center hole 43 Analysis chips 44, 44a, 44b Guide groove 45 Address information recording section 46a, b, c, d Reservoir / injection port 47 Electrophoresis path 47a First migration path 47b Second migration path 53 Protective layer 63 Reflected light 70 incident light control device 72 address reproduction circuit 73 controller 74 servo circuit 75 focus error signal / track error signal 76 light quantity detection signal 77 address information 78 control signal 101 optical pickup 101a actuator 101b optical system 300 substrate surface close to the light emission detector 16

Claims (9)

For analysis that has a sample holder formed by a depression provided on the surface of the base substrate and a cover layer formed so as to cover the depression, and enables optical analysis of the sample held in the sample holder In the substrate,
An analysis substrate comprising antireflection means on the analysis substrate surface.   The analysis substrate according to claim 1, further comprising a reflective layer between the base substrate and the antireflection means.   The analysis substrate according to claim 1, wherein the antireflection means is a black carbon compound layer or a non-light emitting pigment-containing layer.   The analysis substrate according to claim 1 or 2, wherein the antireflection means is a fine concavo-convex structure or a fine porous structure formed on its surface.   The analysis substrate according to claim 1, wherein an opening is provided in the antireflection means located on the sample holding unit.   The analysis substrate according to claim 1, wherein the antireflection means is an antireflection film made of a multilayer film of inorganic oxide.   The analysis substrate according to claim 2, wherein the reflection layer is provided with guide means for guiding incident light for optical analysis. The analysis substrate according to any one of claims 1 to 7,
Light irradiation means for making incident light incident;
An analysis apparatus comprising: a light detection means disposed on the opposite side of the light irradiation means across the analysis substrate and disposed outside the optical path of the incident light. An optical pickup that makes incident light incident on the analysis substrate according to claim 7 and detects reflected light from the analysis substrate;
Drive means for moving the optical pickup in a direction along the guide means;
Control means for controlling the optical pickup so that the incident light follows the guiding means based on a detection signal output from the optical pickup, and
An analysis device comprising:

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