JP4741966B2 - Imaging device, biopolymer analysis chip, gene expression analysis method, and antigen detection method - Google Patents

Description

  The present invention relates to an imaging device, a biopolymer analysis chip, a gene expression analysis method, and an antigen detection method.

  In order to analyze the expression of genes of various biological species, biopolymer analysis chips such as DNA chips and antibody chips and readers thereof have been developed. A biopolymer analysis chip is obtained by aligning and fixing cDNA and antibodies having a known base sequence serving as a probe on a solid support such as a slide glass in a matrix (for example, see Patent Document 1). For example, gene expression analysis using a DNA chip and its reader is performed as follows.

First, a DNA chip is prepared in which a plurality of types of cDNAs having known base sequences (hereinafter referred to as probe DNA) are aligned and fixed on a solid support such as a slide glass. Next, mRNA is extracted from the specimen, cDNA is synthesized using reverse transcriptase, and prepared by labeling with a labeling substance (hereinafter referred to as labeled DNA). Here, as the labeling substance, a fluorescent substance, a chemiluminescent substrate, an enzyme that emits light from the chemiluminescent substrate, or the like can be used.
Next, when the labeled DNA is added onto the DNA chip, the labeled DNA is immobilized on the DNA chip by hybridizing with the complementary probe DNA.

Next, the DNA chip is set in a reader and analyzed by the reader. The reader condenses the light emitted by the labeling substance that scans the DNA chip with a condensing lens and a photomultiplier that moves two-dimensionally with respect to the DNA chip, and condenses the light intensity with the photomultiplier. By measuring the light intensity distribution in the surface of the DNA chip, the light intensity distribution on the DNA chip is output as a two-dimensional image. In the output image, the portion having a high light intensity indicates that a labeled DNA having a base sequence complementary to the base sequence of the probe DNA is included. Therefore, mRNA expressed in the specimen can be identified based on which part of the two-dimensional image has high fluorescence intensity.
JP 2000-1312237 A

  By the way, a biopolymer analysis chip has been developed in which probe molecules such as DNA and antibodies are spotted on a light receiving surface of a solid-state imaging device in which a plurality of photoelectric conversion elements are arranged in a two-dimensional array. In such a biopolymer analysis chip, light generated by a labeling substance that labels a biopolymer such as labeled DNA attached to a spot is measured by each photoelectric conversion element. Spots are scattered on the light receiving surface of the solid-state imaging device, and attenuation of light emitted from the labeling substance can be suppressed because the distance between the biopolymer attached to the light receiving surface and the photoelectric conversion element is short Therefore, there is an advantage that measurement is possible even with a small amount of light.

  In the biopolymer analysis chip as described above, when an enzyme for emitting a chemiluminescent substrate is used as a labeling substance, a solution containing the chemiluminescent substrate is dropped on the spot and excited based on a chemical reaction by the enzyme. The light emitted when the chemiluminescent substrate is returned to the ground state is measured. However, if the chemiluminescent substrate diffuses into the solution during the time until the chemiluminescent substrate excited with the labeling substance returns to the ground state, the signal amount by the photoelectric conversion element decreases, and the S / N ratio decreases.

  An object of the present invention is to solve the above problems and provide an imaging device, a biopolymer analysis chip, a gene expression analysis method, and an antigen detection method with better sensitivity.

In order to solve the above-described problems, the invention described in claim 1 is directed to a photoelectric conversion element, a fluorescent substance that is charged and has a chemical reaction based on an enzyme, or a chemiluminescence that has a charge and generates the phosphor. In order to attract the substrate electrically, an electrode provided on the light receiving surface side of the photoelectric conversion element, and a well provided on the opposite side of the photoelectric conversion element with respect to the electrode and into which a solution is injected. It is an imaging device characterized by comprising.

The invention according to claim 2 is a chemistry for generating a plurality of photoelectric conversion elements arranged in a two-dimensional array and a fluorescent substance having a charge and an enzyme-based chemical reaction, or for generating the fluorescent substance having a charge. In order to electrically attract the luminescent substrate, an electrode provided on the light receiving surface side of each photoelectric conversion element is provided on each side opposite to the photoelectric conversion element, and a solution is injected therein. An imaging device.

  The invention described in claim 3 is characterized in that a probe that binds to a specific biopolymer in the solution injected into the well is provided in the vicinity of the electrode of the imaging device according to claim 1 or 2. This is a biopolymer analysis chip.

  The invention according to claim 4 is the biopolymer analysis chip according to claim 3, wherein the probe is single-stranded DNA having a known base sequence.

  The invention according to claim 5 is the biopolymer analysis chip according to claim 3, wherein the probe is an antibody that binds to a specific antigen.

The invention described in claim 6 is a gene expression analysis method using the biopolymer analysis chip according to claim 4,
A solution containing enzyme-labeled DNA prepared using RNA extracted from any cell as a template is injected into the well,
Next, a part of the enzyme-labeled DNA in the solution is hybridized to the probe,
Thereafter, a solution containing a chemiluminescent substrate that generates a fluorescent substance in an excited state by a chemical reaction in which the enzyme that charges the solution and that labels the enzyme-labeled DNA functions as a catalyst is injected into the well,
Apply a voltage to the electrode to draw the fluorescent material or the chemiluminescent substrate to the electrode side,
Light emitted from the generated fluorescent material in the excited state is detected by the photoelectric conversion element.

The invention according to claim 7 is a method for detecting an antigen using the biopolymer analysis chip according to claim 5,
A solution containing any antigen is injected into the well,
Next, the specific antigen in the solution is bound to the probe,
Next, a solution containing a second antibody labeled with an enzyme and binding to a different antigenic determinant of the same antigen as the probe is injected into the well, and the second antibody is applied to the specific antigen bound to the probe. Binding and removing the second antibody that did not bind to the specific antigen,
Thereafter, a solution containing a chemiluminescent substrate that generates a fluorescent substance in an excited state by a chemical reaction in which an enzyme that charges the second antibody and labels the second antibody functions as a catalyst is injected into the well,
Apply a voltage to the electrode to draw the fluorescent material or the chemiluminescent substrate to the electrode side,
Light emitted from the generated fluorescent material in the excited state is detected by the photoelectric conversion element.

  According to the present invention, the concentration of the fluorescent substance or chemiluminescent substrate in the vicinity of the light receiving surface of the photoelectric conversion element is increased by electrically attracting the fluorescent substance or chemiluminescent substrate generated by the chemical reaction based on the enzyme in the well. It is possible to increase the reaction rate to generate more excited phosphors, or to localize the phosphors to collect the fluorescence, and photoelectrically emit light emitted from the excited phosphors. It can be efficiently captured by the conversion element. Therefore, the total amount of the chemiluminescent substrate to be used can be suppressed.

  The best mode for carrying out the present invention will be described below with reference to the drawings. However, although various technically preferable limitations for implementing the present invention are given to the embodiments described below, the scope of the invention is not limited to the following embodiments and illustrated examples.

<First embodiment>
[1] Overall Configuration of Biopolymer Analysis Chip FIG. 1 is a schematic plan view of the biopolymer analysis chip 1 in the first embodiment to which the present invention is applied, and FIG. FIG.

  The biopolymer analysis chip 1 includes an imaging device including a solid-state imaging device 10 and a grid-frame-shaped partition wall 50 provided on the solid-state imaging device 10 so as to surround the solid-state imaging device 10, and the partition wall 50 and the solid-state imaging device. 10, a plurality of spots 60, 60,... Scattered in each well 52 formed in a matrix. Although FIG. 1 shows a matrix of 2 rows × 2 columns, the present invention may have a matrix composed of more rows and columns.

[2] Solid-State Imaging Device Here, the solid-state imaging device 10 will be described with reference to FIGS. 1 and 2.
The transparent substrate 17 is a glass substrate such as quartz glass or a plastic substrate such as polycarbonate or PMMA, and has a property of transmitting light emitted from a phosphor to be described later (hereinafter referred to as light transmissive property) and insulation. .

  In the solid-state imaging device 10, a double gate type field effect transistor (hereinafter referred to as a double gate transistor) 20 is used as a photoelectric conversion element, and a plurality of double gate transistors 20, 20,. The double gate transistors 20, 20,... Are collectively covered with a protective insulating film 32 such as silicon nitride (SiN).

  FIG. 3 is a plan view showing the double gate transistor 20. As shown in FIG. 3, each of the double gate transistors 20, 20,... Has a semiconductor film 23 that is a light receiving portion, and a bottom gate electrode 21 that is formed below the semiconductor film 23 with the bottom gate insulating film 22 interposed therebetween. The top gate electrode 31 formed on the semiconductor film 23 with the top gate insulating film 29 interposed therebetween, the impurity semiconductor film 25 formed so as to overlap with part of the semiconductor film 23, and another portion of the semiconductor film 23 An impurity semiconductor film 26 formed so as to overlap, a source electrode 27 overlapped with the impurity semiconductor film 25, and a drain electrode 28 overlapped with the impurity semiconductor film 25, and having a level according to the amount of light received by the semiconductor film 23 It converts to an electrical signal.

  The bottom gate electrode 21 is formed on the transparent substrate 17 for each double gate transistor 20. Further, a plurality of bottom gate lines 41, 41,... Extending in the horizontal direction are formed on the transparent substrate 17, and the double gate transistors 20, 20,. Each bottom gate electrode 21 is formed integrally with a common bottom gate line 41. The bottom gate electrode 21 and the bottom gate line 41 have conductivity and light shielding properties, and are made of, for example, chromium, a chromium alloy, aluminum, an aluminum alloy, or an alloy thereof.

The bottom gate electrode 21 and the bottom gate lines 41, 41,... Of the double gate transistors 20, 20,. That is, the bottom gate insulating film 22 is a film formed in common to all the double gate transistors 20, 20,. The bottom gate insulating film 22 has insulating properties and light transmissive properties, and is made of, for example, silicon nitride (SiN) or silicon oxide (SiO ₂ ).

  A plurality of semiconductor films 23 are formed on the bottom gate insulating film 22 so as to be arranged in a matrix. The semiconductor film 23 is formed independently for each double gate transistor 20, and is disposed opposite to the bottom gate electrode 21 in each double gate transistor 20, and the bottom gate insulating film 22 is interposed between the bottom gate electrode 21. Is sandwiched. The semiconductor film 23 has a substantially rectangular shape in plan view, and is a layer formed of amorphous silicon or polysilicon that generates electron-hole pairs in an amount corresponding to the amount of received fluorescence.

  A channel protective film 24 is formed on the semiconductor film 23. The channel protective film 24 is patterned independently for each double gate transistor 20, and is formed on the central portion of the semiconductor film 23 in each double gate transistor 20. The channel protective film 24 has insulating properties and light transmissive properties, and is made of, for example, silicon nitride or silicon oxide. The channel protective film 24 protects the interface of the semiconductor film 23 from an etchant used for patterning. When light enters the semiconductor film 23, an amount of electron-hole pairs according to the amount of incident light is generated around the interface between the channel protective film 24 and the semiconductor film 23. In this case, holes are generated as carriers on the semiconductor film 23 side, and electrons are generated on the channel protective film 24 side.

An impurity semiconductor film 25 is partially formed on one end portion of the semiconductor film 23 so as to overlap the channel protection film 24, and an impurity semiconductor film 26 is partially formed on the other end portion of the semiconductor film 23. The channel protective film 24 is formed so as to overlap. The impurity semiconductor films 25 and 26 are patterned independently for each double gate transistor 20. The impurity semiconductor films 25 and 26 are made of amorphous silicon (n ⁺ silicon) containing n-type impurity ions.

  A source electrode 27 is formed on the impurity semiconductor film 25, and a drain electrode 28 is formed on the impurity semiconductor film 26. The source electrode 27 and the drain electrode 28 are formed for each double gate transistor 20. A plurality of source lines 42, 42,... And drain lines 43, 43,... Extending in the vertical direction are formed on the bottom gate insulating film 22. The source electrodes 27 of the double gate transistors 20, 20,... In the same column arranged in the vertical direction are formed integrally with the common source line 42, and the double gate transistors 20 in the same column arranged in the vertical direction. , 20,... Are formed integrally with a common drain line 43. The source electrode 27, the drain electrode 28, the source line 42, and the drain line 43 have conductivity and light shielding properties, and are made of, for example, chromium, a chromium alloy, aluminum, an aluminum alloy, or an alloy thereof.

  .. Of the double gate transistors 20, 20,... And the source lines 42, 42... And the drain lines 43, 43,. The top gate insulating film 29 is a film formed in common to all the double gate transistors 20, 20,. The top gate insulating film 29 has insulating properties and light transmissive properties, and is made of, for example, silicon nitride or silicon oxide.

  A plurality of top gate electrodes 31 are formed for each double gate transistor 20 on the top gate insulating film 29. The top gate electrode 31 is disposed opposite to the semiconductor film 23 in each double gate transistor 20, and the top gate insulating film 29 and the channel protective film 24 are sandwiched between the top gate electrode 31 and the semiconductor film 23. Further, a plurality of top gate lines 44, 44,... Extending in the horizontal direction are formed on the top gate insulating film 29, and the double gate transistors 20, 20,. The top gate electrode 31 is formed integrally with a common top gate line 44. The top gate electrode 31 and the top gate line 44 are conductive and light transmissive, for example, indium oxide, zinc oxide, tin oxide, or a mixture containing at least one of them (for example, tin-doped indium oxide ( ITO) and zinc-doped indium oxide).

  The top gate electrode 31 and the top gate lines 44, 44,... Of the double gate transistors 20, 20,... Are collectively covered by the protective insulating film 32, and the protective insulating film 32 is common to all the double gate transistors 20, 20,. It is the film | membrane formed in this way. The protective insulating film 32 has insulating properties and light transmittance, and is made of silicon nitride or silicon oxide.

  On the upper surface of the protective insulating film 32, an electrode 33 is provided above the semiconductor film 23 of the double gate transistors 20, 20,. Further, on the upper surface of the protective insulating film 32, a plurality of electrode lines 34, 34,... Extending in the vertical direction are formed on the sides of the double gate transistors 20, 20,. The electrodes 33 and 33 in the same row are formed integrally with a common electrode line 34. The electrode 33 and the electrode line 34 are conductive, and a transparent electrode exhibiting high transparency with respect to the fluorescence of the phosphor 82 described later is preferable. For example, indium oxide, zinc oxide, tin oxide, or any of these It is formed of a mixture containing at least one (for example, tin-doped indium oxide (ITO), zinc-doped indium oxide).

  The electrode 33 and the electrode line 34 are collectively covered with a protective insulating film 35. The protective insulating film 35 has insulating properties and light transmittance, and is made of silicon nitride or silicon oxide.

  The solid-state imaging device 10 configured as described above has the surface of the protective insulating film 35 as a light receiving surface, and is provided so as to convert the amount of light received by the semiconductor film 23 of each double gate transistor 20 into an electrical signal. Yes.

[3] Partition Wall The partition wall 50 is in close contact with the protective insulating film 35, and a well 52 is formed above each of the double gate transistors 20, 20,. Note that two or more double gate transistors 20 may be arranged in one well 52. The partition wall 50 is opaque and prevents light from entering from the well 52 adjacent to the double gate transistor 20.

[4] Spot Next, the spot 60 will be described. As shown in FIG. 1, each well 52 has a spot formed on the upper surface of the solid-state imaging device 10. As shown in FIG. 2, each spot 60 is formed by dropping a solution of cDNA (probe DNA 61) having a known base sequence serving as a probe or an antibody and drying it. Hereinafter, a case where cDNA having a known base sequence is used as a probe will be described.

  In one spot 60, a crowd of a large number of single-stranded probe DNAs 61 having the same base sequence is immobilized on the upper surface of the solid-state imaging device 10, and the probe DNA 61 has a different base sequence for each spot 60. As the probe DNA 61, a DNA having a nucleotide sequence identical to or complementary to a known mRNA nucleotide sequence or a part thereof is used. Specifically, for example, a cDNA library prepared from the same cell specimen as used for enzyme-labeled DNA described later can be used.

  One spot 60 is formed so as to overlap the double gate transistor 20 in each well 52. Note that the number of double gate transistors 20 overlapping one spot 60 may be different.

[5] Analyzer The biopolymer analysis chip 1 is set in the analyzer 70 and the biopolymer analysis chip 1 is used. First, the analyzer 70 will be described with reference to FIG. FIG. 4 is a block diagram showing the configuration of the analyzer 70.

  The analysis apparatus 70 is connected to the biopolymer analysis chip 1, and includes an electrode driver 73, a top gate driver 74, a bottom gate driver 75, and a drain driver 76 that drive the solid-state imaging device 10, a computer 71 that controls these, and a computer. And an output device 77 that performs output (display or print) using the signal output from 71. The output device 77 is a plotter, a printer, or a display.

  When the biopolymer analysis chip 1 is set in the analyzer 70, the electrode line 34 of the solid-state imaging device 10 is connected to the terminal of the electrode driver 73. .. Are the terminals of the top gate driver 74, the bottom gate lines 41, 41,... Are the terminals of the bottom gate driver 75, and the drain lines 43, 43,. Are connected to each other. When the biopolymer analysis chip 1 is set in the analyzer 70, the source lines 42, 42,... Of the solid-state imaging device 10 are connected to a constant voltage source, and in this example, the source lines 42, 42,. It has come to be.

  The electrode driver 73, the top gate driver 74, the bottom gate driver 75, and the drain driver 76 cooperate to drive the solid-state imaging device 10.

  The computer 71 includes a CPU, a RAM, a ROM, and the like (not shown), and outputs control signals to the electrode driver 73, the top gate driver 74, the bottom gate driver 75, and the drain driver 76, whereby the electrode driver 73, the top gate driver 74, The bottom gate driver 75 and the drain driver 76 have a function of driving the solid-state imaging device 10. The computer 71 has a function of causing the output device 77 to output an image according to the input two-dimensional image data.

  Further, the computer 71 has a function of acquiring the light intensity distribution along the light receiving surface of the solid-state imaging device 10 as two-dimensional image data by A / D converting the electric signal input from the drain driver 76.

  Further, the analyzer 70 includes an injection device 79 controlled by the computer 71. The computer 71 drives the injection device 79 and drops a solution containing a chemiluminescent substrate, which will be described later, into each well 52 of the biopolymer analysis chip 1 set in the analysis device 70.

[6] Enzyme labeled DNA
As a sample to be analyzed by the biopolymer analysis chip 1, DNA can be used. As the sample DNA, cDNA obtained by extracting mRNA expressed in an arbitrary cell specimen and using reverse transcriptase can be used. The cDNA is labeled with an enzyme that functions as a catalyst for the reaction of a chemiluminescent substrate described later, such as alkaline phosphatase and peroxidase.

  In order to label the cDNA with an enzyme, for example, an RT-PCR reaction may be carried out using an oligo dT primer labeled with the enzyme or a labeled dNTP mix. Hereinafter, this labeled cDNA is referred to as enzyme-labeled DNA.

[7] Chemiluminescent substrate The chemiluminescent substrate used for detecting the enzyme-labeled DNA will be described. As the chemiluminescent substrate, one that generates a fluorescent substance in an excited state by a chemical reaction using an enzyme used for labeling enzyme-labeled DNA as a catalyst can be used.
Specifically, for example, a dioxetane derivative serving as a substrate for alkaline phosphatase or a luminol compound serving as a substrate for peroxidase can be used.

As an example of the dioxetane derivative, an adamantyl dioxetane chlorinated derivative (C ₁₈ H ₁₉ Cl ₂ O ₇ PNa ₂ ) represented by Chemical Formula 1 will be described as an example. The adamantyl dioxetane chlorinated derivative has a phosphate group and is a substrate for alkaline phosphatase.

Alkaline phosphatase hydrolyzes the phosphate group of an adamantyl dioxetane chlorinated derivative to form an unstable intermediate shown in Formula 2.

The intermediate is spontaneously cleaved and decomposed into adamantanone and phosphor shown in Chemical Formula 3. This phosphor is in an excited state, and energy when the phosphor transitions to the ground state is emitted as light.

  In addition, the chemiluminescent substrate used as the substrate of alkaline phosphatase has a phosphate group and is negatively charged in an aqueous solution. In addition, the phosphor produced by hydrolysis of the chemiluminescent substrate is also negatively charged in the aqueous solution. For this reason, the chemiluminescent substrate and the phosphor move to the positive charge side in the aqueous solution.

[8] Sample Processing A method for processing enzyme-labeled DNA will be described. First, an operator injects a solution containing enzyme-labeled DNA (hereinafter referred to as an enzyme-labeled DNA solution) into each well 52. Before injecting the enzyme-labeled DNA solution, a buffer solution for allowing the enzyme-labeled DNA to migrate may be placed in each well 52, or the enzyme-labeled DNA solution itself may also serve as a buffer solution for electrophoresis. Note that the enzyme-labeled DNA solution may be dropped sequentially or simultaneously on the spots 60, 60,. At this time, the enzyme-labeled DNA solution is heated so that the enzyme-labeled DNA becomes a single strand.

Next, each well 52 of the biopolymer analysis chip 1 is cooled to a predetermined temperature so that the probe DNA 61 and the enzyme-labeled DNA cause hybridization. Then, of the enzyme-labeled DNA in the enzyme-labeled DNA solution injected into each well 52, the DNA complementary to the probe DNA 61 of the spot 60 in the well hybridizes with the probe DNA 61. On the other hand, enzyme-labeled DNA that is not complementary to the probe DNA 61 does not bind to the spot 60.
Thereafter, the enzyme-labeled DNA solution in the well 52 is washed away with a washing buffer solution, and the enzyme-labeled DNA that has not hybridized with the probe DNA 61 is removed from the well 52.

[9] Detection of Sample A method for detecting enzyme-labeled DNA will be described with reference to FIGS.
The solid-state imaging device 10 that has performed the above processing is set in the analyzer 70, the electrode driver 73, the top gate driver 74, the bottom gate driver 75, and the drain driver 76 are connected to the computer 71, and the computer 71 is activated.

Next, the injection device 79 is controlled by the computer 71, and a solution containing the chemiluminescent substrate 81 is injected into each well 52 (FIG. 5). Before injecting the solution containing the chemiluminescent substrate 81, a buffer solution for the chemiluminescent substrate 81 to migrate may be placed in each well 52, and the solution containing the chemiluminescent substrate 81 itself is a buffer solution for electrophoresis. You may also serve.
Next, the computer 71 drives the electrode driver 73 to charge the electrode 33 to a positive charge. Then, the chemiluminescent substrate 81 in the well 52 descends toward the positively charged electrode 33 in the aqueous solution (FIG. 6). For this reason, in the vicinity of the spot 60 on the electrode 33, the concentration of the chemiluminescent substrate 81 is increased, the reaction rate can be increased, and more phosphors 82 can be generated. Therefore, the total amount of the chemiluminescent substrate 81 to be used can be suppressed.

  In the well 52 in which the enzyme-labeled DNA is hybridized to the probe DNA 61 of the spot 60, the chemiluminescent substrate is hydrolyzed by the enzyme 63 that labels the enzyme-labeled DNA 62, and an excited phosphor is generated via an unstable intermediate. . When the excited state phosphor transitions to the ground state, fluorescence (mainly in the visible light wavelength region) is emitted, and the fluorescence enters the double-gate transistor 20 corresponding to the spot 60 that emits fluorescence, so that an electron-hole pair is formed. appear.

Since the phosphor is also electrically negatively charged, the generated phosphor moves to the electrode 33 side to which a positive voltage is applied, that is, the solid-state imaging device 10 side, and does not diffuse throughout the well 52. (FIG. 7). For this reason, more fluorescence enters the double gate transistor 20 and more electron-hole pairs are generated. Therefore, the signal becomes larger than the background noise, and the SN ratio can be improved.
Thereafter, the respective light quantity data of each of the double gate transistors 20, 20,... Is acquired and stored in the RAM.

  The operator performs hybridization of each spot 60, 60,... From image data obtained by outputting the light quantity data of each double gate transistor 20, 20,. The presence or absence can be confirmed. If hybridization has occurred, it can be seen that mRNA having a base sequence complementary to the probe DNA 61 in the spot 60 is expressed in the cell sample.

Second Embodiment
In the first embodiment, DNA synthesized by reverse transcriptase from mRNA expressed in a sample is used as a sample to be analyzed by the biopolymer analysis chip 1. An antigen such as a sugar chain may be used as a sample. In this case, an antibody that binds to an antigen such as a known protein or sugar chain (hereinafter referred to as a probe antibody) is used as a probe.

  Specifically, a solution containing a probe antibody is dropped into each well 52 of the biopolymer analysis chip 1 and dried to form a spot 60. The probe antibodies dropped into each well 52 have different proteins as antigens, and the antibodies forming the same spot 60 recognize the same antigenic determinant. A monoclonal antibody can be used as such a probe antibody.

Next, a solution containing an antigen serving as a sample (hereinafter referred to as a sample solution) is injected into each well 52.
After a sufficient time has passed for the antigen in the sample solution to bind to the probe antibody, the sample solution in the well 52 is washed away with the buffer solution, and the sample solution that has not bound to the probe antibody is removed from the well 52. Remove.

Next, in each well 52, a solution (hereinafter referred to as an enzyme-labeled antibody solution) of an antibody labeled with an enzyme that recognizes a different antigenic determinant of the same antigen as that recognized by the probe antibody (hereinafter referred to as an enzyme-labeled antibody solution). ).
After sufficient time for the antigen bound to the probe antibody to bind to the enzyme-labeled antibody, the enzyme-labeled antibody solution in the well 52 was washed away with a buffer solution, and the enzyme-labeled antibody solution did not bind to the antigen. Things are removed from within the well 52.
Thereafter, similarly to [9] sample detection of the first embodiment, a solution containing a chemiluminescent substrate is injected into each well 52, and the light quantity data is measured by the analyzer 70.

  As shown in FIG. 8, in the well 52 in which the antigen 65 is bound to the probe antibody 64 and the enzyme-labeled antibody 66 is bound to the antigen 65, the chemiluminescent substrate 81 is hydrolyzed by the enzyme 67 of the enzyme-labeled antibody 66, and the intermediate Then, an excited phosphor 82 is generated. The fluorescence emitted from the phosphor 82 is detected by the double gate transistor 20.

  The operator uses the output device 77 to output the light quantity data of each of the double gate transistors 20, 20,... Stored in the RAM and outputs the antigen 65 at each spot 60, 60,. The presence or absence can be confirmed. Therefore, it is possible to directly confirm what kind of antigen is expressed in the specimen according to the type of antibody in the well 52.

  The present invention is not limited to the above embodiment, and various improvements and design changes may be made without departing from the spirit of the present invention.

  For example, in the above embodiment, a single-stranded DNA or antibody having a known base sequence is used as a probe, but other known biopolymers or low molecules may be used. For example, a peptide or protein serving as an antigen, a sugar chain, a low molecular ligand, a known cell, or the like may be used.

1 is a schematic plan view of a biopolymer analysis chip 1 in a first embodiment to which the present invention is applied. It is arrow sectional drawing along the cut surface II of FIG. 2 is a plan view showing an electrode structure of a photoelectric conversion element that is a pixel of the solid-state imaging device 10. FIG. 3 is a block diagram illustrating a configuration of an analysis device 70. FIG. It is explanatory drawing about the detection method of enzyme labeled DNA. It is explanatory drawing about the detection method of enzyme labeled DNA. It is explanatory drawing about the detection method of enzyme labeled DNA. It is explanatory drawing about the detection method of an antigen.

Explanation of symbols

1 Biopolymer Analysis Chip 10 Solid-State Imaging Device 20 Double Gate Transistor (Photoelectric Conversion Element)
33 Electrode 52 Well 60 Spot 61 Probe DNA
62 Enzyme labeled DNA
63, 67 Enzyme 64 Probe antibody 65 Antigen 66 Enzyme labeled antibody 81 Chemiluminescent substrate 82 Fluorescent substance

Claims (7)

A photoelectric conversion element;
To draw a fluorescent substance caused by a chemical reaction based on the charged and enzyme charge, or a charged and chemiluminescent substrate to produce the phosphor charges electrically an electrode provided on the light-receiving surface side of the photoelectric conversion element,
An imaging apparatus comprising: a well provided on a side opposite to the photoelectric conversion element with respect to the electrode, into which a solution is injected. A plurality of photoelectric conversion elements arranged in a two-dimensional array;
Phosphor caused by a chemical reaction based on the charged and enzyme charge or charged and to attract electrically chemiluminescent substrate to produce the phosphor charges, the respectively provided on the light-receiving surface of each photoelectric conversion element electrodes, When,
An imaging device comprising: a well provided on a side opposite to the photoelectric conversion element with respect to each electrode, and a well into which a solution is injected.   A biopolymer analysis chip comprising a probe that binds to a specific biopolymer in a solution injected into the well in the vicinity of the electrode of the imaging device according to claim 1.   The biopolymer analysis chip according to claim 3, wherein the probe is a single-stranded DNA having a known base sequence.   The biopolymer analysis chip according to claim 3, wherein the probe is an antibody that binds to a specific antigen. A gene expression analysis method using the biopolymer analysis chip according to claim 4,
A solution containing enzyme-labeled DNA prepared using RNA extracted from any cell as a template is injected into the well,
Next, a part of the enzyme-labeled DNA in the solution is hybridized to the probe,
Thereafter, a solution containing a chemiluminescent substrate that generates a fluorescent substance in an excited state by a chemical reaction in which the enzyme that charges the solution and that labels the enzyme-labeled DNA functions as a catalyst is injected into the well,
Applying voltage to the electrode to draw the fluorescent material or the chemiluminescent substrate to the electrode side,
A gene expression analysis method, characterized in that light emitted from the generated excited fluorescent substance is detected by the photoelectric conversion element. An antigen detection method using the biopolymer analysis chip according to claim 5,
A solution containing any antigen is injected into the well,
Next, the specific antigen in the solution is bound to the probe,
Next, a solution containing a second antibody labeled with an enzyme and binding to a different antigenic determinant of the same antigen as the probe is injected into the well, and the second antibody is applied to the specific antigen bound to the probe. Binding and removing the second antibody that did not bind to the specific antigen,
Thereafter, a solution containing a chemiluminescent substrate that generates a fluorescent substance in an excited state by a chemical reaction in which an enzyme that charges the second antibody and labels the second antibody functions as a catalyst is injected into the well,
Applying voltage to the electrode to draw the fluorescent material or the chemiluminescent substrate to the electrode side,
A method for detecting an antigen, wherein the photoelectric conversion element detects light emitted from a generated fluorescent substance in an excited state.

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