JP2007010325A - Physiological molecule detecting method and optical reading device - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a physiological molecule detecting method which enables the simple identification of a specific spot from an optical image, collation of a numerical value with a spot image and rapid execution of diagnosis based on an examination result. <P>SOLUTION: The spots 5 fixed on a target physiological molecule detecting chip 1 are arranged in an imaging area 2 on the basis of a preferential order. The imaging area 2 is photographed by one photographing using an imaging means 6 while the detection result of a target physiological molecule is identified from the position of the positive spot 7 of the optical image 3 displayed on an optical image monitor 31 to simply perform rapid diagnosis. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention provides a target living body for detecting a plurality of types of target biomolecules such as target nucleic acid molecules and peptides contained in a biological sample using a DNA microarray or peptide array in which probes that bind to DNA or peptides are immobilized on a chip. The present invention relates to a molecular detection method and an optical reader.

  Nucleic acid molecules such as DNA and RNA have been widely used as molecular markers in biological samples, as well as specific enzyme activities and antigen epitopes and antibodies. By binding a molecular marker such as an antigen or nucleic acid molecule to a known antibody molecule or nucleic acid molecule, the molecular marker in the biological sample can be detected, quantified and used for diagnosis.

  A plurality of molecular markers can be detected simultaneously by using a detection chip in which a probe that binds to a molecular marker is used as a spot and arranged in a matrix on the substrate surface and reacting a biological sample on the detection chip.

  In particular, the use of DNA microarrays in which probe nucleic acids that hybridize with target nucleic acid molecules are formed on a variety of matrices has become active.

  Thousands to tens of thousands of probes are immobilized as spots on the DNA microarray in a matrix, and multiple types of nucleic acid molecules can be detected simultaneously by optically detecting the nucleic acids bound to each probe molecule. I can do things.

  DNA probes immobilized on DNA microarrays can be obtained by photopolymerizing one base at a time on a substrate using semiconductor photolithographic technology, or by dissolving DNA molecules synthesized in advance to a length of several tens to several hundred bases. A method is adopted in which the droplets are adsorbed and fixed by spotting on the substrate.

  As a method for detecting a nucleic acid molecule, a probe nucleic acid that forms a complementary base pair with a target nucleic acid molecule is immobilized on a substrate surface, and a target nucleic acid molecule previously labeled with a label molecule such as a fluorescent molecule or RI is hybridized with a probe, A method of detecting whether or not a labeled molecule is bound to the substrate surface with a fluorescence detector or the like is used.

The target nucleic acid molecule can be labeled with a labeled molecule by directly labeling a fluorescent molecule using an enzymatic reaction or the like on a DNA molecule extracted from a biological sample, or by synthesizing cDNA from the extracted RNA and amplifying it by PCR. In this case, a method of incorporating fluorescent molecules is used.
A method for detecting a target nucleic acid molecule at high speed and with high sensitivity using such a DNA microarray has been developed (see Patent Document 1).

  Hereinafter, such a hybridization reaction detection method and apparatus will be described with reference to FIG.

As shown in FIG. 16, beams emitted from a plurality of laser light sources 101, 102, 103... Enter a multi-beam splitter 104, are divided into two, pass through a multi-lens 105, pass through a multi-prism 106, and enter a multi-pinhole 107. Is incident on. The multi-beam 108 emitted from the multi-pinhole 107 irradiates the DNA chip 109, and the fluorescence on the DNA chip 109 is transmitted through the objective lens 110, and the light of the fluorescence wavelength is reflected by the wavelength selection beam splitter 111. Detected by. By relatively moving the multi-beam 108 and the DNA chip 109 and constructing a fluorescence image from the obtained data, the fluorescence image of the DNA chip can be detected with high speed and high sensitivity.
JP 2002-55050 A

  In such a conventional detection method, in order to determine the inspection result from the obtained optical image, the numerical value obtained by analyzing the optical image must be compared with the spot of the probe, and the number of spots is reduced. In many cases, collation becomes complicated, and it is difficult to identify a specific spot from the constructed optical image, and it is difficult to compare the numerical value with the actual optical image. There is a problem that it takes time to perform the detection, and there is a demand for a detection method in which a specific spot can be easily identified from an optical image, a numerical value and a spot image can be easily collated, and diagnosis based on a test result can be performed quickly.

  The present invention solves such a conventional problem. A specific spot can be easily identified from an optical image, a numerical value and a spot image can be easily collated, and diagnosis based on a test result can be quickly performed. It aims at providing the biomolecule detection method which can be performed easily.

  In addition, the apparatus used for such a conventional detection method is large-sized and has a problem that it is limited to a place having a large space as an installation place, and is small and easy to carry. An easy-to-use detection device is required.

  An object of the present invention is to solve such a conventional problem, and to provide a detection device that is small in size and can be easily carried and is easy for a tester to use for diagnosis.

  In order to achieve the above object, the biomolecule detection method of the present invention is a target biomolecule detection method using a target biomolecule detection chip on which one or more types of probes that bind to the target biomolecule are immobilized, The probe is immobilized as one or more types of independent spots on the target biomolecule detection chip, and the spots are arranged on the target biomolecule detection chip based on priority order. The above spot is arranged in an imaging area of the target biomolecule detection chip, and the biomolecule is detected by optically imaging the imaging area at one time. It is what.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  The biomolecule detection method of the present invention is the biomolecule detection method according to claim 1, wherein the spots are circular in order to achieve the above object.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  3. The biomolecule detection method according to claim 1, wherein a target biomolecule detection chip having a circular imaging area is used to achieve the above object. It is a thing.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  The biomolecule detection method of the present invention uses a target biomolecule detection chip having a donut-shaped imaging area in which no spot is arranged at the center in order to achieve the above object. The biomolecule detection method according to any one of the above is used.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  5. The biomolecule detection method of the present invention is characterized in that spots are arranged so that priorities are arranged from the center of the imaging area toward the circumference in order to achieve the above object. The biomolecule detection method described in 1. is used.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  The biomolecule detection method of the present invention uses a target biomolecule detection chip comprising a flat substrate whose imaging area has a smooth surface in order to achieve the above object. This is a biomolecule detection method described in the above.

  By this means, an optical image can be obtained with high accuracy, a specific spot can be easily identified from the optical image, a numerical value and a spot image can be easily collated, and a diagnosis can be quickly made based on a test result. This is a molecular detection method.

  In the biomolecule detection method of the present invention, in order to achieve the above object, the imaging area is constituted by a membrane, and the imaging area is pressed against a smooth reference surface having a smooth surface to perform imaging. The biomolecule detection method according to any one of Items 1 to 5.

  By this means, an optical image can be obtained with high accuracy, a specific spot can be easily identified from the optical image, a numerical value and a spot image can be easily collated, and a diagnosis can be quickly made based on a test result. This is a molecular detection method.

  The biomolecule detection method according to the present invention is characterized in that, in order to achieve the above object, the priorities are ranked according to the progress of the target diagnosis result. This is a biomolecule detection method.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  3. The biomolecule detection method according to claim 1, wherein the biomolecule detection method of the present invention uses a target biomolecule detection chip having a square imaging area in order to achieve the above object. It is a thing.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  10. The biomolecule detection method according to claim 9, wherein the biomolecule detection method of the present invention has spots arranged so that the priorities are arranged along one side of the imaging area in order to achieve the above object. This is a detection method.

  By this means, a specific spot can be easily identified from the optical image, the collation of the numerical value and the spot image can be simplified, and a biomolecule detection method capable of promptly diagnosing based on the test result.

  The optical reading device of the present invention is an optical reading device that images a target biomolecule detection chip in which one or more types of probes that bind to a target biomolecule are immobilized as one or more types of independent spots in order to achieve a superior purpose. A target biomolecule detection chip immobilization means for immobilizing the target biomolecule detection chip, and one or a plurality of light sources for irradiating excitation light in a predetermined wavelength range on the biomolecule detection chip And a light receiving means that is a photoelectric conversion element that receives light in a predetermined wavelength range emitted by the excitation light on the target biomolecule detection chip, and according to a priority order based on the importance of the target biomolecule An optical reading device characterized in that one or more types of the spots arranged in the imaging area are optically captured by one imaging. It is.

  By this means, the detection device is small and easy to carry, and is easy to use for diagnosis by the tester.

  An optical reading apparatus according to the present invention includes an optical image monitor that displays an optical image picked up to achieve the above object, and displays the optical image picked up by one photographing on a single screen. The optical reading device according to claim 11 is obtained.

  By this means, the detection device is small and easy to carry, and is easy to use for diagnosis by the tester.

  The optical reading device according to any one of claims 11 and 12, further comprising a condensing lens for condensing light emitted on the target biomolecule detection chip in order to achieve the above object. It is what.

  By this means, the detection device is small and easy to carry, and is easy to use for diagnosis by the tester.

  The optical reading device of the present invention is the optical reading device according to any one of claims 11 to 13, wherein the light receiving means is constituted by a two-dimensional array in order to achieve the above object. .

  By this means, the detection device is small and easy to carry, and is easy to use for diagnosis by the tester.

  The optical reader according to the present invention is an optical reader according to any one of claims 11 to 14, wherein the light source is an LED in order to achieve the above object.

  By this means, the detection device is small and easy to carry, and is easy to use for diagnosis by the tester.

  According to the present invention, a biomolecule detection method can easily identify a specific spot from an optical image, can easily collate a numerical value with a fluorescent image of the spot, and can quickly make a diagnosis based on a test result. Can provide.

  In addition, it is possible to provide a detection device that is small in size and can be easily carried, and that is easy for an examination performer to use for diagnosis.

  The invention according to claim 1 of the present invention is a target biomolecule detection method using a target biomolecule detection chip on which one or more types of probes that bind to a target biomolecule are immobilized, wherein the probe is the target biomolecule. Each of the spots is immobilized on the molecule detection chip as one or more independent spots, the spots are arranged on the target biomolecule detection chip based on priority, and the one or more spots are It is arranged in the imaging area of the target biomolecule detection chip, and is a biomolecule detection method characterized by detecting the biomolecule by optically imaging the imaging area at one time, The target biomolecule is reacted with an immobilized probe limited to the imaging area on the target biomolecule detection chip, the target biomolecule is bound in the imaging area, By optically capturing the combined target biomolecules in a single image, multiple types of target biomolecules can be detected rapidly at the same time, and spots are arranged in the imaging area based on priority. Therefore, the spot can be easily identified visually by checking the optical image obtained by photographing, and the inspection result can be quickly judged based on the arranged priority. It has the effect | action used as the biomolecule detection method which can perform a diagnosis rapidly.

  The invention according to claim 2 of the present invention is the biomolecule detection method according to claim 1, wherein the spots are circular, and the circular spots are arranged in the imaging area. As a result, a large number of spots can be efficiently formed in the imaging area, and the target biomolecules bound in the imaging area are imaged by one imaging so that the target biomolecules can be detected simultaneously. Since the spots are arranged in the imaging area based on the priority order, the spot can be easily identified visually by confirming the optical image obtained by photographing, and the spots are arranged in the priority order. Since a test result can be quickly determined based on this, it has an effect of a biomolecule detection method capable of performing a diagnosis quickly.

  The invention according to claim 3 of the present invention is the biomolecule detection method according to any one of claims 1 and 2, characterized in that a target biomolecule detection chip whose imaging area is circular is used. When the imaging area is optically imaged, the lens can be used to efficiently collect light from the circular imaging area, so the sensitivity is increased, and the target biomolecule bound in the imaging area is imaged by one imaging. Therefore, target biomolecules can be detected at the same time, and the spots are arranged in the imaging area based on the priority order. Therefore, it is easy to visually recognize the spot by checking the optical image obtained by photographing. The biomolecule detection method can perform diagnosis quickly because the identification of the test results can be performed and the test result can be quickly determined based on the arranged priority. It has the effect of becoming.

  The invention according to claim 4 of the present invention uses a target biomolecule detection chip in which an imaging area where no spot is arranged at the center is a donut shape. This is a biomolecule detection method. When optically imaging an imaging area, a spot is placed in the center even if a lens with a different light collection rate is used in the vicinity of the center of the imaging area. Since spots are not captured, it is possible to detect spots with high accuracy by photographing areas where the light collection rate is stable. This is because spots are arranged in the imaging area based on priority. Because it is possible to easily identify spots visually by checking the optical images obtained by photographing, and to quickly determine the inspection results based on the priorities arranged. An effect that the biomolecules detection method can be carried out quickly diagnose.

  The invention according to claim 5 of the present invention is characterized in that the spots are arranged so that the priorities are arranged from the center of the imaging area toward the circumferential portion. Since the positional relationship of the peripheral part with respect to the central part can be easily determined visually, the spot can be easily identified by checking the optical image obtained by photographing, and Since the test result can be quickly determined based on the arranged priority order, the biomolecule detection method can be performed more quickly.

  The invention according to claim 6 of the present invention uses the target biomolecule detection chip formed of a flat substrate whose imaging area has a smooth surface. This is a molecular detection method, and since the imaging area is a flat plate, it is possible to shoot without any pintomer when optically imaging the imaging area, so that a spot image in the imaging area can be obtained with high accuracy. The biomolecule detection method can be performed.

  The invention according to claim 7 of the present invention is characterized in that the imaging area is composed of a membrane, and the imaging area is pressed against a smooth reference surface having a smooth surface to capture an image. This is a biomolecule detection method according to any one of the membranes, wherein the surface with the spot of the imaging area can be easily flattened by pressing the smooth reference surface from the back side of the imaging area of the membrane. When the imaging area is optically photographed, it can be easily photographed without a pintomer, so that the biomolecule detection method can obtain a spot image in the imaging area with high accuracy.

  The invention according to claim 8 of the present invention is characterized in that the priority is ranked according to the degree of progress of the target diagnosis result, and the biomolecule detection method according to any one of claims 1 to 7 In the captured optical image, spots are arranged according to the degree of progress of the target diagnosis result, so it is easy to see the position of the spot where the target biomolecule is bound. The degree of progress can be determined, and the biomolecule detection method is capable of quickly determining the test result and diagnosing it as the degree of progress.

  The invention according to claim 9 of the present invention is the biomolecule detection method according to claim 1, characterized in that a target biomolecule detection chip having a square imaging area is used. When a rectangular light receiving element such as a CCD camera is used for shooting, a spot can be efficiently arranged in the imaging area, so that target biomolecules can be detected simultaneously, and the spot is captured in the imaging area. Since it is arranged in the order of priority, it is possible to easily identify the spot visually by checking the optical image obtained by photographing, and the inspection result can be quickly based on the arranged priority. Therefore, it has an effect of a biomolecule detection method capable of quickly making a diagnosis.

  The invention according to claim 10 of the present invention is the biomolecule detection method according to claim 9, wherein the spots are arranged so that the priorities are arranged along one side of the imaging area. Because the spots are arranged linearly along the sides of the square, the spots can be easily identified visually, and the test results can be quickly judged based on the priorities arranged. Therefore, the biomolecule detection method can perform diagnosis more quickly.

  The invention according to claim 11 of the present invention is an optical reader for imaging a target biomolecule detection chip in which one or more types of probes that bind to a target biomolecule are immobilized as one or more types of independent spots. A target biomolecule detection chip immobilization means for immobilizing the target biomolecule detection chip, one or a plurality of light sources for irradiating excitation light in a predetermined wavelength range on the biomolecule detection chip, and the target A light receiving means that is a photoelectric conversion element that receives light in a predetermined wavelength range that is emitted by the excitation light on the biomolecule detection chip, and is arranged in the imaging area according to the priority order based on the importance of the target biomolecule An optical reading device characterized in that one or more kinds of the spots are optically captured by one imaging, and the target biological component The target biomolecule detection chip that is bound to the probe is fixed to the target biomolecule detection chip fixing means, and the target biomolecule detection chip emits light by irradiating the target biomolecule detection chip with the excitation light from the light source. In addition, it is possible to quickly detect a plurality of types of target biomolecules at the same time by imaging the light emission of the imaging area by a single photographing with the photoelectric conversion element, and the spots are based on the priority order in the imaging area. Therefore, the spot can be easily identified visually by checking the optical image obtained by photographing, and the inspection result can be quickly judged based on the priority of arrangement. Therefore, the diagnosis can be performed quickly, and it is small and easy to carry. To.

  According to a twelfth aspect of the present invention, an optical image monitor that displays a captured optical image is provided, and the optical image captured by one photographing is displayed on a single screen. The optical reading device is described. By projecting the captured optical image on the optical image monitor, the spot image in the imaging area is displayed in the optical image monitor, and the spot image can be easily identified from the optical image monitor. It is possible to quickly determine the inspection result based on the priority order, so that the diagnosis can be performed quickly, and it is small and easy to carry. It has the action.

  The invention according to claim 13 of the present invention is the optical reading device according to claim 11, further comprising a condensing lens for condensing light emitted on the target biomolecule detection chip. Yes, because the light in the imaging area is collected by the condenser lens, the spot can be detected with high sensitivity, the target biomolecule can be detected with higher sensitivity, and it is small and easy to carry. Has an effect of becoming an optical reading device that is easy to use for diagnosis.

  The invention according to claim 14 of the present invention is the optical reading device according to any one of claims 11 to 13, characterized in that the light receiving means is constituted by a two-dimensional array. A plurality of types of target biomolecules can be detected at the same time by the light receiving means configured as described above, and the optical reading device is small and easy to carry.

The invention according to claim 15 of the present invention is the optical reading device according to any one of claims 11 to 14, characterized in that the light source is an LED, and the target biomolecule detection chip is lowered by the LED. Because it can irradiate with energy and suppress the discoloration caused by excitation light irradiation, the imaging area can be imaged, so that target biomolecules can be detected stably and reproducibly, and it is small and easy to carry. It has an effect of becoming an easy-to-use optical reader.
Hereinafter, embodiments of the present invention will be described with reference to the drawings.

(Embodiment 1)
FIG. 1 is a diagram showing a biomolecule detection method described in Embodiment 1 of the present invention. FIG. 1 (A) shows a target biomolecule detection chip 1 used for the biomolecule detection method, FIG. 1 (B) shows an imaging area 2, and FIG. 1 (C) is for detecting the target biomolecule. 1 (D) shows a biomolecule detection method using the target biomolecule detection chip 1, and FIG. 1 (E) shows an optical image 3 obtained by photographing the imaging area 2. .

  As shown in FIG. 1A, probes 4 that bind to the target biomolecule are immobilized on the target biomolecule detection chip 1 in the imaging area 2 as independent spots 5. For example, when DNA is detected as the target biomolecule, a nucleic acid containing a complementary base sequence that hybridizes with the target DNA is used as the probe 4. For example, a synthesized DNA molecule is used as the probe 4 and has a base sequence complementary to DNA or RNA derived from an organism in order to detect genetic material derived from a specific organism. In order to increase the properties, DNA molecules having a length of 20 to several hundred bases are used. For example, in order to detect a specific bacterium, the probe 4 can be detected by including a base sequence complementary to the bacterium genome based on the bacterium genome information. For example, tens to hundreds of nucleotide sequences including ITS on the genome of Dehalococides ethenegenes 195R are used as probe 4 to detect Deococcoides ethenegenes 195R, which is a VOC-degrading bacterium. In addition, in order to simultaneously detect a plurality of types of target nucleic acid molecules contained in the same sample, a plurality of types of probes 4 having base sequences complementary to the plurality of types of target nucleic acid molecules are used as the target biomolecule detection chip 1. It is fixed as a spot 5 in an independent area. The spot 5 refers to a defined area such as a circle or a rectangle, and refers to a region distributed in a two-dimensional plane that is not coupled to each other. For example, a DNA microarray in which several tens to several hundreds of circular spots 5 are arranged, A microwell plate is utilized. In the case of a DNA microarray, circular spots 5 having a diameter of several tens of μm to several hundreds of μm are arranged on a two-dimensional plane with a gap of several tens of μm to several hundreds of μm. As the target biomolecule detection chip 1, glass having a flat surface, resin, metal or the like is used. In the case of a DNA microarray, a slide glass is used. As a method for immobilizing the probe 4 to the target biomolecule detection chip 1, a method in which a solution containing DNA as the probe 4 is dropped on the target biomolecule detection chip 1 and dried is used, and an ink jet or a micro spotting device is used. A small amount of the DNA solution is dropped onto the slide glass as small droplets by using 10 to 10000 spots 5 on a 25 mm × 75 mm slide glass. In addition, the surface of the slide glass as the target biomolecule detection chip 1 is treated with various silane coupling agents having an amino group, an aldehyde group, an epoxy group, etc. A probe DNA is synthesized on the target biomolecule detection chip 1 by introducing a functional group using a molecular species such as an aldehyde group, an aldehyde group, an SH group, biotin or a thiol, and performing a coupling reaction using a photopolymerization reaction. The method to do is used.

  In addition, when detecting an antigen such as a protein or sugar chain as a target biomolecule, an antibody or a lectin that specifically binds to the protein or sugar chain is used. For example, to detect the presence of hepatitis B virus An anti-HBS antibody against HBS antigen, which is an antigen of hepatitis B virus, is used as a probe, and for example, AFP lectin is used as a probe to detect α-fetoptothein isoform, which is a liver cancer marker. In addition, as long as it is a molecule | numerator which can be couple | bonded with a specific target biomolecule as a probe 4, if it is a molecule | numerator which can couple | bond with a specific target biomolecule, protein fragments, peptides, epitopes, receptors, antigens, aptamer-allergens, etc. Can also be used. In this case as well, 0.1 μg / ml to 0.1 mg / ml anti-HBS monoclonal antibody or the like is diluted in a buffer solution such as PBS and dropped on the slide glass as 0.1 μl to 10 μl microdroplets to form a solid phase. Can be

  As shown in FIG. 1B, spots 5 are arranged in the imaging area 2, and the spots 5 are arranged based on priority. The priorities are ranked based on the importance of one or more types of the target biomolecules. For example, as shown in FIG. 1B, twelve spots 5 are arranged in the order of priority from the upper left to the upper right, and the first stage is set as the first stage, and the fifth spot 5 is arranged immediately below the first stage and directed to the right. Are arranged in the second row, and the fourth row is arranged from the left to the right under the second row to form the third row. For example, when diagnosing infectious diseases, the diseases to be detected simultaneously include plague, Marburg disease, Lassa fever, cholera, bacterial dysentery, typhoid fever, paratyphoid fever, enterohemorrhagic Escherichia coli infection, amoeba dysentery, echinococcosis, parrot disease, When detecting viral hepatitis, although the frequency of occurrence of each infectious disease is different, the frequency of occurrence of each infectious disease can be set as an importance and a priority can be set. According to the 2002 cumulative report disclosed by the National Center for Infectious Diseases Information Center, 0 plagues, 0 Marburg disease, 0 Lassa fever, 51 cholera, 699 bacterial dysentery, 63 typhoid fever , 35 cases of paratyphoid, 3183 cases of enterohemorrhagic Escherichia coli infection, 465 cases of amoeba dysentery, 10 cases of echinococcosis, 54 cases of parrot disease, 948 cases of viral hepatitis. E. coli infection, 2) viral hepatitis, 3) bacterial dysentery, 4) amoeba dysentery, 5) typhoid, 6) parrot disease, 7) cholera, 8) paratyphoid, 9) echinococcosis, 10) plague, 11) Marburg disease, 12) Lassa fever. By arranging as shown in FIG. 1B based on this priority, the target biomolecule detection chip 1 in which probes are arranged based on the onset frequency is obtained. In this case, the probe 4 is a cDNA or oligo synthetic probe for the bacterial or viral gene to be infected.

  As shown in FIG. 1 (C), the target biomolecule is extracted from the sample, then the extracted biomolecule is purified, and then the purified biomolecule is labeled with the labeled molecule, and then the labeled molecule is labeled. The target biomolecule thus formed is bound to the probe 4 on the target biomolecule detection chip 1, and then the extra substance not bound to the probe 4 on the target biomolecule detection chip 1 is washed, and then the target biomolecule detection is performed By detecting the labeled molecule bound on the chip 1, the target biomolecule is detected. For example, when detecting infected bacterial or viral DNA as a target biomolecule, sodium lauryl sulfate, N-lauroyl sarcosine sodium, lauryl phosphate, caprylate salt, collate salt, sulfone, etc. from peripheral blood as a biological sample RNA was extracted by treatment with an anionic surfactant at 37 ° C. for 10 minutes, the extracted RNA was purified using, for example, a hydroxyapatite column, and 100 μg of the purified RNA was M-MuLV reverse transcriptase. A probe containing a base sequence such as a bacterium or a virus, which is labeled in a 40 μl labeling reaction solution using Cy3-dCTP and Cy5-dCTP, and the labeled DNA is immobilized on the target biomolecule detection chip 1 In hybridization reaction for 2 hours at 4 and 50 ° C Hybridization reaction, binding, washing with 2XSCC buffer at 52 ° C., and detection of Cy3 and Cy5 of the target biomolecule bound on the target biomolecule detection chip 1 using a fluorescence microscope or a fluorescence scanner Can do.

  As shown in FIG. 1D, when detecting the target biomolecule bound on the target biomolecule detection chip 1, the imaging area 2 to which the target biomolecule is bound is imaged by one imaging using the imaging means 6. By doing so, a plurality of target biomolecules can be detected simultaneously and uniformly. For example, when detecting DNA labeled with Cy3-dCTP and Cy5-dCTP, a fluorescence scanner can be used as the imaging means 6, and when detecting Cy3, for example, an LED that emits light having a wavelength of excitation light of 550 nm is excited. By using it as a light source, Cy3 reflects light having a wavelength of 570 nm as reflected light. The reflected light that has been reflected can be quantified by, for example, photoelectric conversion using a CCD after passing through a band-pass filter. When fluorescent molecules are detected by irradiating excitation light in this way, it may be difficult to detect with high accuracy due to the fading of the fluorescent molecules, and when measuring by scanning excitation light, excitation light is measured. Although the accuracy is further deteriorated by irradiating non-fluorescent molecules, it is possible to image all the spots 5 and detect the distribution of the fluorescent molecules on the target biomolecule detection chip 1 by one imaging. As a result, the target biomolecule can be produced without degrading accuracy due to the fading of the fluorescent molecule.

  As shown in FIG. 1D, the optical image 3 captured by the imaging unit 6 indicates whether or not the target biomolecule is bound to the spot 5 arranged based on the priority order in the imaging area 2. . Spot 5 to which the target biomolecule is bound is observed as positive spot 7. For example, if it is detected that the target biomolecule is bound on up to the seventh spot 5 of the spots 5 arranged in the priority order 1 to 12, and the positive spot 7 is observed up to the seventh priority order, the priority order It is possible to judge at a glance that there are up to the seventh target biomolecules. For example, priority is given to the frequency of occurrence 1) enterohemorrhagic Escherichia coli infection, 2) viral hepatitis, 3) bacterial dysentery, 4) amoeba dysentery, 5) typhoid, 6) parrot disease, 7) cholera, 8) paratyphoid 9) Echinococcus disease, 10) Pest, 11) Marburg disease, 12) When using the target biomolecule detection chip 1 arranged with Lassa fever, it is at first glance that it is infected with an infectious disease having a higher incidence of cholera or higher. Can be judged. In addition, if the binding of the target biomolecule is observed only on one point of the 12 spots 5 arranged, it is a person who does not know the frequency of onset of the infectious disease and the statistical data of the onset frequency. Can also be judged immediately.

  In the above configuration, the spot 5 can be easily identified visually by confirming the optical image 3 obtained by photographing by the imaging means 6, and the test result can be quickly determined based on the priority order from the position of the positive spot 7. This makes it possible to make a quick diagnosis.

  Note that the shape of the spot 5 is not limited to a circle, and a spot having a shape such as a quadrangle or a hexagon can be constructed in order to increase the density of the spot 5 in the imaging area 2. For example, the target biomolecule detection chip 1 is irradiated with light so as to form a square or hexagon using a photomask of photolithography which is a semiconductor manufacturing technology, and is made into a square or hexagon using a photoreaction. The probe 4 can be immobilized by polymerizing DNA. In addition, when the probe 4 solution is drawn on the target biomolecule detection chip 1 by the ink jet method, it can be drawn on various spots 5 such as a quadrangle and a hexagon, and the spot 5 can be arranged in the imaging area 2. it can.

  Alternatively, the probe 4 immobilized in one spot 5 may be a mixed probe 4 for a target biomolecule of the same genera- tion, and may be a single spot 5 that can detect the biotarget biomolecule of the same genera. For example, Pythium family bacteria are widely distributed in environments such as soil and rivers, and it is known that there are more than 100 types of various bacterial species. As a mixed probe 4 for determining whether or not these Pythium family species are present, P. thidium P. aphanidermatum, P.A. arrhenomanes, P.M. gramicola, Pmyriotylum, P.M. sulcatum, P.M. torulosum, P.M. DNA for seven types of vanterpooli is mixed as a probe 4 and immobilized in one spot 5. If any one of the above seven species is present in the sample, the presence of a Pythium family bacterium is detected because the mixed probe 4 binds to the immobilized spot 5 Can do. In addition, as an allergen for mites present in the living environment, allergens derived from mushrooms and mites are present, but antibodies against the allergens derived from each mite are mixed and immobilized in the same spot 5 The spot 5 can be detected whether or not a tick is present.

(Embodiment 2)
FIG. 2 is a diagram showing the biomolecule detection method described in the second embodiment of the present invention. The same parts as those in the first embodiment are denoted by the same reference numerals, and detailed description thereof is omitted. 2A shows the imaging area 2 in which circular spots 8 are arranged, and FIG. 2B shows the imaging area 2 in which square spots 9 are arranged.

  As shown in FIG. 2A, when the imaging area 2 is circular, the circular spot 8 can be used to easily and efficiently arrange the imaging area 2. The circular shape is a shape composed of an arc that does not include a vertex, and is preferably a perfect circle, and it is desirable that the probe 4 is fixed to the entire inner side of the circle. Alternatively, it may be a ring-shaped spot 5 in which the probe 4 is fixed only at the outer circumference of the circular spot 8 and the probe 4 is not fixed at the center of the spot 5. When the square spots 9 are arranged at the same interval, the protruding portion 10 that protrudes from the same imaging area 2 may occur. Therefore, when the imaging area 2 is made circular and small, it is desirable that the arranged spots 5 are circular. When the shape of the imaging area 2 is changed to various shapes, the shape of the spot 5 is changed depending on the shape of the imaging area 2 in order to arrange the spots 5 so as to have the highest density in the imaging area 2. I have to let it. For example, a rectangular spot 5 is optimal for a rectangular imaging area 2, and a circular or hexagonal spot 5 is appropriate for a circular imaging area 2. When the imaging area 2 having various shapes is created, the shape of the spot 5 needs to correspond to the shape of the imaging area 2, but if the shape of the spot 5 is circular, the various imaging areas 2 It is possible to correspond to the most dense arrangement corresponding to.

  The circular spot 8 can be formed by, for example, dropping a solution containing an antibody or a DNA probe onto the target biomolecule detection chip 1 made of a slide glass as a small droplet by inkjet or the like and physically adsorbing the solution. it can. In addition, a method in which a fixed amount of a solution containing an antibody, a DNA probe, or the like is discharged to the pin tip using a micro spotter and the pin is attached to the biomolecule detection chip 1 is also used. It can be easily used as a method for forming the film. As the size of the circular spot 8, for example, in the case of a DNA microarray, a diameter of several tens of μm to several hundreds of μm is common, and the circular spot 8 is arranged on a two-dimensional plane with a gap of several tens of μm to several hundreds of μm.

  In the above configuration, a large number of spots 5 can be efficiently formed in the imaging area 2 by arranging the circular spots 8.

(Embodiment 3)
FIG. 3 is a diagram showing the biomolecule detection method according to the third embodiment of the present invention. The same parts as those in the first and second embodiments are denoted by the same reference numerals and detailed description thereof is omitted. FIG. 3A shows a case in which when the 14 circular spots 8 are arranged, the imaging area 2 is made circular and the circular imaging area 11 is formed. FIG. 3B shows a case in which when the 14 circular spots 8 having the same size are arranged, the imaging area 2 is made into a quadrangular imaging area 12. 3C shows a light irradiation area 13 that emits light when photographing the circular imaging area 11, and FIG. 3D shows a light irradiation area that emits light when photographing the rectangular imaging area 12. 13 is shown.

  The length a of the circular imaging area 11 shown in FIG. 3A is smaller than the length b of the rectangular imaging area 12 shown in FIG. 3B, and imaging is performed when the circular spot 8 having the same size is used. Area 2 can be made smaller. When the imaging area 2 has the same area, the number of circular spots 8 to be arranged can be increased, or the circular spots 8 themselves can be enlarged.

  As shown in FIGS. 3C and 3D, when the circular imaging area 11 is optically photographed, the imaging means 6 irradiates the light irradiation area 13 with light. In order to image the inside of the imaging area 2, the light irradiation area 13 needs to be larger than the imaging area 2, but when the circular imaging area 11 is irradiated with light as shown in FIG. It is not necessary to make 13 too large. When irradiating light to the rectangular imaging area 12 as shown in FIG. 3 (D), the light irradiation area 13 is circular, so that the entire rectangular imaging area 12 is covered unless the light irradiation area 13 is greatly expanded. Therefore, a portion where the circular spot 8 does not exist in the light irradiation area 13 becomes large. As described above, the circular imaging area 11 as the imaging area 2 can irradiate light more efficiently than the rectangular imaging area 12. Further, since the lens is used when photographing the imaging area 2 using the optical lens, the optical image 3 that can be acquired is circular. Therefore, as in the case of light irradiation, regarding the condensing at the time of imaging, the circular imaging area 11 can be more efficiently condensed and the sensitivity can be improved than the imaging area 2 is made the rectangular imaging area 12. The circular imaging area 11 is, for example, a circle having a diameter of 0.1 mm to 4.0 mm, in which spots having a diameter of several tens of μm to several hundreds of μm are arranged. Even if the circular imaging area 11 is colored in advance with a fluorescent paint or the like so as to be easily understood visually, a step is provided on the target biomolecule detection chip 1, for example, a chip in which the circular imaging area 11 is lowered one step may be used. Absent.

  In the above configuration, when the imaging area 2 is the circular imaging area 11, when the imaging area 2 is photographed using a lens, the light can be efficiently collected from the circular imaging area 11, so that the sensitivity increases, and the circular imaging area. Since a large number of circular spots 8 can be efficiently arranged in the area 11, target biomolecules can be detected simultaneously with high accuracy by one imaging.

(Embodiment 4)
FIG. 4 is a diagram showing the biomolecule detection method described in the fourth embodiment of the present invention. The same parts as those in the first to third embodiments are denoted by the same reference numerals, and detailed description thereof is omitted. 4A shows the donut-shaped imaging area 14, and FIG. 4B shows the light irradiation amount in the light irradiation area 13 when the donut-shaped imaging area 14 is optically imaged.

  As shown in FIG. 4A, for example, when 100 circular spots 8 are arranged on the target biomolecule detection chip 1, the circular spots 8 are arranged around the area centroid 15 of the arranged area. Instead, the area where the circular spots 8 are arranged is arranged in a donut shape, and the donut-shaped arrangement area is defined as a donut-shaped imaging area 14. The area center-of-gravity point 15 is a center-of-gravity point with respect to the area of the donut-shaped imaging area 14, and indicates a point that becomes the center of gravity with respect to the area of the area where the spot 5 is arranged even when the imaging area 2 is not circular.

  As shown in FIG. 4B, when irradiating the donut-shaped imaging area 14 with light, the amount of light in the vicinity of the optical center point 16 of the light irradiation area 13 increases, while the peripheral portion of the light irradiation area 13 has a light amount. Becomes weaker. The closer to the optical center point 16, the more light is irradiated, and the further away from the optical center point 16, the less. However, in the vicinity of the middle between the center and the outer periphery, a stable region is generated with a small difference in the amount of light to be irradiated. An area where the circular spot 8 is arranged is a donut-shaped imaging area 14 as shown in FIG. 4A, and the optical center point 16 of the light irradiation area 13 irradiated to the donut-shaped imaging area 14 is an area centroid point 15. By overlapping, it is possible to reduce unevenness in the amount of light applied to the area where the circular spot 8 is arranged. If there is a difference in the amount of light to be irradiated, the sensitivity will be different between the circular spots 8 and the measurement accuracy will be deteriorated. However, by making the imaging area 2 a donut-shaped imaging area 14, the amount of light irradiation will be reduced. By reducing the difference and reducing the sensitivity difference between the circular spots 8, it is possible to measure with high accuracy.

  In the above configuration, by making the donut-shaped imaging area 14, it is possible to photograph a region where the irradiation amount and the light collection rate are stable, and it is possible to detect the spot with high accuracy.

(Embodiment 5)
FIG. 5 shows the biomolecule detection method according to the fifth embodiment of the present invention. The same parts as those in the first to fourth embodiments are denoted by the same reference numerals, and detailed description thereof is omitted. FIG. 5A shows a case where spots are arranged in the circular imaging area 11 so that the priorities are arranged from the center toward the circumference. A spot 5 having a high priority is arranged at the center of the circular imaging area 11, and a spot 5 having a low priority is arranged on the circumference. When the target biomolecule is detected by the method shown in the first embodiment, for example, as shown in FIG. 5B, when the center of the circular imaging area 11 is the positive spot 7 and the circumferential portion is the negative spot 17, By examining the optical image 3, the examination executor can easily diagnose that only target biomolecules with high priority are present and target biomolecules with low priority are not present. On the other hand, when the center is the negative spot 17 and the circumferential portion is the positive spot 7, there is no target biomolecule with a high priority, and it is easy if only a target biomolecule with a low priority exists. Can be diagnosed.

  FIG. 5C shows another pattern arranged from the center toward the circumference. By arranging spirally from the center toward the circumference, it becomes easier to identify the spot, and the priority order of existing target biomolecules can be judged at a glance by how many spirals of the positive spot 7 are rotating. It becomes easy to do.

  In the above configuration, by arranging the spots in the circular imaging area 11 so that the priorities are arranged from the center toward the circumferential portion, the specific spot 5 can be easily identified from the optical image 3 and based on the inspection result. Diagnosis can be performed quickly.

(Embodiment 6)
FIG. 6A shows a view of the target biomolecule detection chip 1 configured on the flat substrate 18 used in the biomolecule detection method according to the sixth embodiment of the present invention as viewed from the side. The same parts as those in the first to fifth embodiments are denoted by the same reference numerals, and detailed description thereof is omitted. FIG. 6B shows that the target biomolecule detection chip 1 configured on the base 18 shown in FIG. 6A is imaged by the imaging means 6.

  As the substrate 18, for example, a substrate made of glass, resin, metal or the like, which is formed into a flat plate shape and whose surface roughness is adjusted is used. For example, when a DNA microarray is used as the target biomolecule detection chip 1, the probe 4 is immobilized on the base 18 of the slide glass by the method described in the first embodiment, and the surface roughness of the slide glass is Japan. It is defined in the industrial standard JIS B0610. As shown in FIG. 6B, the imaging means 6 is installed so as to be parallel to the flat substrate 18 and the imaging area 2 is photographed, so that all the spots 5 arranged on the imaging area 2 are captured. The optical image 3 that is focused on can be captured. When the substrate 18 is distorted or the surface roughness is high, the optical image 3 cannot be imaged while focusing on all the spots 5 on the imaging area 2. Since the target biomolecule detection method of the present invention is characterized by detecting a plurality of target biomolecules by one imaging, it is important to keep the imaging area 2 flat.

  In the above configuration, by forming the imaging area 2 with the flat base 18 having a smooth surface, an optical image can be obtained with high accuracy, and a specific spot can be easily identified from the optical image. It is possible to easily collate images and to quickly make a diagnosis based on the inspection result.

(Embodiment 7)
FIG. 7A shows a view of the target biomolecule detection chip 1 composed of the membrane 19 used in the biomolecule detection method described in the seventh embodiment of the present invention as viewed from the side. The same parts as those in the first to sixth embodiments are denoted by the same reference numerals, and detailed description thereof is omitted. FIG. 7 (B) presses the target biomolecule detection chip 1 constituted by the membrane 19 shown in FIG. 7 (A) against the smooth reference surface 20, thereby making the imaging area 2 a flat plate having a smooth surface. Shows how.

  As shown in FIG. 7A, by providing the imaging area 2 on the membrane 19 and arranging the probe 4, the target biomolecule detection chip 1 can be deformed, and the handling becomes easy. As the membrane, a nylon membrane can be used, and an antibody or oligo DNA probe immobilized on a nylon membrane can be used. Since the target biomolecule detection chip 1 is the membrane 19, when reacting with the target biomolecule, the reactivity with the probe 4 can be promoted by applying vibration to the membrane 19 using a piezoelectric element or the like, for example. Moreover, many 0.2-0.5 micrometer holes can be made in the membrane 19, and it can also be set as the state which can filter a solution. By using the target biomolecule detection chip as a filter membrane, the reaction liquid can be filtered after reacting the target biomolecule, and the operability can be improved by filtering in the same manner when washing with the washing liquid. As a method of pressing the membrane 19 against the smooth reference surface 20, there is a method of pulling both ends of the membrane 19 from the left and right with tweezers or the like and pressing it against the smooth reference surface 20 as it is. Further, as shown in FIG. 7C, the membrane 19 is sandwiched between the outer periphery of the membrane 19 with a jig 21 having the shape of the outer periphery, and the membrane kit 22 is pressed against the smooth reference surface 20. A smooth surface can be formed. Here, as described in Embodiment 6, for example, when used as a DNA microarray, the smooth surface is nylon by using a smooth reference surface 20 whose surface roughness satisfies the requirements of Japanese Industrial Standard JIS B0610. Since the membrane itself is thin and tough, it is possible to easily form a smooth surface that satisfies the JIS B0610 standard.

  In the above configuration, it is easy to handle and an optical image can be obtained with high accuracy, and a diagnosis based on a test result can be performed quickly.

(Embodiment 8)
FIG. 8 shows the biomolecule detection method according to the eighth embodiment of the present invention. The same parts as those in the first to seventh embodiments are denoted by the same reference numerals, and detailed description thereof is omitted. FIG. 8A shows gene products that function in the progression process of the cell cycle, and FIG. 8B shows the order of gene products that function in the progression process as the degree of progression. 8 (C) shows an arrangement in which circular spots 8 are arranged in the circular imaging area 11 on the basis of the priorities ranked in accordance with the degree of progress. FIG. 8 (D) is a diagram of FIG. 8 (C). The optical image 3 which image | photographed the circular imaging area 11 is shown.

  As shown in FIG. 8A, since eukaryotic cells divide and proliferate, they have a cell cycle that replicates DNA and divides cells. In the cell cycle, genes working in each process are known, and P27, cyclin E, Cdc6, Wee1, securin, cyclin A, cyclin B, and cyclin D act in the G1, S, G2, and M phases, respectively. I know.

  As shown in FIG. 8B, for example, the priority order of gene products acting as the cell cycle progresses is 1) P27, 2) cyclin E, 3) Cdc6, 4) Wee1, 5) securin, 6 ) Cyclin A, 7) Cyclin B, 8) Cyclin D.

  As shown in FIG. 8C, P27, cyclin as a target biomolecule of an animal cell using the target biomolecule detection chip 1 arranged in the circular imaging area 11 on the basis of the priority ranking corresponding to the degree of progress. E, Cdc6, Wee1, suculin, cyclin A, cyclin B, and cyclin D are detected.

  As shown in FIG. 8D, if the positive spot 7 is at the position of priority 1) and the positions 2) to 8) are the negative spot 17 in the optical image 3, the detection result is P27. As can be seen, the tester can easily diagnose that the animal cell to be tested is in the G1 phase.

  In the above configuration, the priorities are ranked according to the progress of the target diagnosis results, and the spots are arranged according to the progress of the target diagnosis results by arranging the spots 5 based on the priorities. It is possible to easily determine the degree of progress by observing the position of the spot to which the target biomolecule is bound, and to quickly determine the diagnosis and make a diagnosis.

(Embodiment 9)
FIG. 9 shows a biomolecule detection method according to the ninth embodiment of the present invention. The same parts as those in the first to eighth embodiments are denoted by the same reference numerals, and detailed description thereof is omitted. FIGS. 9A and 9B show an example of the target biomolecule detection chip 1 in which the imaging area 2 is a rectangular imaging area 12 having a square shape. FIG. 9C shows a method for imaging the rectangular imaging area 12 by the imaging means 6.

  When the circular spots 8 are arranged in the quadrangular imaging area 12, they can be arranged so that the priorities are arranged from the center toward the outer periphery as shown in FIG. 9A, and FIG. As can be seen, they can be arranged in a straight line from left to right. By arranging as shown in FIGS. 9A and 9B, the imaging area 2 becomes a square and forms a rectangular imaging area 12. When this imaging area is imaged by the imaging means 6, the imaging means 6 is imaged parallel to the target biomolecule detection chip 1 as shown in FIG. When a two-dimensional array is used, the optical image 3 is rectangular as shown in FIG. 9D, and a rectangular CCD light receiving element can be used without waste.

  In the above configuration, when the imaging area 2 is the rectangular imaging area 12 and the optical image 3 is obtained using the rectangular light receiving means 23 such as the CCD used for the imaging means 6, the light receiving element is used without waste. The spot 5 in the rectangular imaging area 12 can be well imaged. As the imaging means 6, it is sufficient if imaging can be performed, and there is a fluorescent scanner or the like.

(Embodiment 10)
FIG. 10 shows the biomolecule detection method according to the tenth embodiment of the present invention. The same parts as those in the first to ninth embodiments are denoted by the same reference numerals, and detailed description thereof is omitted. FIGS. 10A and 10B show the spots arranged so that the priorities are arranged along one side of the rectangular imaging area 12.

  FIG. 10 (A) is arranged along the X-axis, and since the priorities are arranged in one direction, the diagnosis can be immediately made according to the position of the positive spot 7.

  FIG. 10B shows an example of arrangement along the Y axis and the X axis. 1) 2) 3) is arranged from the top to the bottom of the Y axis, and 1) toward 2) 3). 4), 5) and 6) are arranged along the X axis. By arranging in stages in this way, the priorities from 1) to 9) are divided into three levels, 1) to 3) are advanced, 4) to 6) are moderate, and 7) to 9) are Can be divided into low. For example, when the positive spot 7 is at the position 4), it can be easily determined that the diagnostic result is important even in the middle.

  In the above configuration, by arranging the spots 5 so that the priorities are arranged along one side of the rectangular imaging area 12, the spots 5 can be easily identified visually. Based on this, it becomes possible to quickly determine the inspection result.

(Embodiment 11)
FIG. 11 shows an optical reading device 24 described in the eleventh embodiment of the present invention. The same parts as those in Embodiments 1 to 10 are denoted by the same reference numerals and detailed description thereof is omitted.

  As shown in FIG. 11, the optical reading device 24 of the present invention includes at least one light source 25, light receiving means 23, target biomolecule detection chip fixing means 26, spot position display means 27, and control unit 28. As the light source 25, a laser, an LED, or the like is used, and is usually adjusted using an optical filter so that light having a predetermined wavelength is irradiated. As an optical filter to be used, a short wavelength cut filter, a long wavelength cut filter, or a band pass filter is used. As the wavelength region to be adjusted, green light of about 530 nm and red light of about 630 nm are used singly or in plural. However, even in other wavelength regions, the wavelength region can interfere with fluorescent molecules. Any wavelength range can be used. A target biomolecule detection chip 1 including a plurality of spots 5 is fixed to a target biomolecule detection chip fixing means 26, and a predetermined wavelength range from a light source 25 to an imaging area 2 of the fixed target biomolecule detection chip 1. Then, the excitation light 29 is irradiated, the imaging area 2 of the target biomolecule detection chip 1 is irradiated with the excitation light 29, and the light receiving means 23 receives the reflected light 30 emitted by the excitation light 29. As the light receiving means 23, a photomultiplier tube or a charge coupled device (CCD) is used. For example, when a DNA microarray is used as the target biomolecule detection chip 1, DNA molecules labeled with fluorescent molecules such as Cy3 and Cy5 are independently bound to the slide glass at regular intervals, and these fluorescent molecules or luminescent molecules Each has a unique absorption spectrum and fluorescence wavelength. The excitation light source that interferes with the fluorescent molecule uses a light source 25 corresponding to the absorption spectrum of the fluorescent molecule, and the light receiving means 23 selects the target wavelength from the reflected light. An optical filter for limiting is used in combination. For example, since the fluorescence reflected from Cy3 and Cy5 is 570 nm and 670 nm, respectively, an optical filter that transmits light having a wavelength characteristic of 570 nm or 670 nm is used in combination. In addition, by setting the fixed time for receiving the reflected light 30 emitted and emitted from the light source 25 to, for example, 1 second to 10 seconds, light is received correspondingly from when the fluorescence amount of the spot 5 is too strong to when it is weak. be able to. The set light reception time can be adjusted according to the amount of fluorescence of the spot 5, and can be set to 10 seconds or more when the fluorescence is weak, and can be set to 1 second or less when the fluorescence is too strong. Can be set. The control unit 28 controls on / off or strength of the excitation light 29 by the light source 25. The control unit 28 is for controlling the light source 25, the light receiving unit 23, and the spot position display unit 27. For example, a CPU or the like is used. The target biomolecule detection chip immobilization means 26 is composed of a jig for sandwiching and fixing a slide glass for fixing a DNA microarray formed on the slide glass, and has a recess for fixing a microwell or the like. The holding jig etc. are used. The spot position display means 27 displays the optical image 3 and includes, for example, a liquid crystal monitor. The control unit 28 stores the captured optical image 3, and the measurer can also call the stored optical image 3 and display it on the spot position display means 27.

  In the above configuration, the measurer fixes the target biomolecule detection chip 1 to the target biomolecule detection chip immobilization means 26 and operates the control unit 28, thereby exciting light 29 in the imaging area 2 of the target biomolecule detection chip 1. Is reflected, and the reflected light 30 is received by the light receiving means 23 and displayed as the optical image 3 on the spot position display means 27 by one photographing. As described above, the light emission of the imaging area 2 is captured by the light receiving means 23 by one imaging, so that a plurality of types of target biomolecules can be quickly detected simultaneously, and the spot 5 is located in the imaging area 2. Since it is arranged based on the priority order, the spot 5 can be easily identified visually by checking the optical image 3 obtained by photographing, and the inspection result can be quickly determined based on the arranged priority order. Will be able to judge.

  In addition, the content displayed on the spot position display means 27 can display a diagnosis result based on the optical image 3 instead of the captured optical image 3, and can be used promptly for diagnosis by the user. The control unit 28 determines that the optical image 3 captured by the light receiving unit 23 is fluorescing when the light amount of the spot 5 is within the set threshold range, and the spot 5 has too little fluorescence. Alternatively, it can be determined that the spots 5 other than the too many spots 5 are not fluorescent, and the presence of the spots 5 can be displayed on the spot position display means 27. For example, the threshold is set as a diagnostic criterion. When the brightness of each pixel constituting the optical image 3 is displayed in a stage of 1 to 255, a pixel having a brightness value of stage 10 is set as a threshold value as a signal having a high possibility of a non-specific signal to be a back in advance. A pixel having a luminance value of 200 or higher as a signal having a high possibility of an abnormal signal is set to a threshold value or higher. Can be hidden, also displays the diagnosis result on the spot position display means 27 may display the diagnosis result to the test executor. For example, the control unit 28 recognizes that the spot 5 within the set threshold value has been imaged, and the name of the target biomolecule bound to the spot 5 detected by the spot display means 27 is the spot position display means 27. Can be displayed. The control unit 28 includes a microcomputer programmed with the above flow and contents. For example, a target biomolecule detection chip 1 having a spot 5 in which an antibody against mite allergen is immobilized as a probe 4, a spot 5 in which an antibody against cocoon is immobilized as a probe 4, and a spot 5 in which an antibody against a cedar pollen antigen is immobilized as a probe 4 When the fluorescence intensity is within the threshold value set only in the spot 5 in which the antibody against the mite is immobilized as the probe 4 in the picked optical image 3, the tick is detected in the spot position display means 27. A diagnostic result “tick +” that means “can be displayed. By displaying the diagnosis result on the spot position display means 27 in this way, the operation of the diagnosis by the examination executor based on the detection result can be omitted, and the diagnosis result can be obtained more quickly.

(Embodiment 12)
FIG. 12 shows an optical reading device 24 according to the twelfth embodiment of the present invention. The same parts as those in Embodiments 1 to 11 are denoted by the same reference numerals and detailed description thereof is omitted. The difference from the contents described in the eleventh embodiment is that the spot position display means 27 is replaced with the optical image monitor 31.

  The optical image monitor is, for example, a color liquid crystal monitor. By copying the optical image 3 picked up by the optical reading device 24, the position of the positive spot 7 on the target biomolecule detection chip 1 and the color are displayed to the test performer. Can provide information on the shades of For example, when a DNA microarray is used as the target biomolecule detection chip 1, for example, by displaying the spot 5 fluoresced red or green such as Cy3 or Cy5 on the optical image monitor according to the fluorescence intensity, It is possible to know the intensity and color of the fluorescence of the spot 5. Since the spots 5 are arranged based on the priority order, it is easier to diagnose by combining with the fluorescence color and the intensity of the fluorescence of the spots 5.

  In the above configuration, since the optical image monitor 31 is attached to the optical reading device 24, there is no need to install another device such as a computer or an image output device in order to measure the target biomolecule detection chip 1, and the imaging area The optical image 3 of the spot 5 in the image 2 is displayed in the optical image monitor 31, the image of the spot 5 can be easily identified from the optical image monitor 31, and the inspection result can be quickly judged based on the priority order. Therefore, the diagnosis can be performed quickly, and the optical reader is small and easy to carry.

(Embodiment 13)
FIG. 13 shows an optical reading device 24 according to the thirteenth embodiment of the present invention. The same parts as those in the first to twelfth embodiments are denoted by the same reference numerals and detailed description thereof is omitted. The difference from the content described in the twelfth embodiment is that a condensing lens 32 for condensing light on the target biomolecule detection chip 1 is further provided.

  As shown in FIG. 13, as the condenser lens 32, a straight type lens or a multi-branch type glass lens is used. Since the reflected light 30 from the imaging area 2 is normally diffused in various directions, the amount of light received by the light receiving means 23 is a part of the reflected light emitted from the imaging area 2. By providing the condenser lens 32, the reflected light 30 that could not reach the light receiving means 23 can be collected and received by the light receiving means 23, so that even when the reflected light 30 is small, it can be detected.

  In the above configuration, the condensing lens 32 is arranged in front of the light receiving means 23, whereby the optical reading device 1 with high sensitivity is obtained. In addition, since the optical reader is small and easy to carry and easy to use for diagnosis by the tester, the diagnosis using the target biomolecule detection chip becomes simple.

(Embodiment 14)
FIG. 14 shows an optical reading device 24 described in Embodiment 14 of the present invention. FIG. 14A is different from the content described in Embodiment 13 in that the light receiving means 23 is a two-dimensional array 33. It is a point that is configured. FIG. 14B shows the two-dimensional array 33. The same parts as those in the first to thirteenth embodiments are denoted by the same reference numerals and detailed description thereof is omitted.

  As the two-dimensional array 33, for example, a CCD is used, and an imaging region 2 of 1.0 mm ↑ 2 to 20.0 mm ↑ 2 can be simultaneously imaged using a CCD of 100,000 to 1 million pixels. For example, when a DNA microarray is used, since the spot 5 has a diameter of several tens to several hundreds of micrometers, about 20 to 1,000 spots should be arranged in the imaging area 2 of 1.0 mm ↑ 2. Therefore, 20 to 20,000 spots can be simultaneously imaged by using the light receiving means 23 of a two-dimensional array.

  With the above-described configuration, a plurality of types of target biomolecules can be simultaneously detected by the light receiving means 23 configured by the two-dimensional array 33. The optical system is small and easy to carry, and is easy for a tester to use for diagnosis. It becomes a reading device.

(Embodiment 15)
FIG. 15 shows an optical reading device 24 described in the fifteenth embodiment of the present invention. The difference from the content described in the fourteenth embodiment is that the light source 25 is composed of LEDs 34. The same parts as those in Embodiments 1 to 14 are denoted by the same reference numerals and detailed description thereof is omitted.

  The light receiving means 23 receives the reflected light 30 emitted from the LED 34 and emitted from the imaging area 2 by the excitation light 29. As the LEDs 34, blue, green, yellow, red LEDs 34 or the like are used singly or in plural, and the LED 34 has a double heterojunction structure or a quantum well junction structure. For example, when detecting a DNA microarray, the spot 5 labeled with Cy3 or Cy5 is imaged. When the fluorescent molecule is Cy3, the green LED 34 is used, and when the fluorescent molecule is Cy5, the red LED 34 is used. In addition, in order to further limit the wavelength of the excitation light and specifically recognize the fluorescence signal, the wavelength of the excitation light 29 can be limited using an optical filter.

In the above configuration, the LED 34 can irradiate the excitation light 29 with a small current consumption and image the imaging area 2, and the target biomolecule detection chip 1 can be remeasured because the target biomolecule on the imaging area 2 is unlikely to deteriorate. In addition, when the LED 34 irradiates light, the heat generation amount is small and the burden on the device is small, so that the device life and the life of the DNA microarray are prolonged. In addition, since the light source 25 itself is small, the fluorescence reading device 1 itself can be miniaturized, and it is small in size and can be easily carried, so that the tester can easily use it for diagnosis.

  By using the biomolecule detection method and optical reading apparatus of the present invention, a biomolecule detection device such as a DNA microarray can be efficiently detected, and a diagnosis can be made quickly and easily from the detection result. Moreover, it becomes an inexpensive apparatus configuration and can be widely spread as an optical reading apparatus that can be used by anyone. In addition, the quality of diagnostic services improves as diagnostic efficiency increases.

The figure which shows the biomolecule detection method of Embodiment 1 of this invention. The figure which shows the biomolecule detection method of Embodiment 2 of this invention. The figure which shows the biomolecule detection method of Embodiment 3 of this invention. The figure which shows the biomolecule detection method of Embodiment 4 of this invention. The figure which shows the biomolecule detection method of Embodiment 5 of this invention. The figure which shows the biomolecule detection method of Embodiment 6 of this invention. The figure which shows the biomolecule detection method of Embodiment 7 of this invention. The figure which shows the biomolecule detection method of Embodiment 8 of this invention. The figure which shows the biomolecule detection method of Embodiment 9 of this invention. The figure which shows the biomolecule detection method of Embodiment 10 of this invention. The figure which shows the optical reader of Embodiment 11 of this invention. The figure which shows the optical reader of Embodiment 12 of this invention. The figure which shows the optical reader of Embodiment 13 of this invention. The figure which shows the optical reader of Embodiment 14 of this invention. The figure which shows the optical reader of Embodiment 15 of this invention. A diagram showing a conventional hybridization reaction detection method and apparatus

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Target biomolecule detection chip 2 Imaging area 3 Optical image 4 Probe 5 Spot 6 Imaging means 7 Positive spot 8 Circular spot 9 Square spot 10 A protruding part 11 Circular imaging area 12 Rectangular imaging area 13 Light irradiation area 14 Donut-shaped imaging area DESCRIPTION OF SYMBOLS 15 Area center-of-gravity point 16 Optical center point 17 Negative spot 18 Base 19 Membrane 20 Smooth reference plane 21 The jig | tool of the outer peripheral part shape 22 Membrane kit 23 Light-receiving means 24 Optical reader 25 Light source 26 Target biomolecule detection chip immobilization means 27 Spot position display means 28 Control unit 29 Excitation light 30 Reflected light 31 Optical image monitor 32 Condensing lens 33 Two-dimensional array 34 LED

Claims (15)

A target biomolecule detection method using a target biomolecule detection chip on which one or more types of probes that bind to a target biomolecule are immobilized, wherein each of the probes is independent on the target biomolecule detection chip The spots are fixed as described above, the spots are arranged on the target biomolecule detection chip on the basis of priority, and one or more types of the spots are arranged in the imaging area of the target biomolecule detection chip. A biomolecule detection method, wherein the biomolecule is detected by optically imaging the imaging area at one time. The biomolecule detection method according to claim 1, wherein the spot is circular. The biomolecule detection method according to claim 1, wherein a target biomolecule detection chip having a circular imaging area is used. The biomolecule detection method according to any one of claims 1 to 3, wherein a target biomolecule detection chip in which an imaging area in which no spot is arranged at the center has a donut shape is used. The biomolecule detection method according to claim 3, wherein the spots are arranged so that the priorities are arranged from the center of the imaging area toward the circumferential portion. The method for detecting a biomolecule according to any one of claims 1 to 5, wherein a target biomolecule detection chip comprising a flat substrate having an imaging area having a smooth surface is used. The biomolecule detection method according to claim 1, wherein the imaging area is formed of a membrane, and the imaging area is pressed against a smooth reference surface having a smooth surface. The biomolecule detection method according to any one of claims 1 to 7, wherein the priorities are ranked according to the degree of progress of a target diagnosis result. The biomolecule detection method according to claim 1, wherein a target biomolecule detection chip having a square imaging area is used. The biomolecule detection method according to claim 9, wherein the spots are arranged so that the priorities are arranged along one side of the imaging area. An optical reader for imaging a target biomolecule detection chip in which one or more types of probes that bind to a target biomolecule are immobilized as one or more independent spots, and immobilizes the target biomolecule detection chip Target biomolecule detection chip immobilization means, one or more light sources that irradiate excitation light in a predetermined wavelength range on the biomolecule detection chip, and light emission by the excitation light on the target biomolecule detection chip Light receiving means that is a photoelectric conversion element that receives light in a predetermined wavelength range, and optically receives one or more types of spots arranged in an imaging area according to a priority order based on the importance of the target biomolecule An optical reading device characterized in that an image is picked up once. The optical reading apparatus according to claim 11, further comprising an optical image monitor that displays the picked-up optical image, wherein the optical image picked up by one photographing is displayed on a single screen. The optical reading device according to claim 11, further comprising a condensing lens that condenses the light emitted on the target biomolecule detection chip. 14. The optical reading device according to claim 11, wherein the light receiving means is constituted by a two-dimensional array. The optical reading device according to claim 11, wherein the light source is an LED.

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