WO2004063731A1 - Capteur optique - Google Patents

Capteur optique Download PDF

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Publication number
WO2004063731A1
WO2004063731A1 PCT/JP2004/000318 JP2004000318W WO2004063731A1 WO 2004063731 A1 WO2004063731 A1 WO 2004063731A1 JP 2004000318 W JP2004000318 W JP 2004000318W WO 2004063731 A1 WO2004063731 A1 WO 2004063731A1
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WO
WIPO (PCT)
Prior art keywords
light source
sample
led light
light
semiconductor light
Prior art date
Application number
PCT/JP2004/000318
Other languages
English (en)
Japanese (ja)
Inventor
Toshinobu Niimura
Akihiro Namba
Takami Shibazaki
Original Assignee
Olympus Corporation
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Olympus Corporation filed Critical Olympus Corporation
Priority to JP2005508020A priority Critical patent/JPWO2004063731A1/ja
Publication of WO2004063731A1 publication Critical patent/WO2004063731A1/fr

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • G01N21/6452—Individual samples arranged in a regular 2D-array, e.g. multiwell plates

Definitions

  • the present invention relates to a photodetection device applied to an inspection device for a biological substance used for analysis of nucleic acids such as DNA and DNA, antigen-antibody reaction, and protein binding reaction. '' Background technology
  • the most commonly used genetic testing methods include extracting nucleic acids from biological samples and using PCR (Polymerase Chain Reaction) or NASBA (Nucleic Acid Sequence-Based Amplification).
  • the target gene is amplified by amplifying the nucleic acid labeled with a radioisotope or a fluorescent dye using the method of amplification (based on J), and the base sequence of the target gene or Methods for measuring the concentration are known.
  • the electrophoresis method is used for analysis of gene expression and mutation analysis.
  • the electrophoresis method has problems such as that it takes time and effort to perform the measurement, and there is a limit to the measurement that can be performed at one time. Therefore, recently, capillary electrophoresis, which allows a fluorescently labeled nucleic acid to react in a plurality of cavities and rapidly process many samples at once, has been widely used. According to the capillary electrophoresis method, the measurement can be performed in a shorter time and more easily than the method using the conventional electrophoresis method.
  • the DNA chip has a large number of DNA probes immobilized on the surface of a glass substrate, and a large number of oligo probes synthesized on a small area on a silicon wafer by applying a semiconductor manufacturing process. There is a DNA chip.
  • the test method using a DNA chip as described above, multiple nucleotide sequences and expression levels of DNA in a sample can be determined simultaneously. Furthermore, many gene expression levels / multiple mutations can be determined by applying a DNA chip. It is possible to carry out various tests, such as analysis of data. Furthermore, from data obtained using a DNA chip, many genes can be classified into multiple loops, and information on gene fluctuations associated with development and differentiation can be obtained.
  • the gene analysis method using a DNA chip has the following problems. It has the advantage of performing many tests at once, but requires long test times. Since there are many inspection steps throughout and a complicated operation is required, it is difficult to obtain a reproducible detection result.
  • a gene inspection device using a DNA microarray has recently been considered.
  • genetic analysis is performed using fluorescence emitted from reaction products generated in the reaction chamber of the DNA microarray.
  • This device is a microscope-based device.
  • a hybridization reaction was caused between the target nucleic acid and the nucleic acid probe, which had been fluorescently labeled in advance, in the reaction tank of the DNA microarray, and the reaction was captured by the DNA microarray.
  • a fluorescence signal from a fluorescent substance is acquired, and a fluorescence image is obtained based on the signal.
  • the DNA microarray has a reaction tank (liquid storage section).
  • a fluorescent substance is efficiently excited to generate stable fluorescence from a reaction product generated in a reaction chamber of the DNA microarray. This is extremely important in performing highly accurate gene analysis.
  • LED Light Emitting Diode
  • Excitation light in this case is used to irradiate the fluorescent substance with the excitation light to excite the fluorescent substance, and to provide a filter means for selectively transmitting the wavelength of the incident light with respect to the fluorescence emitted by the excited fluorescent substance.
  • An apparatus has been proposed in which a fluorescent signal is received by a body imaging device and a fluorescent image is obtained based on the fluorescent signal (for example, see Japanese Patent Application Laid-Open No. 10-132744). .
  • the light from the LED and the like is irradiated to the well as excitation light via a dichroic mirror and an objective lens, and the fluorescent light emitted from the sample is passed through the objective lens and the dichroic mirror.
  • An apparatus has also been proposed in which a photomultiplier tube is used for detection via a pinhole cutout (see Japanese Patent Application Laid-Open No. 200-11616).
  • An apparatus has been proposed (see Japanese Patent Publication No. 7-122628).
  • LED light sources do not have sufficient light intensity to excite the fluorescent material and cannot generate sufficient fluorescence from the fluorescent material. For this reason, when an LED is used as a light source, it is often difficult to receive light with a commonly used photodetector such as a CCD camera. Therefore, when an LED light source is used as a light source for exciting a fluorescent substance, the output intensity should be as low as possible. In addition to selecting a model with a large surface area, bundle two or more LEDs so that they are as close to the sample surface as possible, and make sure that a sufficient amount of excitation light is uniformly applied to the fluorescent material. It is necessary to take measures such as doing so.
  • a photodetector capable of efficiently irradiating sufficient light with high efficiency and performing highly reliable fluorescence detection.
  • a light detection device includes: a semiconductor light source unit that emits light for irradiating a labeled substance of a sample; and a device that collects light emitted by irradiating the labeled substance with the irradiation light.
  • a condenser lens for emitting light, a filter for selectively transmitting light emitted when the labeling substance is irradiated with light by the irradiation light, and a light for transmitting light passing through the condenser lens.
  • a photodetector having a photodetector for detection, wherein the optical path of the excitation light is different from the optical path of the photodetector.
  • a light detection device includes: a semiconductor light source unit that emits light for irradiating a labeled substance of a sample; and the semiconductor light source unit includes a semiconductor light emitting element and an optical element.
  • a condenser lens for condensing light emitted by irradiating the labeling substance with the irradiation light, and selectively condensing light emitted by irradiating the labeling substance with the irradiation light.
  • a photodetector having a photodetector that detects light passing through the light-collecting lens.
  • FIG. 1 is a schematic diagram of an optical inspection apparatus according to the first embodiment of the present invention. The figure which shows schematic structure.
  • FIG. 2 is a diagram showing a schematic configuration of an example of a DNA reaction vessel used in the first embodiment.
  • FIG. 3 is a diagram showing a schematic configuration of another example of the DNA reaction vessel used in the first embodiment.
  • FIGS. 4A and 4B are views for explaining light irradiation on a sample according to the first embodiment.
  • FIG. 5 is a block diagram showing the entire optical inspection apparatus according to the first embodiment.
  • FIG. 6 is a diagram showing a schematic configuration of an LED light source unit according to a first modification of the first embodiment.
  • FIG. 7 is a diagram showing a schematic configuration of an LED light source unit according to a second modification of the first embodiment.
  • FIG. 8 is a diagram showing a schematic configuration of an LED light source unit according to a third modification of the first embodiment.
  • FIG. 9 is a diagram showing a schematic configuration of a main part of an optical inspection device according to a second embodiment of the present invention.
  • FIG. 10 is a view showing a schematic configuration of an optical inspection device according to a modification of the second embodiment.
  • FIGS. 11A and 11B are diagrams showing a schematic configuration of an optical inspection device according to a third embodiment of the present invention.
  • FIG. 12 is a diagram showing a schematic configuration of a measurement optical system used in the third embodiment.
  • FIG. 13 is a block diagram showing an LED light source unit drive circuit used in the third embodiment.
  • FIG. 14 is a diagram illustrating a schematic configuration of an LED light source driving circuit used in the third embodiment.
  • FIG. 15 is a diagram showing a schematic configuration of an optical inspection device according to a fourth embodiment of the present invention.
  • FIG. 16 is a diagram showing a schematic configuration of an optical inspection device according to a first modification of the fourth embodiment.
  • FIG. 17 is a diagram illustrating a schematic configuration of an optical inspection device according to a second modification of the fourth embodiment.
  • FIG. 18 is a diagram showing a schematic configuration of an optical inspection device according to a fifth embodiment of the present invention.
  • FIG. 19 is a diagram illustrating a schematic configuration of an optical inspection device according to a modification of the fifth embodiment.
  • a fluorescence detection apparatus that irradiates a fluorescent substance with excitation light.
  • a fluorescent detector that inspects fluorescence from the fluorescent substance will be described. It is of course applicable to a detection device that detects light or reflected light (hereinafter, these are collectively referred to as a “light detection device”).
  • Various fluorescent dyes and fluorescent glass particles can also be used as the fluorescent substance.
  • metal particles or dielectric particles are used as a labeling substance when detecting with scattered light or reflected light.
  • fine particles such as silver, platinum, and silicon, and latex particles can be used.
  • fine particles of metals such as gold, silver, and platinum have a particle size of 10 to 100 nm.
  • the speed of the particles in motion is optimal.
  • Latex particles having a particle size of 0.1 to 1 m are also particularly preferred because the speed of the particles in motion is also optimal.
  • the appropriate particle size is determined by the specific gravity of the particles and the speed of the browning motion.
  • the motion state of the particles includes, for example, Brownian motion and vibration.
  • telomere binding substance refers to hormones, tumor markers, enzymes, antibodies, antigens, abzymes, other proteins, nucleic acids, cDNA, DNA, RNA, and PNA. And the like, meaning a substance that can specifically bind to a biological substance, and is called a probe.
  • a “biological substance” is a substance that specifically binds to a known specific binding substance placed at a predetermined position on a substrate on which a probe is immobilized, and is extracted and isolated from a living body.
  • the term refers to substances that have been subjected to chemical treatment, chemical treatment, chemical modification, and the like, as well as substances directly extracted from living organisms.
  • substances such as hormones, tumor markers, enzymes, antibodies, antigens, abzymes, other proteins, nucleic acids, cDNA, DNA, RNA, and PNA.
  • Specific binding between a biological substance and a “specific binding substance” means that an unstable double strand is formed between complementary nucleotide sequences found in DNA, RNA, etc. Or highly specific binding, such as in the case of hybridization (hybridization) or selectively reacting only with a specific substance, such as an antigen and an antibody or a biotin and avidin. (First Embodiment)
  • FIG. 1 is a diagram showing a schematic configuration of an optical inspection device according to a first embodiment of the present invention.
  • An optical inspection device according to each embodiment of the present invention includes a light-collecting lens, a sample holding unit, a light source, and a photodetector.
  • the optical inspection apparatus includes a sample stage 1, and a DNA reaction container 2 as a subject is placed on the sample stage 1.
  • FIG. 2 is a diagram showing a DNA slide glass reaction vessel 3 as a specific example of the DNA reaction vessel 2.
  • the DNA slide glass reaction vessel 3 has a sample tank 301 arranged in a slide glass-like reaction vessel. In the sample tank 301, a DNA microarray 302 is laid.
  • a sample solution test sample
  • a hybrid is formed between the nucleic acid probe and the target nucleic acid previously fluorescently labeled in the DNA microarray 302. Redidation reaction occurs.
  • a fluorescent substance for example, a fluorescent dye substance
  • captured by the DNA microarray emits fluorescence.
  • the sample solution that did not contribute to the reaction is washed together with the buffer.
  • FIG. 3 is a diagram showing a specific example of another DNA reaction container 4.
  • the DNA reaction container 4 is formed of a plastic material.
  • a plurality of sample vessels 401 are arranged in the DNA reaction vessel 4, and a DNA microarray 402 is laid in the sample vessel 401.
  • a hybridization reaction occurs between the target nucleic acid and the nucleic acid probe that have been fluorescently labeled in advance.
  • a fluorescent substance for example, a fluorescent dye, captured in the DNA microarray by the hybridization reaction emits fluorescence. In this case, the sample solution that did not contribute to the reaction is washed together with the buffer.
  • an objective lens 5 is arranged as a focusing lens.
  • the objective lens 5 is positioned on a perpendicular line from one surface of the sample stage so that its optical axis 5a coincides with the center of the sample vessel 201.
  • the objective lens 5 is held by an objective lens holding mechanism 6.
  • the objective lens holding mechanism 6 has a cylindrical holding member 600.
  • the base end of the objective lens 5 is fitted inside the holding member 600.
  • a plurality of screw holes 62 are formed on the peripheral surface of the holding member 61 at equal intervals along the circumferential direction.
  • a fixing member 603 is provided so that the peripheral surface of the holding member 601 abuts.
  • the fixing member 603 is fixed to a device main body (not shown), and an elongated hole 604 along the optical axis direction of the objective lens 5 is formed in the fixing member 603.
  • a position adjusting screw 605 is screwed into the screw hole 602 on the peripheral surface of the holding member 601 via the elongated hole 604. By loosening the screws for position adjustment 605, the objective lens 5 moves along with the holding member 601 along the elongated hole 604 in the direction of the optical axis 5a. This allows you to adjust the focus.
  • a filter 7, an imaging lens 8, and a light detection device that together with the objective lens 5 constitute a light detection unit.
  • Container 9 is placed on the optical axis 5 a of the objective lens 5, a filter 7, an imaging lens 8, and a light detection device that together with the objective lens 5 constitute a light detection unit.
  • Container 9 is placed on the optical axis 5 a of the objective lens 5, a filter 7, an imaging lens 8, and a light detection device that together with the objective lens 5 constitute a light detection unit.
  • Container 9 is placed on the optical axis 5 a of the objective lens 5, a filter 7, an imaging lens 8, and a light detection device that together with the objective lens 5 constitute a light detection unit.
  • Container 9 is placed on the optical axis 5 a of the objective lens 5.
  • the filter 7 selectively transmits the fluorescence emitted when the fluorescent substance labeled on the sample 202 is excited.
  • the imaging lens 8 forms the fluorescent light selected by the filter 7 on the detection surface of the photodetector 9.
  • the imaging lens 8 may be a glass lens such as BK7 used for a normal lens as a material, but the imaging lens 8 may be quartz glass, a plastic lens, or Diffractive optical elements, liquid crystal lenses, etc. Elements and materials capable of condensing ordinary visible light can be used.
  • LED light source units 11 for example, four LED light source units 11; in FIG. 1, three LED light source units 11 are provided as semiconductor light source means. (Shown) is arranged.
  • the LED light source unit 11 has a cylindrical LED light source holder 111.
  • the LED light source holder 1101 has an LED light source 1102 disposed therein.
  • a non-pass filter 1103 is disposed on the optical path of the light emitted from the LED light source 1102 disposed inside the LED light source holder 1101.
  • the non-linear filter 1103 mainly passes near the wavelength range of the light emitted from the LED light source 1102.
  • the spectral characteristics of the emission wavelengths of the LED light sources 1102 included in the plural LED light source units 11 are the same.
  • the light from the LED light source unit 11 is supplied to the sample tank on the sample stage 1.
  • the 201 sample 202 is irradiated as excitation light.
  • Each LED light source unit 11 is mounted so that no excitation mura occurs.
  • the central axis of the light of each LED light source unit 11 corresponds to the periphery a, b, c of the sample 202 surface.
  • the mounting position is set. In this case, if it is possible to prevent the occurrence of excitation mura on the sample 202 surface, even if the position is off the center axis of the sample 202 surface, It is also possible to set the mounting position of each LED light unit 11 so that it intersects at one point of the upper or lower spatial position.
  • the LED light source unit 11 moves the sample 20
  • the excitation light applied to the sample 2 is not limited to the sample
  • the main part of the optical inspection device thus configured is installed in the light-shielding box 12 and is shielded from the outside.
  • Figure 5 shows a block diagram of the entire optical inspection system.
  • the computer 10 has a monitor 13 as a display means.
  • the computer 10 includes a fluorescence detection unit 14 including a main part of the apparatus shown in Fig. 1 and an LED light source unit driving circuit as a driving means for driving each LED light unit 11. 1 5 and are connected.
  • the LED light source drive circuit 15 is driven by one of the LED light source units 11 in the fluorescence detection unit 14.
  • the drive current of the LED light source unit 11 is set and the drive current is supplied to the LED light unit 11.
  • the LED light source unit drive circuit 15 responds to the content of the command at this time, and the fluorescence detection unit 1 4 Determine the LED light source unit 11 driven from the power of the LED light source unit 11 in 1 and at the same time determine the magnitude of the drive current of the LED light source unit 11 Supply drive current to unit 11.
  • each LED light source 1 102 As shown in Fig. 4A and Fig. 4B, the central axis of each light passes through the periphery a, b, c, d of the specimen 202 surface so that excitation unevenness does not occur on the surface.
  • the LED light sources 1102 are arranged so as to intersect at one point e (or ⁇ ) of a spatial position vertically above (or below) the substantially central axis of the sample 202 surface.
  • the fluorescent substance in the sample 202 emits fluorescence.
  • the fluorescent light is condensed by the objective lens 5, passes through the filter 7 and the imaging lens 8, and forms an image on the detection surface of the photodetector 9.
  • the photodetector 9 detects the intensity of the fluorescent light, converts it into an electric signal, and outputs the electric signal to the computer 10.
  • the computer 10 performs image processing and signal analysis such as contour enhancement, contrast correction, color correction, etc., and displays it on the monitor 13 as a fluorescent image.
  • a plurality of LED light source units 11 (for example, four LED light source units) having the LED light sources 1102 having the same emission wavelength spectrum are used.
  • the unit 1 1) was arranged side by side around the objective lens 5, and the excitation light was irradiated on the sample 202 simultaneously by the multiple LED light source units 1 1. Excitation light of sufficient intensity to excite the fluorescent substance in 2 can be efficiently obtained.
  • the center axis of each light passes through the periphery of the sample 202 surface and the sample 202 so that the light from the plurality of LED light sources 1102 becomes uniform on the sample 202 surface. Since they are set to intersect at one point vertically above or below the space on the approximate center axis of the two surfaces, The shadow and excitation unevenness on the surface of the sample 202 can be eliminated, and more stable fluorescence can be generated than the fluorescent substance generated in the sample tank 201.
  • the sample 202 Specularly reflected light from the surface does not pass through the light receiving optical path of the objective lens 5 as it is and enters the photodetector 9 as noise light, so that noise light can be reduced.
  • the LED is used as an excitation light source for exciting a fluorescent substance, it is inexpensive, has a long life, generates little heat, and is safe. In addition, power consumption can be reduced. In addition, the device configuration can be made small and highly portable.
  • the LED light source unit 11 has only the band-pass filter 1103 as an optical element other than the LED light source 1102, the configuration is simple. Not only is it easy to assemble and adjust the optical axis, but it is also useful because it leads to cost reduction.
  • the LED light source unit 11 by disposing the LED light source unit 11 around the objective lens 5, the optical path of the excitation light and the optical path having the objective lens 5 for fluorescence detection are separated from each other. There is no need to install extra optical elements such as a Crook Mirror, and a simple device configuration can be achieved in this regard. In addition, since there is no optical element such as a die-cloth mirror, reflected light and scattered light generated on the surface of these optical elements, and light passes through these optical elements As a result, the loss of the light intensity detected by the photodetector 9 can be minimized because the fluorescence intensity does not attenuate.
  • FIG. 6 shows a first embodiment of the first embodiment.
  • FIG. 9 is a diagram showing a schematic configuration of an LED light source unit 16 according to a modification.
  • the LED light source unit 16 has a cylindrical LED light source holder 1601.
  • the LED light source holder 1601 has an LED light source 1602 disposed therein.
  • a non-pass filter 1603, a diffusion plate 1604, A light lens 1605 is arranged on the optical path of the light emitted from the LED light source 1602 disposed inside the LED light source holder 16-011.
  • the Nordpass filter 1603 mainly transmits near the wavelength range of the light emitted from the LED light source 1602.
  • the diffuser plate 164 is used to suppress the non-uniformity (intensity unevenness) of the light intensity from the LED light source 1602 that has passed through the non-pass finoletor 1603.
  • the diffusion plate 1604 for example, "frosted glass” or a translucent plastic plate is used.
  • the condenser lens 1605 is the light diffused by the diffuser 1604. And focuses the diffused light at a position determined by the focal length.
  • the positions of the non-pass filter 1603 and the diffuser 1604 are the same as the diffuser 1604, the nozzle 1601 and the condenser lens.
  • the order may be 1 6 0 5.
  • the LED light source unit 16 has a condensing lens 1605 on the optical path of light emitted from the LED light source 1602. It can irradiate a focused beam.
  • a condensing lens 1605 having a NA of 0.95 or more is used, a very small area on the sample 202 surface, for example, a diameter of 0.5.
  • Light can be collected in the range of about ⁇ m, and a specific extremely narrow range of excitation can be achieved.
  • the excitation light from the LED light source 1602 becomes a condensed beam and the area of the irradiation cross section is small, a portion other than the sample 202 surface, such as the side wall of the sample tank 201, is used.
  • each LED light source 1602 is adjusted so that the excitation light from the LED light source 1602 is focused on one point in the plane of the sample 202, the LED light source 16 It is possible to irradiate the excitation light from O2 only to a desired specific portion in the surface of the sample 202. As a result, it is possible to efficiently irradiate the excitation light from the LED light source 1602 to a desired portion in the plane of the sample 202.
  • FIG. 14 is a diagram showing a schematic configuration of an LED light source unit 17 according to a second modification.
  • the LED light source unit 17 has a cylindrical LED light source holder 1701.
  • the LED light source holder 1701 has an LED light source 1702 disposed therein.
  • a bandpass filter 1701 on the optical path of the light emitted from the LED light source 1702 disposed inside the LED light source holder 1701, a bandpass filter 1701, a diffusion plate 1704, and a There is a lens 1705.
  • Band No. The finoletor 1703 and the diffuser 1704 are the same as those described with reference to FIG.
  • the collimating lens 1705 converts the light diffused by the diffusion plate 1704 into parallel light, and emits collimated light. In this case.
  • the positions of the band-noise filter 1703 and the diffuser 1704 are the same as those of the diffuser 1704, the non-pass filter 170, and the coil.
  • the order of the lenses 1705 can be used.
  • the LED light source unit 17 has a collimated lens 105 disposed on the optical path of light emitted from the LED light source 1702, so that the sample It is possible to irradiate the flat collimated light onto the 202 surface.
  • This makes it easy to identify the irradiation position in the plane of the sample 202, making it easy to program a program for automatically adjusting the irradiation position by computer control.
  • it is easy and useful to predict the irradiation surface position and its illuminance.
  • the excitation light can be accurately operated at a desired irradiation position on the sample 202 surface. .
  • since light is not diffused, it is possible to prevent erroneous irradiation of light in a portion of the sample tank 201 other than the sample 202 surface, for example, including a side wall. it can.
  • FIG. 14 is a diagram illustrating a schematic configuration of an LED light source unit 18 according to a third modification of the first embodiment.
  • the LED light source unit 18 has a cylindrical LED light source holder 1801.
  • the LED light source holder 1801 has an LED light source 1802 disposed therein.
  • a non-pass filter 1803 and a diffusion plate 1804 are disposed on the optical path of the light emitted from the LED light source 1802 disposed inside the LED light source holder 1801.
  • the band pass filter 1803 and the diffusion ⁇ 1804 are the same as those described with reference to FIG. In this case, the position of the non-pass filter 1803 and the diffusion plate 1804 may be in the order of the diffusion plate 1804 and the bandpass filter 1803.
  • the LED light source unit 18 according to the third modification has the diffusion plate 1804 arranged on the optical path of the light emitted from the LED light source 1802. Irradiate diffuse light without unevenness You can do it. As a result, it is possible to irradiate almost uniformly the excitation light over the entire surface of the sample 202 without unevenness, and it is possible to receive stable and excellent fluorescence with excellent reproducibility. .
  • LED light source units 11, 16, 16, 17 can be used properly according to the size and structure of the sample 202 and the sample tank 201. That is, the LED light source unit 1 1
  • Each of the LED light sources unit 16, 17, and 18 is composed of an LED light source and an optical element such as a non- zero filter, a lens, and a diffusion plate.
  • the excitation light can be condensed, collimated, or diffused in units of light. This makes it possible to control the beam pattern of the excitation light to the sample 202 surface in various ways, and the sample tank 201 of various shapes has more unevenness. It can respond to a certain sample.
  • the LED light source unit 16 (17) it can be used according to the application. Either the filter 1603 (1703) or the diffusion plate 1604 (1704) may be used, or neither of them may be used.
  • Scanners 1103, 1603, 1703, and 1803 are different depending on the spectrum of the emission wavelength of the fluorescent dye used as the sample. ° Needless to say, it may be a filter or a low-pass filter.
  • the condenser lens 1705 6 The collimating lens 1705 may be a glass lens such as BK7 used for ordinary lenses as a material, but the focusing lens 16 0 5 ⁇ Collimate lens 1 7 0 5 is quartz glass or plastic lens or diffractive optical element Elements and materials that can condense ordinary visible light, such as liquid crystal lenses, can be used.
  • the first embodiment the case where a plurality of LED light source units 11 are arranged around the objective lens 5 has been described. In 1, only one LED light source unit 11 disposed around the objective lens 5 may be used. Even in this case, the same effect as described above can be expected.
  • the second embodiment is an embodiment in which the angle of light irradiation can be adjusted by changing the inclination angle of the LED light source unit with respect to the sample surface.
  • FIG. 9 is a diagram illustrating a schematic configuration of the second embodiment.
  • the objective lens 21 is held by an objective lens holding mechanism 22.
  • the objective lens holding mechanism 22 has a cylindrical holding member 222.
  • the objective lens 21 is fitted inside the holding member 222.
  • an LED light source unit holder 23 is provided as a semiconductor light source means holding member.
  • the LED light source unit holder 23 is provided with a plurality of (for example, about 8 to 12) LED light source unit storage holes 2301 at even intervals along the circumferential direction. . These LED light source cut-out holes 2301 are arranged to be inclined toward the surface of the sample 202 on the sample stage 1. The tilt angle is about 45 ° to 60 ° with respect to the optical axis 21a of the objective lens 21 with respect to the axis.
  • the LED light source unit holder 23 has an L A screw hole 2302 penetrating the ED light source unit storage hole 2301 is provided. The two screw holes 2302 are formed at predetermined intervals in the direction along each LED light source unit accommodation hole 2301.
  • the LED light source unit 24 is accommodated in the LED light source unit accommodating hole 2301.
  • the LED light source unit 24 has a cylindrical holoreder 2401, and the LED illuminator 2402 is disposed inside the holoreder 2401. Then, such an LED light source unit 24 is housed in the LED light source unit housing hole 2301 via an O-ring 25. In this case, the O-ring 25 is located between the two screw holes 2302.
  • the LED light source unit storage hole 2301 has a bandpass filter 240 on the optical path of the light emitted from the LED light source 2402 of the LED light source unit 24. 3 is located.
  • the non-phosphorescent light source 2403 mainly passes near the wavelength range of light emitted from the LED light source 2402 power.
  • a spacer 26 is disposed between the hologram 2404 of the LED light source unit 24 and the finolators 2403 of the LED light source unit.
  • the spacer 26 is composed of an LED light source 2402 and a band laser. Performs positioning of the fin locator 2403.
  • a metal such as aluminum or brass or a plastic is used.
  • a tilting screw 27 as a position adjusting means is screwed.
  • Fan screw 27 is for LED light source unit 2 Press the two points across the O-ring 25 on the side of the holder 2 4 0 1.
  • the light irradiation angle can be adjusted by changing the tilt angle of the LED light source unit 24 with respect to the sample 202 surface by adjusting the pressing force of the lifting screw 27 according to the screwing amount. .
  • the irradiation angle of the light emitted from the LED light source 2402 adjusted by these two tilting screws 27 is ⁇ 2 to 3. It is on the order.
  • a plurality of screw holes 222 are formed on the peripheral surface of the holding member 222 at even intervals along the circumferential direction. Further, a fixing member 222 is provided so that the peripheral surface of the holding member 222 contacts.
  • the fixing member 2203 is fixed to the apparatus main body 28 side, and has an elongated hole '224' formed along the optical axis direction of the objective lens 21.
  • a positioning screw 222 is screwed into the screw hole 222 on the peripheral surface of the holding member 222 through the elongated hole 222.
  • the objective lens 21 is moved along with the holding members 222 along the elongated holes 222. Move in the optical axis 21a direction. This allows the focus to be adjusted.
  • the LED light sources 2402 of each LED light source unit 24 are turned on, and the excitation light emitted from these LED light sources 2402 is applied to the sample 202 surface on the sample stage 1.
  • the light emission spectrum of the light emitted from the LED light source 2402 is all the same.
  • Focus adjustment is performed by moving the optical axis in the 2 la direction (the direction of arrow A in the figure).
  • each LED light source unit 24 passes through the periphery of the sample 202 surface and is vertically above or below the substantially central axis of the sample 202 surface. Set to intersect at one point of the position. This makes it possible to illuminate the sample 202 surface uniformly and uniformly.
  • the screwing amount of the tilting screw 27 is manually adjusted.
  • the screwing of the tilting screw 27 is performed.
  • a motor at the position corresponding to the adjustment of the control amount and to operate this motor automatically by linking it with the computer.
  • the LED light source unit 24 may be configured to be capable of adjusting the position in the height direction and adjusting at least one of the inclination angles.
  • FIG. 10 is a diagram showing a schematic configuration of an optical inspection device according to a modification of the second embodiment, and the same parts as those in FIG. 1 are denoted by the same reference numerals. In this modification, the block diagram of the optical inspection device described with reference to FIG. 5 is used.
  • a light detector 31 is disposed on the sample table 1 to which light from the plurality of LED light source units 11 is irradiated, instead of the sample tank 201 as light intensity detecting means.
  • a solid-state image sensor CCD camera or CMOS sensor
  • an image pickup tube or the like is used as the light detector 31.
  • the photodetector 31 individually detects the intensity of light from each LED light source unit 11. The detection output of the photodetector 31 is taken into the computer 10 described in FIG.
  • the computer 10 analyzes the variation of the light intensity from the output of the photodetector 31 corresponding to the intensity of the light from each LED light source unit 11 and outputs the LED light source unit drive circuit 15. And individually controls the current supplied to the LED light source 1102 of each LED light unit 11.
  • the magnitude of the current supplied to each LED light source 1102 can be controlled based on the light intensity obtained near the sample surface. This makes it possible to illuminate the specimen surface with uniform brightness. As a result, illumination unevenness on the sample 202 surface can be suppressed, the fluorescent substance in the sample tank 201 can be excited almost uniformly, and highly reliable fluorescence detection can be performed. And can be.
  • each LED light source unit 11 obtained by the photodetector 31 was adjusted.
  • the LED light source unit drive circuit 15 is manually adjusted based on the intensity of light from the LED light source 1102 to adjust the light from the LED light source 1102 to the same brightness.
  • the photodetector 31 may be removed, and the sample tank 201 may be re-installed at this position.
  • an imaging means for example, a CCD camera or a CMOS sensor (both not shown)
  • the sample tank 201 is placed on the sample table 1 as it is, and The light from the LED light source 111 of the LED light source unit 11 is individually radiated to the sample tank 201, and the fluorescent light from the sample tank 201 is objective lens 5, filter 7, imaging lens 8 , Each of which is taken by the image pickup means, guided to the computer 10 and displayed on the monitor 13, and the brightness of each pixel of the fluorescent image on the monitor 13 is set to the computer 10.
  • the LED light source unit drive circuit 15 is adjusted, and each of the LED light source unit drive circuits 15 is adjusted.
  • the current supplied to the LED light sources 11.02 can be individually controlled.
  • the light from each LED light source 1102 can be adjusted to the same brightness, so that uneven illumination on the sample 202 surface can be suppressed, and the sample tank can be controlled.
  • the fluorescent substance in 201 can be excited almost uniformly.
  • the third embodiment is an embodiment in which a plurality of types of fluorescent dyes are excited using two or more LED light sources having different peak emission wavelengths, and a fluorescent signal can be detected.
  • FIGS. 11A and 11B are diagrams showing a schematic configuration of the third embodiment, and the same parts as those in FIG. 9 are denoted by the same reference numerals.
  • the block diagram of the optical inspection apparatus described with reference to FIG. 5 is used.
  • the first to third LED light source units 41, 42, and 43 having three types of LED light sources having different peak emission wavelengths are mounted on the LED light source unit holder 23.
  • the emission spectrum of the LED light source has a mountain-shaped structure, and has one peak wavelength.
  • the first LED light source unit 41 uses an LED light source having a light emission peak wavelength of 490 nm
  • the second LED light source unit 42 uses the light emission peak wavelength.
  • An LED light source having a wavelength of 52 O nm is used, and an LED light source having a peak emission wavelength of 63 O nm is used as the third LED light source unit 43.
  • the first to third LED light source units 41, 42, and 43 are L Four ED light source unit holders 23 are arranged symmetrically around the objective lens 21 along the periphery. In other words, these first to third LED light source units 41, 42, and 43 have LED light sources having different emission wavelength spectral characteristics, respectively, and these different spectra
  • the first to third LED light source units 41, 42, and 43 having the same characteristics are arranged at regular intervals along the periphery of the LED light source unit holder 23.
  • the first LED light source unit 41, the second LED light source unit 41, and the third LED light source unit 43 are repeatedly arranged in this order.
  • the first to third LED light source units 41, 42, and 43 have the same spectral characteristics at positions facing each other across the objective lens 21. It is arranged as follows.
  • the first to third LED light source units 41, 42, and 43 are mounted in front of the respective LED light sources (in the drawing, the first LED light source unit). 4)
  • the LED light source 4101 in front of the LED light source 4101 (only the filter 4102 is shown) transmits the light near the peak wavelength of the emission of each LED light source best. t that is made as having the property of, Roh down de path full I filter 4 1 0 2 of the first LED light source Interview two Tsu DOO 4 1, the wavelength to be the most good rather permeation 4 9 O n ra
  • the second LED light source unit is set to a nearby position.
  • the filter has the best transmission wavelength set around 520 nm
  • the third LED light source unit 43 has a non-pass filter with the best transmission wavelength around 630 nm.
  • Fig. 12 shows the schematic configuration of the photodetection optical system of the device configured as described above.
  • the LED light source unit having the above three different wavelengths at the peak emission wavelength is shown. 4 1, 4 2,
  • Figure 4 shows the case where the sample is irradiated with excitation light.
  • two dichroic mirrors 45 and 46 are arranged on the optical axis above the objective lens 21.
  • the dichroic mirror 45 is set so that the reflected light travels in a direction of approximately 45 ° with respect to the optical axis of the objective lens 21, and the peak emission of the fluorescent dye FITC is performed.
  • a transflective spectral characteristic that reflects light having a wavelength slightly longer than the wavelength of 520 nm, for example, light having a wavelength of 550 nm or less, and transmits light having a wavelength longer than that. have.
  • the dichroic mirror 46 is installed so that the reflected light travels in a direction of approximately 45 ° with respect to the optical axis of the objective lens 21, and the fluorescent dye Cy 3 Peak emission wavelength of 5 6
  • transflective spectral characteristic that reflects light having a wavelength slightly longer than 5 nm, for example, light having a wavelength of 62 nm or less, and transmits light having a wavelength longer than that. I have.
  • a CCD camera 48 is disposed on the reflection optical path of the die mirror 45 through a condenser lens 47.
  • a CCD camera 50 is arranged via a condenser lens 49 in the reflected light path of the die-cloth mirror 46, and a condensing lens 51 is arranged in the transmitted light path.
  • CCD camera 52 is arranged in the transmitted light path.
  • the outputs of the CCD cameras 48, 50, 52 are sent to the computer 10.
  • three types of target DNA are now set for one sample, and different fluorescent dyes, FITC (Forescein-isothiocyanate), Cy3, and Cy5, are used for each of them.
  • Label Next, a sample solution containing DNA labeled with these fluorescent dyes is dropped into the sample tank, DNA hybridization is performed, and the labeling substance that has not contributed to the reaction is removed with a buffer solution (PBS (PBS)). Wash with a mixture of phosphoric acid buffer), EDTA (ethylene sodium acetate sodium nitrate), and NaCl (pH 7.4).
  • PBS buffer solution
  • the sample obtained in this way is set on the sample stage 1 shown in FIGS. 11A and 11B. Then, the sample tank 201 is irradiated with excitation light from each LED light source unit 41, 42, 43. At this time, the light from the LED light source having three different wavelengths as the peak emission wavelengths is simultaneously irradiated onto the sample surface.
  • fluorescence is emitted from the fluorescent dye labeled on the sample that has reacted with the DNA probe by the DNA hybridization, and the light passes through the objective lens 21 shown in Fig. 12 and is diced. Reutz Miller 4 5 is reached.
  • the fluorescence from the fluorescent dye FITC is reflected by the dichroic mirror 45, passes through the lens 47, enters the CCD camera 48, and is guided to the computer 10. It is obtained as a green fluorescent image by FITC.
  • the fluorescence of the fluorescent dye Cy 3 passes through the dichroic mirror 45, is reflected by the dichroic mirror 46, passes through the lens 49, and passes through the lens 49.
  • FIG. 13 shows a block diagram of the drive circuits of the first to third LED light source units 41, 42, and 43.
  • an LED light source unit driving circuit 54 and an LED light source unit driving circuit 55 are respectively provided.
  • An LED light source unit driving circuit 56 is provided.
  • the LED light source unit drive circuit 54 is connected to the first LED light source unit 41 and the LED light source unit drive circuit 55 is connected to the second LED light source unit 42.
  • An LED light source unit drive circuit 56 is connected to the third LED light source unit 43.
  • a common power supply 53 is connected to the LED light source unit driving circuit 54, the LED light source unit driving circuit 55, and the LED light source unit driving circuit 56, and as described in FIG. Combiner 10 is connected.
  • the LED light source unit drive circuit 54, the LED light source unit drive circuit 55, and the LED light source unit drive circuit 56 receive power from a common power supply device 53. Also, LED light source- The LED drive circuit 54, the LED light source unit drive circuit 55, and the LED light source unit drive circuit 56 are controlled based on a command from the computer 10, and are controlled by the first LED light source unit. A driving current is supplied to each of the LED light sources of the light source 41, the second LED light source unit 42, and the third LED light source unit 43 to generate excitation light of three kinds of fluorescent dyes.
  • the three types of fluorescence signals obtained by these excitation lights are respectively transmitted by the CCD cameras 48 and 46 by the dichroic mirrors 45 and 46, respectively. , 50 and 52 separately, and the image output signals from these CCD cameras 48, 50 and 52 are guided to the computer 10, where image analysis is performed, and three types of images are obtained. A composite image using the fluorescent dye is generated and output.
  • FIG. 14 shows a specific circuit configuration of the power supply unit 53 and the LED light source unit drive circuits 54, 55, and 56.
  • the power supply device 53 guides the output from the AC power supply 57 of 100 V to the transformer circuit 58 composed of a transformer to adjust the voltage.
  • the output from the transformer circuit 58 is sent to a bridge rectifier circuit 59 composed of four diodes to perform full-wave rectification, and then smoothed by a smoothing capacitor 60.
  • the smoothed output is led to a constant voltage circuit 6 4 composed of two power transistors 6 16 2 connected in Darlington and an OP amplifier 63, and a constant voltage is output. Is output.
  • the terminal voltage of the Zener diode 65 is used as a reference voltage, and a variable reference voltage is generated by a variable reference voltage generation circuit 67 composed of circuit elements such as a P-amplifier 66 and a resistor. are doing.
  • a variable reference voltage generation circuit 67 composed of circuit elements such as a P-amplifier 66 and a resistor. are doing.
  • variable reference resistor 68 By changing the value of the variable reference resistor 68 connected to the amplifier 63, the voltage output of the constant voltage circuit 64 is adjusted in relation to the variable reference voltage of the variable reference voltage generation circuit 67. Then, a drive current is supplied to the LED light source unit drive circuit 54, the LED light source unit drive circuit 55, and the LED light source unit drive circuit 56. In other words, by changing the value of the variable reference resistor 68, the driving current to the LED light source unit driving circuit 54, the LED light source unit driving circuit 55, and the LED light source unit driving circuit 56 is increased. Can be adjusted at the same time.
  • a FET (field effect transistor) 70 generates an abrupt current in the event of a short-circuit when the current is short-circuited, and flows through the OP amplifier 63. It plays a role in preventing accidental damage.
  • the power can be turned on and off by the power switch 69.
  • the power supply stitch 69 is connected to the computer 10, and the power input is also adjusted by the computer 10.
  • the power supply switch 69 may be manually switched.
  • LED light source unit driving circuits 54, 55, 56 are connected in parallel.
  • the LED light source unit drive circuit 54 serves as a drive circuit for the four first LED light source units 41 described in FIGS. 11A and 11B, and includes a variable resistor 71 and a variable resistor 71. scan I Tutsi 7 2 t Similarly the series circuit are provided separately to each, LED light source unit drive circuit 5 5 also as the second LED light source Interview two Tsu DOO 4 2 of the driving circuit of the four partial
  • a series circuit of a variable resistor 71 and a switch 72 is separately prepared, and the LED light source unit driving circuit 56 is also equipped with a third LED light source unit for four.
  • As a drive circuit for the unit 43 a series circuit of a variable resistor 71 and a switch 72 is separately provided.
  • the first to third LED light source units 4 are controlled by adjusting the resistance values of the respective variable resistors 71, 73, and 75.
  • the drive current supplied to each of the LED light sources 1, 42, and 43 can be adjusted, and the output light intensity can be controlled.
  • the adjustment of the output light intensity may be performed manually, or may be automatically performed automatically by a computer. This makes it possible to individually control the drive current of each of the first to third LED light source units 41, 42, and 43.
  • the LED light source unit driving circuits 54, 55, and 56 control the on / off of the switches 72, 74, and 76 to control the first to third LED light source units 4.
  • the LED light sources 1, 42, and 43 can all be turned on and off independently. These on / off controls may be performed automatically by computer control or manually.
  • the first to third LED light source units 4 As described with reference to FIG.
  • the light intensity from each LED light source of 1, 42, 43 is detected, the detection signal is led to a computer, and the light intensity is analyzed by a computer, and the variable resistors 71, 73, 7 Adjust the resistance of 5.
  • the resistance values of the variable resistors 71, 73, and 75 may be manually adjusted based on the light intensities from the individual LED light sources obtained by this detection. This Thereby, it is possible to adjust so that the light from the LED light source is almost uniformly irradiated to a part of the reaction tank of the DNA microarray.
  • variable resistances 71, 73, and 75 can control the magnitude of the drive current flowing through all LED light sources, so that the intensity of the excitation light can be increased or reduced all at once. You can do it. As a result, it is possible to irradiate each sample with excitation light of an appropriate intensity, and it is possible to efficiently detect fluorescence.
  • three types of fluorescent dyes can be formed by using the first to third LED light source units 41, 42, and 43 having three types of LED light sources having different peak emission wavelengths. Since simultaneous excitation is possible, fluorescence images from these three types of fluorescent dyes can be acquired, and these can be synthesized or displayed individually.
  • the individual LED light sources constituting the first to third LED light source units 41, 42, and 43 can easily be turned on and off according to the fluorescent dye to be excited.
  • the drive current can be individually adjusted by the variable operation of the variable resistance, so that each fluorescent dye can be irradiated with excitation light of the correct intensity, and the efficiency can be improved. Fluorescence can be detected well.
  • the present invention is not limited to this, and two or more types of fluorescent dyes may be obtained in the same manner.
  • the fluorescent dye is not limited to FITC, Cy3, and Cy5 shown in the examples, but may be a load-min green (Rhodamine G), a Texas-red (Texas). Red), RITC (Rhodamine B-isothiocya), etc. may be used.
  • the number of LED light source units is not particularly limited, and a plurality of LED light source units are individually controlled by a plurality of drive circuits.
  • the present invention can be applied even if the output wavelengths of a plurality of LED light source units are the same or different. It is also possible to irradiate the sample with one type of light from LED light sources having different peak emission wavelengths, and to repeat the light several times at different wavelengths. For example, it is possible to use a turret in which filters with different transmission wavelengths are set and automatically switch by computer in conjunction with LED light sources of different wavelengths. . .
  • a reflecting member is arranged on the optical path of light emitted from the LED light source unit, and the light in the optical path is reflected, so that the optical axis angle of the excitation light can be increased and the light of the objective lens can be increased. This is an embodiment that approaches an axis angle.
  • FIG. 15 is a diagram showing a schematic configuration of the fourth embodiment, and the same parts as those in FIG. 1 are denoted by the same reference numerals.
  • LED light source units 81 as semiconductor light source means (four LED light units in the illustrated example) are used.
  • the LED light source unit 81 has a cylindrical LED light source holder 8101.
  • the LED light source holder 8101 has an LED light source 8102 disposed therein.
  • a bandpass filter 8103, a diffusion plate 8104, a condenser lens 8 1 0 5 is arranged on the optical path of light emitted from the LED light source 8102 inside the LED light source holder 8101.
  • the non-noise filter 8103 mainly passes near the wavelength range of light emitted from the LED light source 8102.
  • reflectors 82 are provided as reflecting members.
  • the reflecting plate 82 reflects light from the LED light source unit 81 and irradiates the sample 202 in the sample tank 201 on the sample stage 1 as excitation light.
  • the reflector 82 reduces the angle ⁇ formed by the optical axis of the excitation light with respect to the optical axis of the objective lens 5 so that the optical axis angle of the excitation light approaches the optical axis angle of the objective lens. I'm doing it.
  • the respective mounting positions of the reflectors 8 2 reflect the light from the LED light source unit 81 and irradiate the sample 20 ′ 2 surface with uniform brightness and irradiate the light.
  • shadows and excitation unevenness also occur on the sample 202 having a three-dimensional part on part or the entire surface. It is adjusted so that it does not occur.
  • a reflector 82 is arranged on the optical path of the light emitted from the LED light source unit 81 to reflect the light in the optical path, so that the optical axis angle of the excitation light can be adjusted by the objective lens 5. It can approach the optical axis angle. This means that the excitation light can be radiated from a position close to the top of the sample surface 202 (the limit that does not enter the observation field of view of the objective lens 5), so that the LED light source unit The difference between the shortest distance a and the longest distance b of the optical path from 81 to the sample 202 surface through the reflector 82 can be reduced. As a result, since the intensity of the excitation light is inversely proportional to the distance of the optical path, the difference in the intensity of the excitation light on the surface of the sample 202 can be reduced. The two surfaces can be excited efficiently and uniformly.
  • the reflection plate 82 is arranged as a reflection member on the optical path of the light emitted from the LED light source unit 81 to reflect the light in the optical path. If such a method is used, the degree of freedom in arranging the LED light source units 81 increases, so that a large number of LED light source units 81 are arranged around the objective lens 5. It can be.
  • FIG. 16 is a diagram showing a schematic configuration of an optical inspection apparatus according to a first modification of the fourth embodiment. The same parts as those in FIG. The label is attached.
  • a plurality of LED light source units 81 as semiconductor light source means are arranged in multiple layers (two layers in the illustrated example).
  • reflecting plates 82, 82 ' (corresponding to the LED light source units 81 of each layer) as reflecting members. (Corresponding to 2 layers in the example shown).
  • the reflectors 82 and 82 ' reflect light from the LED light source unit 81 of each layer and serve as excitation light for the sample 202 in the sample tank 201 on the sample stage 1. And irradiate.
  • the light from the LED light source unit 81 is reflected to irradiate the sample 202 with uniform brightness even at the respective mounting positions of the reflectors 82 and 82 '.
  • the sample 202 having a three-dimensional portion on part or the entire surface is also shaded.
  • Fig. 15 is the same as Fig. 15 for other adjustments so that no excitation unevenness occurs.
  • the light from the light source unit 81 of each layer is reflected by using the reflectors 82, 82 '.
  • LED light source units 81 since the number of LED light source units 81 can be greatly increased, these LED light source units 81 are divided into a plurality of sets, and the emission wavelength spectrum is increased for each set. By using LED light sources with different torque characteristics, it is possible to excite a plurality of types of fluorescent dyes and detect fluorescent signals in the same manner as described in the third embodiment. And
  • the reflection plate 82 is disposed as a reflection member on the optical path of the light emitted from the LED light source unit 81 to reflect the light in the optical path.
  • a die crack mirror can be used as a member.
  • FIG. 17 is a diagram showing a schematic configuration of the second modification, and the same parts as those in FIG. 15 are denoted by the same reference numerals.
  • the LED light source unit 8 1 has an LED light source 8 1 0 2 disposed inside a cylindrical LED light source holder 8 1 0 1 and an LED light source 8 1 0 inside the LED light source holder 8 1 0 1.
  • a diffusing plate 8104 and a condensing lens 8105 are arranged on the optical path of the light emitted from 2, and the above-described bandpass finoleta 8103 is omitted.
  • a die-cloth mirror 83 is arranged as a reflecting member.
  • the dichroic mirror 8 3 mainly reflects near the wavelength range of the light emitted from the LED light source 8 102 of the LED light source unit 81. It has such characteristics.
  • the die-cloth mirror 83 reflects the light from the LED light source unit 81 and applies excitation light to the sample 202 in the sample tank 201 on the sample stage 1. And irradiate. Also in this case, the angle between the optical axis of the excitation light and the optical axis of the objective lens 5 is reduced so that the optical axis angle of the excitation light approaches the optical axis angle of the objective lens.
  • the die-cloth mirror 83 has a characteristic of mainly reflecting near the wavelength range of light emitted from the LED light source 8102 of the LED light source unit 81.
  • the LED it is possible to omit the bandpass finolators built in the LED light source unit 81. As a result, the cost of the LED light source unit 81 can be reduced and the size can be reduced.
  • the light guide member is arranged on the optical path of the light emitted from the LED light source unit, and the optical path is bent, so that the optical axis angle of the excitation light is increased and the optical axis angle of the objective lens is increased. This is an embodiment in which the distance is approached.
  • FIG. 18 shows a schematic configuration of the fifth embodiment, and the same parts as those in FIGS. 1 and 15 are denoted by the same reference numerals.
  • the light source unit 81 is arranged around the objective lens 5.
  • the LED light source unit 81 has a cylindrical LED light source holder 8101.
  • the LED light source holder 8101 has an LED light source 8102 disposed therein.
  • a bandpass filter 8103, a diffusion plate 8104, 810 5 are arranged on the optical path of the light emitted from the LED light source 8102 inside the LED light source HONOLEDA 8101.
  • the non-pass filter 8103 mainly transmits light near the wavelength range of the light emitted from the LED light source 8102.
  • a light incident end 84a of an optical fiber 84 is arranged separately as a light guide member.
  • the optical fiber 84 can bend the optical path of the light from the LED light source unit 81 freely.
  • the light from the light emitting end 84b is irradiated as the excitation light onto the sample 202 in the sample tank 201 on the sample stage 1, and at the same time, the light from the objective lens 5
  • the angle formed by the optical axis of the excitation light with respect to the optical axis can be reduced so that the optical axis angle of the excitation light approaches the optical axis angle of the objective lens.
  • the position of the light emitting end 84b of these optical fibers 84 is caused by the fact that the emitted light is applied to the sample 202 surface with uniform brightness and the incident angle of the applied light. Adjustment so that excitation unevenness does not occur on the sample 202 surface, and also so that shadows and excitation unevenness do not occur on the sample 202 having a three-dimensional part on all or part of the surface. Have been.
  • the same effects as in the first embodiment can be expected.
  • the optical path is bent by the optical fiber 84 arranged on the optical path of the light emitted from the LED light source unit 81, the optical axis angle of the excitation light is adjusted by the objective lens. It can approach the optical axis angle.
  • the difference in the intensity of the excitation light on the sample 202 surface can be reduced, and the sample 202 surface can be efficiently used. It can be excited uniformly.
  • the optical guide member 84 is used as the light guide member.
  • a material obtained by molding glass rod-shaped acrylic material is applied. You can also do it.
  • a control method for illuminating a sample uniformly and uniformly with light as described in the modification of the second embodiment and the third embodiment. Can be applied.
  • the optical fiber 84 is arranged as a light guide member on the optical path of the light emitted from the LED light source unit 81, and the optical path is bent. If such a method is used, the degree of freedom in arranging the LED light source units 81 increases, so that it is possible to arrange a large number of LED light source units 81 around the objective lens 5. it can.
  • FIG. 19 is a diagram showing a schematic configuration of an optical inspection device according to a modification of the fifth embodiment, and the same parts as those in FIG. 18 are denoted by the same reference numerals.
  • a plurality two in the illustrated example
  • a plurality of LED light source units 81 as semiconductor light source means are arranged around the objective lens 5.
  • a light incident end 84a of an optical canopy refino 84 as a light guide member is separately arranged.
  • the optical fiber 84 allows the light path of the light from the LED light source unit 81 to be freely bent, and the light from the light emitting end 84b is transferred to the sample on the sample stage 1.
  • the sample 202 in the tank 201 is irradiated as excitation light.
  • the position of the light emitting end 84b of the optical fiber 84 is such that the emitted light is applied to the surface of the sample 202 with uniform brightness, and the light to be applied is In order to prevent excitation unevenness from occurring on the sample 202 surface due to the incident angle, no shadow or excitation unevenness occurs on the sample 202 having a three-dimensional part on all or part of the surface. It has been adjusted accordingly.
  • the optical path of the light from the light source unit 81 is bent by using the optical fin 84, so that the position of each LED light source unit 81 is automatically determined.
  • the degree of freedom can be increased. By increasing the degree of freedom in the location of the LED light source units 81, it is possible to use more LED light source units 81 than in the fifth embodiment. And are possible. This makes it possible to easily obtain the amount of light required to irradiate the sample 202 surface, and to illuminate the sample 202 surface without unevenness in the light amount.
  • the LED light source units 81 are divided into a plurality of sets, and the spectral characteristics of the emission wavelength are set for each set. By using different LED light sources, it is also possible to excite a plurality of types of fluorescent dyes and detect fluorescent signals as in the third embodiment.
  • optical fiber 84 is used as the light guide member.
  • a material obtained by molding a glass rod material may be used. it can.
  • the present invention is not limited to the above-described embodiment, and can be variously modified in the implementation stage without departing from the spirit of the invention.
  • the sample applied in each of the above-described embodiments is not limited to an example of a DNA microarray, but can be applied to all reaction vessels called so-called DNA microarrays.
  • the present invention is widely applicable not only to DNA but also to detection and measurement dealing with various other biological materials.
  • the above-described embodiment includes various stages of the invention, and various inventions can be extracted by appropriately combining a plurality of disclosed constituent elements. For example, even if some components are deleted from all the components shown in the embodiments, the problem described in the section of the problem to be solved by the invention can be solved, and the problem described in the section of the effect of the invention can be solved. If the effect obtained is obtained, a configuration from which this component is deleted can be extracted as an invention.
  • light is applied to the labeled substance of the sample. Since the optical path for irradiation and the optical path of the fluorescence detecting means having the focusing lens are separated, optical elements such as dichroic mirrors that attenuate light can be omitted, and the detected light intensity Loss can be minimized.
  • a plurality of semiconductor light source means are arranged along the periphery of the converging lens and light is irradiated simultaneously from these, so that the labeling substance is irradiated with light. It is possible to efficiently obtain light of sufficient intensity to carry out.
  • the direction of the light emitted from the semiconductor light source means and the direction of inclination of the condenser lens with respect to the optical axis can be adjusted, respectively, so that the desired specific Light can be condensed at the location, and efficient irradiation of the labeling substance with light can be performed.
  • the drive current to the semiconductor light source means can be individually controlled based on the light intensity obtained near the sample, the light from the semiconductor light source means can be controlled by the same light. Brightness can be adjusted, and the sample can be illuminated with uniform brightness.
  • the present invention relates to a photodetection device that can efficiently irradiate sufficient light and can perform highly reliable photodetection.

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Abstract

L'invention porte sur un capteur optique comprenant: une source lumineuse à semi-conducteur éclairant la substance marquée d'un échantillon, une lentille focalisant la lumière produite sur la substance marquée, un filtre transmettant sélectivement la lumière produite lorsque la substance marquée est éclairée, et un capteur optique dont l'élément photosensible détecte la lumière ayant traversé la lentille de focalisation. La trajectoire optique de la lumière d'éclairage diffère de celle du capteur optique.
PCT/JP2004/000318 2003-01-16 2004-01-16 Capteur optique WO2004063731A1 (fr)

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WO2004063731A1 true WO2004063731A1 (fr) 2004-07-29

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Cited By (13)

* Cited by examiner, † Cited by third party
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JP2013089167A (ja) * 2011-10-21 2013-05-13 Toppan Printing Co Ltd 識別装置
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JP2017173820A (ja) * 2016-03-21 2017-09-28 カール ツァイス マイクロスコピー ゲーエムベーハーＣａｒｌ Ｚｅｉｓｓ Ｍｉｃｒｏｓｃｏｐｙ Ｇｍｂｈ 光シート顕微鏡および光シート顕微鏡を作動するための方法
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EP3330697A1 (fr) * 2016-11-30 2018-06-06 Bayer Aktiengesellschaft Dispositif de détermination de l'action d'agents actifs sur nématodes et autres organismes dans des essais aqueux
JP2019002933A (ja) * 2009-09-21 2019-01-10 アコーニ バイオシステムズ インコーポレイテッド 光学システム
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CN113167462A (zh) * 2019-07-19 2021-07-23 高科技器械私人有限公司 光学系统和照射样品平面的方法
KR20220005179A (ko) * 2020-07-06 2022-01-13 한림대학교 산학협력단 스테인드 글라스 방식의 다중채널 형광 검출 장치
JP2022088444A (ja) * 2017-03-07 2022-06-14 イルミナ インコーポレイテッド 単一の光源、２光学チャネル配列決定

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EP1774295A1 (fr) * 2004-08-02 2007-04-18 Inodiag Dispositif de lecture pour lames portant des micro depots supports de reaction biologique
JP2019002933A (ja) * 2009-09-21 2019-01-10 アコーニ バイオシステムズ インコーポレイテッド 光学システム
US10363558B2 (en) 2010-08-31 2019-07-30 Canon U.S. Life Sciences, Inc. System and method for serial processing of multiple nucleic acid assays
JP2013544490A (ja) * 2010-08-31 2013-12-19 キヤノン ユー．エス． ライフ サイエンシズ， インコーポレイテッド 高分解能熱融解の検出のための光学システム
US10266873B2 (en) 2010-08-31 2019-04-23 Canon U.S. Life Sciences, Inc. Optical system for high resolution thermal melt detection
JP2013089167A (ja) * 2011-10-21 2013-05-13 Toppan Printing Co Ltd 識別装置
JPWO2013137247A1 (ja) * 2012-03-12 2015-08-03 三菱レイヨン株式会社 蛍光検出装置及び蛍光検出方法
WO2013137247A1 (fr) * 2012-03-12 2013-09-19 三菱レイヨン株式会社 Dispositif de détection de fluorescence et procédé de détection de fluorescence
CN104204778A (zh) * 2012-03-12 2014-12-10 三菱丽阳株式会社 荧光检测装置及荧光检测方法
US9395303B2 (en) 2012-03-12 2016-07-19 Mitsubishi Rayon Co., Ltd. Fluorescence detection device and fluorescence detection method
JP2019505794A (ja) * 2016-01-13 2019-02-28 インスティトゥート ドクトル フェルスター ゲゼルシャフト ミット ベシュレンクテル ハフツング ウント コンパニー コマンディトゲゼルシャフト インジケータの発光を発生させて測定するためのデバイスを含む爆発物を検出するための可搬デバイス
JP2017173820A (ja) * 2016-03-21 2017-09-28 カール ツァイス マイクロスコピー ゲーエムベーハーＣａｒｌ Ｚｅｉｓｓ Ｍｉｃｒｏｓｃｏｐｙ Ｇｍｂｈ 光シート顕微鏡および光シート顕微鏡を作動するための方法
CN107526156A (zh) * 2016-03-21 2017-12-29 卡尔蔡司显微镜有限责任公司 光片显微镜以及用于运行光片显微镜的方法
WO2018069358A1 (fr) * 2016-10-11 2018-04-19 Cork Institute Of Technology Système de détection de fluorescence
US10859495B2 (en) 2016-10-11 2020-12-08 Cork Institute Of Technology Fluorescence sensing system
JP2020501138A (ja) * 2016-11-30 2020-01-16 バイエル、アクチエンゲゼルシャフトＢａｙｅｒ Ａｋｔｉｅｎｇｅｓｅｌｌｓｃｈａｆｔ 水性試験における線虫および他生物に対する活性成分の効果を立証するための装置
EP3330697A1 (fr) * 2016-11-30 2018-06-06 Bayer Aktiengesellschaft Dispositif de détermination de l'action d'agents actifs sur nématodes et autres organismes dans des essais aqueux
CN109997027A (zh) * 2016-11-30 2019-07-09 拜耳股份公司 记录具有一个或多个空腔的细胞培养板的全区域图像的装置
JP2022088444A (ja) * 2017-03-07 2022-06-14 イルミナ インコーポレイテッド 単一の光源、２光学チャネル配列決定
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JP2022543930A (ja) * 2019-07-19 2022-10-17 アドバンスド インストゥルメント プライベート リミテッド 光学システム、及びサンプル面の照明方法
JP7357618B2 (ja) 2019-07-19 2023-10-06 アドバンスド インストゥルメント プライベート リミテッド 光学システム、及びサンプル面の照明方法
KR20220005179A (ko) * 2020-07-06 2022-01-13 한림대학교 산학협력단 스테인드 글라스 방식의 다중채널 형광 검출 장치
KR102376680B1 (ko) * 2020-07-06 2022-03-18 한림대학교 산학협력단 스테인드 글라스 방식의 다중채널 형광 검출 장치

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