WO2008126672A1 - Microscope et cube à fluorescence à placer à l'intérieur - Google Patents

Microscope et cube à fluorescence à placer à l'intérieur Download PDF

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Publication number
WO2008126672A1
WO2008126672A1 PCT/JP2008/055627 JP2008055627W WO2008126672A1 WO 2008126672 A1 WO2008126672 A1 WO 2008126672A1 JP 2008055627 W JP2008055627 W JP 2008055627W WO 2008126672 A1 WO2008126672 A1 WO 2008126672A1
Authority
WO
WIPO (PCT)
Prior art keywords
optical
microscope
optical system
fluorescent
microscope apparatus
Prior art date
Application number
PCT/JP2008/055627
Other languages
English (en)
Japanese (ja)
Inventor
Toshiaki Nihoshi
Original Assignee
Nikon 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
Priority claimed from JP2007329046A external-priority patent/JP5286774B2/ja
Application filed by Nikon Corporation filed Critical Nikon Corporation
Publication of WO2008126672A1 publication Critical patent/WO2008126672A1/fr
Priority to US12/563,141 priority Critical patent/US8014065B2/en

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Classifications

    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/16—Microscopes adapted for ultraviolet illumination ; Fluorescence microscopes
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/0004—Microscopes specially adapted for specific applications
    • G02B21/002—Scanning microscopes
    • G02B21/0024—Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
    • G02B21/0032—Optical details of illumination, e.g. light-sources, pinholes, beam splitters, slits, fibers

Definitions

  • the present invention relates to a microscope apparatus capable of fluorescence observation and a fluorescent cube used in the microscope apparatus.
  • Confocal microscopes and total reflection fluorescence microscopes are widely used as methods for observing living cells, etc., and both are common in that they use a laser light source, and can be used as confocal microscopes and total reflection fluorescence microscopes.
  • Has been proposed see, for example, Japanese Patent Laid-Open No. 2 0 0 5-1 2 1 7 96).
  • a first aspect of the present invention includes an illumination optical system that irradiates a sample with a ray beam from a laser light source, a fluorescence detection optical system that detects fluorescence from the sample, and the illumination optical system.
  • a microscope apparatus provided in the optical path and having a plurality of fluorescent cubes for guiding the laser beam to the specimen, and an objective lens, at least one of the fluorescent cubes is configured to illuminate a chief ray of the laser beam.
  • An optical means for condensing the laser light beam at a predetermined position away from the optical axis of the pupil position of the objective lens is provided so as to be substantially parallel to the optical axis of the optical system.
  • Obvious A microscopic device is provided.
  • a second aspect of the present invention is a fluorescent cube that is replaceably disposed in the optical path of the illumination optical system of the fluorescence microscope, and when placed in the optical path of the fluorescence microscope, the second aspect of the present invention is configured to pass through an object lens.
  • the principal beam of the laser beam irradiated onto the specimen is made substantially parallel to the optical axis of the illumination optical system, and the laser beam is collected at a predetermined position away from the optical axis of the pupil position of the objective lens.
  • a fluorescent cube characterized by having an optical means for emitting light.
  • the microscope apparatus which can be switched from a confocal microscope to a total reflection fluorescence microscope can be provided by switching the fluorescence cube currently used for the fluorescence microscope.
  • a fluorescent cube that can be switched from a confocal microscope to a total reflection fluorescent microscope.
  • FIG. 1 is a schematic configuration diagram of a microscope apparatus according to the first embodiment.
  • FIG. 2 shows an optical system of the microscope apparatus according to the first embodiment. .
  • FIG. 3 shows the optical system of the microscope apparatus when the optical system of the microscope apparatus according to the first embodiment is switched to the total reflection microscope.
  • FIGS. 4A, 4B, and 4C are diagrams for explaining the operation of the optical means in the microscope apparatus according to the first embodiment.
  • FIG. 4A shows a confocal scanning illumination state or an epi-illumination state
  • FIG. 4C is a diagram for explaining the action of the wedge prism.
  • FIG. 5 shows an optical system of a microscope according to the second embodiment.
  • FIG. 6 shows the optical system of the microscope apparatus when the optical system of the microscope according to the second embodiment is switched to a total reflection microscope.
  • FIG. 1 is a schematic configuration diagram of a microscope apparatus according to the first embodiment.
  • FIG. 2 shows an optical system of the microscope apparatus according to the first embodiment.
  • Figure 3 shows the optical system of the microscope device when switched to the total reflection microscope.
  • 4A, 4B, and 4C are diagrams for explaining the operation of the optical means.
  • FIG. 4A shows the confocal scanning illumination state or the epi-illumination state
  • FIG. 4B shows the total reflection illumination state
  • FIG. 4C FIG. 6 is a diagram for explaining the operation of a wedge prism.
  • the transmission illumination optical system described later is omitted.
  • the microscope apparatus 1 includes an inverted fluorescent microscope main body 2 (hereinafter simply referred to as a microscope) and a control apparatus 3 (hereinafter referred to as a personal computer for controlling various devices mounted on the microscope 2). , PC)).
  • the microscope 2 illuminates the specimen 1 2 placed on the stage 1 1 with the light from the transmitted illumination light source 1 3 through the transmitted illumination optical system 1 4, and transmits the light transmitted through the specimen 1 2 to the Liporva 1 5. Light is collected with the mounted objective lens 16.
  • the light condensed by the objective lens 16 is imaged on the primary image surface 17 a via the imaging lens 18 of the imaging optical system 17 and the mirror M 1.
  • the image of specimen 1 2 imaged on the primary image plane 1 7 a is secondary through relay lens 1 7 b, mirror M 2, relay lens 1 7 c, and lens 1 9 a of eyepiece tube 1 9
  • the image is formed on the image plane 1 8 b and is observed by an observer through an eyepiece (not shown).
  • the fluorescent cube 21 in the fluorescent cube holder 20 shown in FIG. 1 is removed from the optical path.
  • the prism 2 2 is exchangeably arranged in the optical path, and is exchanged for a parallel plane plate of the same thickness when the transmission image of the sample 1 2 is observed.
  • the microscope apparatus 1 can be used as a transmission microscope.
  • a confocal scanning observation system 3 1 and an epi-illumination system 4 1 are arranged via a common illumination optical system 51.
  • the microscope apparatus 1 is used as a scanning microscope (scanning fluorescence microscope, confocal scanning microscope) will be described with reference to FIG.
  • the confocal scanning observation system 3 1 guides laser light from a laser light source (not shown) with an optical fiber 3 2, and laser light emitted from the end face of the optical fiber 3 2. Is made into a substantially parallel laser beam by the collector lens 3 3 and is incident on the two-dimensional scanner 3 4 that scans the sample 1 2 two-dimensionally.
  • the laser light emitted from the two-dimensional scanner 34 is focused on the image plane I P 1 by the pupil relay lens 35.
  • the laser beam emitted from the image plane IP 1 is converted into a laser beam substantially parallel to the optical axis by the imaging lens 52 of the illumination optical system 51, and is incident on the fluorescent cube 21 arranged in the optical path in an exchangeable manner. .
  • the dichroic mirror 1 4 4 that is detachably inserted in the optical path of the illumination optical system 51 used in the epi-illumination system 41 described later is used for illumination. It is removed from the optical path of the optical system 51.
  • the fluorescent cube 21 includes a wavelength selection filter 21a, a dichroic mirror 21b, and an emission filter 21c.
  • the laser light incident on the fluorescent cube 21 is selected by the wavelength selection filter 21 a and the Dyke mouth mirror 2 1 b, and is reflected in the direction of the objective lens 1 6. Incident on 6 and focused on specimen 1 2.
  • Fluorescence expressed in the sample 1 2 excited by the laser light is collected by the objective lens 16, enters the fluorescent cube 21, and the predetermined fluorescence is selected by the emission filter 21 in the fluorescent cube 2 1. Transparent.
  • the transmitted predetermined fluorescence is imaged on the image sensor 2 3 via the prism 2 2 detachably disposed on the imaging lens 18 and the imaging optical system 17, and the image sensor 2 3 emits fluorescence.
  • An image is taken.
  • the image captured by the image sensor 23 is subjected to image processing by the PC 3 shown in FIG. 1 and displayed on the monitor 3a.
  • the microscope apparatus 1 can be used as a scanning fluorescence microscope.
  • the laser beam reflected by the sample 1 2 is collected by the objective lens 16, reflected by the dichroic mirror 2 1 b of the fluorescent tube 2 1, and then reverses the illumination optical system 5 1.
  • the light is incident on the two-dimensional scanner 34, descanned, reflected by the beam splitter 36, and incident on the light receiving element 39 such as PMT through the imaging lens 37 and the pinhole 38.
  • a two-dimensional image is generated by PC 3 based on the intensity of each point received by light receiving element 39 and displayed on monitor 3a.
  • the microscope apparatus 1 can be used as a confocal scanning microscope.
  • the microscope apparatus 1 it is possible to observe the fluorescent image captured by the image sensor 23 and the confocal image received by the light receiver 39 by displaying them on the monitor 3a. .
  • the light from the light source (not shown) of the epi-illumination system 41 is guided by the optical fiber 42, and the light emitted from the end face of the optical fiber 42 is made almost parallel by the collector lens 43. Then, the light enters the dichroic mirror 44, which is detachably disposed on the illumination optical system 51 via the field stop 45. The light reflected by the dichroic mirror 4 4 is collected by the imaging lens 5 2 of the illumination optical system 51, and enters the fluorescent cube 21 that is arranged in the optical path as light substantially parallel to the optical axis. To do.
  • a light source not shown
  • a laser light source a high-pressure mercury lamp, a xenon lamp, or the like can be used.
  • the laser light incident on the fluorescent cube 21 is selected by the wavelength selection filter 21 a and the Dyke mouth Ic-mill 1 2 b, and is reflected in the direction of the objective lens 16. It enters 1 6 and is focused on sample 1 2.
  • Fluorescence expressed in the sample 1 2 excited by this light is collected by the objective lens 16, enters the fluorescent cube 21, and the predetermined fluorescence is emitted in the emission cube 21 of the fluorescent cube 21. Is selectively transmitted.
  • the transmitted predetermined fluorescence is imaged on the image sensor 23 via the imaging lens 18 and the prism 22 disposed in the imaging optical system 17 so as to be removable, and a fluorescent image is captured.
  • the image picked up by the image pickup device 23 is subjected to image processing by the PC 3 shown in FIG. 1 and displayed on the monitor 3a. In this way, the microscope apparatus 1 It can be used as an emission fluorescence microscope.
  • the illumination when the microscope apparatus 1 is used as a total reflection microscope uses the laser light of the confocal scanning observation system 31 described above. Further, a fluorescent cube 61 containing an optical member 60 to be described later for achieving total reflection illumination can be used as a total reflection microscope by exchanging and inserting it into the optical path.
  • laser light from a laser light source (not shown) of the confocal scanning observation system 31 is guided by an optical fiber 32, and laser light emitted from the end face of the optical fiber 32 is a collector lens. 3 3
  • the laser light is almost parallel and enters the two-dimensional scanner 3 4.
  • the tilt of each XY mirror of the two-dimensional scanner 34 is controlled by the control unit of the PC 3 shown in FIG.
  • the laser beam after the optical axis shift emitted from the two-dimensional scanner 34 is imaged on the image plane IP 1 by the pupil relay lens 35 and enters the optical path through the imaging lens 52 of the illumination optical system 51.
  • the light enters the arranged fluorescent cube 61.
  • the fluorescent cube 61 is composed of an optical member 60 composed of a wedge prism 62 and a condensing lens 63 such as a convex lens, a dichroic mirror 21 b, and an emission filter 21 c. It is built in the fluorescent cube holder 20 shown in FIG.
  • the wedge prism 6 2 and the condenser lens 6 3 are arranged such that the optical axis I 1 is shifted by a distance “d” from the optical axis of the illumination optical system 51 as shown in FIGS. 3 and 4B. .
  • This distance “d” corresponds to the position of NA which is the total reflection condition of the objective lens 16.
  • the laser light incident on the fluorescent cube 61 becomes laser light in which the principal ray is shifted by the distance “d” from the optical axis I 1 of the illumination optical system 51 by the wedge prism 62.
  • the light is collected by the lens 6 3 at the pupil position P of the objective lens 16 in the annular total reflection condition region.
  • the laser beam focused on the total reflection condition area is transmitted from the objective lens 16. It enters sample 1 2 at an incident angle that is totally reflected at the interface between sample 1 2 and the glass substrate that supports sample 1 2.
  • the laser light incident on the sample 12 at the total reflection angle generates an evanescent wave at the boundary surface, and fluorescence excited by the evanescent wave is generated near the boundary surface of the sample 12. Since the wavelength of the laser light is selected by a laser light source (not shown), the wavelength selection filter 21a shown in FIG. 2 is not necessary at this time.
  • Fluorescence expressed in the specimen 1 2 excited by the evanescent wave is collected by the objective lens 16, enters the fluorescent tube 61, and the predetermined fluorescence is emitted from the fluorescent cube 6 1 at 2 1 c. Is selectively transmitted.
  • the transmitted predetermined fluorescence is imaged on the image sensor 2 3 via the prism 2 2 detachably disposed in the optical path of the imaging lens 18 and the imaging optical system 17, and the image sensor 2 3 emits fluorescence.
  • An image is taken.
  • the image captured by the image sensor 23 is subjected to image processing by the PC 3 shown in FIG. 1 and displayed on the monitor 3a.
  • the microscope apparatus 1 replaces the fluorescent cube 21 described above with the fluorescent cube 61 and moves the optical axis I 1 of the laser light by a distance “d” from the optical axis of the illumination optical system 51 by a distance “d”. It can be used as a total reflection fluorescent microscope by moving it in parallel with the cana 3 4 and wedge prism 6 2. It is also possible to control the laser light to scan the annular total reflection condition region at the pupil position P of the objective lens 16 with the two-dimensional scanner 34 during total reflection illumination. By scanning the laser beam in the annular total reflection condition area, it is possible to perform satisfactory total reflection illumination.
  • Fig. 4A shows the state of illumination light when used as a scanning microscope or an epi-illumination microscope.
  • the objective lens in any case of the (+) maximum field angle light beam indicated by the broken line incident on the imaging lens 52, the image center light beam indicated by the solid line, and (1) the maximum field angle light beam indicated by the one-dot chain line At the pupil position P of the lens 16, it is a substantially parallel light beam that is not condensed.
  • the optical axis of the laser beam is illumination optics. It is shifted from the optical axis of system 51 by a distance “d” above the plane of the paper in FIG. 4B.
  • the laser light incident on the imaging lens 52 is incident such that the optical axis I 1 of the laser light is inclined by an angle with respect to the optical axis of the illumination optical system 51 as shown in FIG. 4C.
  • the wedge prism 6 2 sets the tilt angle ⁇ to be substantially zero so that the optical axis of the illumination optical system 51 and the optical axis I 1 of the laser beam are separated by a distance “d” and are substantially parallel to each other. .
  • the laser beam is condensed by the condenser lens 63 into the annular total reflection condition region at the pupil position P of the objective lens 16. As a result, total reflection illumination is possible.
  • ⁇ ( ⁇ 1 1) X ⁇ .
  • ⁇ is the refractive index of the medium constituting the wedge prism 62.
  • the wedge prism 6 2 having an apex angle ⁇ 5 corresponding to the inclination angle of the optical axis I 1 of the laser beam corresponding to the numerical aperture ( ⁇ ⁇ ) of the objective lens 16, and the condensing of the focal length f
  • the fluorescent cube 6 1 having the optical member 60 combined with the lens 63 is placed in the fluorescent cube holder 20 shown in FIG. 1, and is exchanged and inserted into the fluorescent cube 6 1 corresponding to the objective lens 16. It becomes possible to achieve total reflection illumination easily.
  • the movement amount “d” of the optical axis I 1 of the laser beam is also determined according to the objective lens 16 inserted in the optical path from the control unit of the PC 3 shown in FIG. By controlling the tilt of the laser beam, it is possible to set the optical axis position of the laser light according to the switching of the objective lens 16 and the fluorescent cube 61.
  • the laser beam is illuminated by controlling the two-dimensional scanner 3 4 of the confocal scanning observation system 3 1 to a predetermined inclination.
  • the fluorescent cube 61 placed in the fluorescent cube holder 20 that is configured to be able to exchange multiple fluorescent cubes in the optical path is exchanged and inserted into the optical path, enabling total reflection fluorescence observation can do.
  • FIG. 5 shows an optical system of a microscope according to the second embodiment.
  • FIG. 6 shows the optical system of the microscope apparatus when the optical system of the microscope apparatus according to the second embodiment is switched to a total reflection microscope.
  • symbol is attached
  • the usage method when the microscope apparatus 100 is used as a transmission microscope is the same as that in the first embodiment, and a description thereof will be omitted.
  • the confocal scanning observation system 3 1 guides laser light from a laser light source (not shown) with an optical fiber 3 2 and is emitted from the end face of the optical fiber 3 2.
  • the laser beam is converted into a substantially parallel laser beam by the collector lens 33 and is incident on the two-dimensional scanner 34 that scans the sample 12 two-dimensionally.
  • the laser beam emitted from the two-dimensional scanner 34 is imaged on the image plane I P 1 by the pupil relay lens 35.
  • the laser beam emitted from the image plane IP 1 is reflected by a dichroic mirror 71 1 that can be inserted into and removed from the optical path, collected by an imaging lens 18, and incident on the objective lens 16 as a substantially parallel laser beam. And collected on sample 1 2.
  • the fluorescent cubes 21 arranged so as to be exchangeable in the optical path are removed from the optical path.
  • Fluorescence expressed in the sample 12 excited by the laser beam is collected by the objective lens 16, and predetermined fluorescence is selectively transmitted through the dichroic mirror 72 and the emission filter 73.
  • the transmitted predetermined fluorescence is imaged on the image sensor 23 by the imaging lens 74, and a fluorescence image is captured by the image sensor 23.
  • the image captured by the image sensor 23 is subjected to image processing by the PC 3 shown in FIG. 1 and displayed on the monitor 3a. In this way, the microscope The device 101 can be used as a scanning fluorescence microscope.
  • the laser beam reflected by the specimen 12 is collected by the objective lens 16, travels back in the optical path, and enters the two-dimensional scanner 3 4 via the imaging lens 18 and the dichroic mirror 7 1. After being descanned, it is reflected by the beam splitter 36 and enters the light receiving element 39 such as PMT through the imaging lens 37 and the pinhole 38. Based on the intensity of each point received by the light receiving element 39, a two-dimensional image is generated by PC3 and displayed on monitor 3a.
  • the microscope apparatus 101 can be used as a confocal scanning microscope. When the specimen image is observed through the eyepiece tube 19 with the above-mentioned transmitted illumination, the dichroic mirrors 7 1, 7 2 and the fluorescent cube 2 1, or the fluorescent cube 8 1 described later, are in the optical path. Has been removed from.
  • the fluorescence image captured by the image sensor 2 3 and the confocal image received by the light receiving element 3 9 are displayed on the monitor 3 a shown in FIG. It is possible to observe.
  • the microscope apparatus 101 is used as an epifluorescence microscope
  • the confocal scanning observation system 3 1 and the epi-illumination system 4 1, except for a part, are arranged independently.
  • the epi-illumination system 4 1 guides light from a light source (not shown) by the optical fiber 4 2, makes light emitted from the end face of the optical fiber 4 2 light almost parallel by the collector lens 4 3, and The light is condensed by the imaging lens 52 through the stop 45 and is incident on the fluorescent cube 21 arranged in the optical path in a replaceable manner.
  • the fluorescent cube 21 includes a wavelength selection film 21a, a dichroic mirror 21b, and an emission filter 21c.
  • the laser light incident on the fluorescent cube 21 is selected by the wavelength selection filter 21 a and the Dyke mouth mirror 2 1 b, and is reflected in the direction of the objective lens 1 6. Incident on 6 and focused on specimen 1 2.
  • Fluorescence expressed in the sample 12 excited with this light is collected by the objective lens 16 and A predetermined fluorescence is selected by the dichroic mirror 7 2 and the emission filter 7 3, imaged on the image sensor 2 3 by the imaging lens 7 4, and a fluorescence image is captured by the image sensor 2 3.
  • the image captured by the image sensor 23 is subjected to image processing by the PC 3 shown in FIG. 1 and displayed on the monitor 3a.
  • the microscope apparatus 101 can be used as an epifluorescence microscope.
  • a light source not shown
  • a laser light source a high-pressure mercury lamp, a xenon lamp, or the like can be used.
  • the illumination when the microscope apparatus 10 1 is used as a total reflection microscope uses the above-described confocal scanning observation system 3 1, and fluorescent light containing an optical member 80 described later for achieving total reflection illumination.
  • the cube 81 can be used as a total reflection microscope by inserting it into the optical path.
  • the confocal scanning observation system 3 1 guides laser light from a laser light source (not shown) by an optical fiber 3 2 and collects laser light emitted from the end face of the optical fiber 3 2 as a collector lens 3. 3 makes the laser beam almost parallel and enters the two-dimensional scanner 34.
  • the inclination of each XY mirror of the two-dimensional scanner 34 is controlled by the control unit of PC 3 shown in FIG. .
  • the laser light emitted from the two-dimensional scanner 3 4 is imaged on the image plane I P 1 by the pupil relay lens 3 5.
  • the laser beam emitted from the image plane IP 1 is reflected by the dichroic mirror 71 and converted into a laser beam substantially parallel to the optical axis by the imaging lens 18, and is exchanged in the fluorescent cube 8 1 arranged in the optical path. Incident.
  • the fluorescent cube 81 is composed of an optical member 80 composed of a parallel flat glass 82 and a condensing lens 83 such as a convex lens arranged at a predetermined angle with respect to the optical axis, and is shown in FIG. It is built in the fluorescent cube holder 20 and is arranged so as to be exchangeable in the optical path.
  • the optical axis of the laser light emitted from the imaging lens 18 is shifted to the optical axis I 1 moved by a distance “d” from the optical axis by the parallel flat glass 82 inclined with respect to the optical axis. This The distance “d” corresponds to the position of NA which is the total reflection condition of the pupil position P of the objective lens 16.
  • the laser beam whose optical axis has been moved by the parallel plane glass 8 2 of the fluorescent cube 8 1 is condensed by the condenser lens 8 3 onto the annular total reflection condition region at the pupil position P of the objective lens 16. .
  • the laser beam focused on the total reflection condition region is incident on the sample 1 2 at an incident angle that is totally reflected from the objective lens 16 at the boundary surface between the sample 1 2 and the glass substrate supporting the sample 1 2. .
  • the laser light incident on the sample 12 at the total reflection angle generates an evanescent wave at the boundary surface, and fluorescence excited by the evanescent wave is generated near the boundary surface of the sample 12. Since the wavelength of the laser beam is selected by a laser light source (not shown), the wavelength selection filter 21 a shown in FIG. 5 is not necessary at this time.
  • the fluorescence expressed in the specimen 12 excited by the evanescent wave is collected by the objective lens 16 and selectively transmitted through the dichroic mirror 7 2 and the emission filter 7 3.
  • the transmitted predetermined fluorescence is imaged on the image sensor 23 by the imaging lens 74, and a fluorescence image is captured by the image sensor 23.
  • the image picked up by the image pickup device 23 is subjected to image processing by PC 3 shown in FIG. 1, and is displayed on the monitor 3a.
  • the microscope apparatus 101 replaces the above-described fluorescent cube 21 with the fluorescent cube 81 and changes the optical axis of the laser light so that it matches the optical axis of the confocal scanning observation system 31.
  • the three-dimensional scanner 34 it can be used as a total reflection fluorescent microscope.
  • a fluorescent cube 8 having an optical member 8 0 composed of a parallel plane glass 8 2 for shifting the optical axis in accordance with the numerical aperture (NA) of the objective lens 16 and a condensing lens 8 3 having a focal length f.
  • the two-dimensional scanner 34 of the confocal scanning observation system 31 is controlled so that the laser beam coincides with the optical axis.
  • the total reflection fluorescence observation can be performed.

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  • Physics & Mathematics (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Optics & Photonics (AREA)
  • Microscoopes, Condenser (AREA)

Abstract

L'invention vise à obtenir un microscope à fluorescence et à réflexion totale à partir d'un microscope confocal dont on a changé les cubes à fluorescence. Le microscope (1) comprend un système d'éclairage optique destiné à exposer un échantillon aux flux lumineux laser émis par des sources de lumière laser (32 et 42), un système optique de détection qui détecte la fluorescence de l'échantillon, une pluralité de cubes à fluorescence (61) disposée dans le chemin optique du système d'éclairage optique pour guider les flux lumineux laser vers l'échantillon, et une lentille d'objectif (16). Un ou plusieurs cubes à fluorescence comprennent des moyens optiques (60) grâce auxquels l'éclairage principal des flux lumineux laser est sensiblement parallèle à l'axe optique du système d'éclairage optique. Ces moyens optiques (60) regroupent les flux lumineux laser à un emplacement prédéterminé qui ne coïncide pas avec l'axe optique de l'emplacement de pupille (P) de la lentille d'objectif.
PCT/JP2008/055627 2007-04-11 2008-03-18 Microscope et cube à fluorescence à placer à l'intérieur WO2008126672A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
US12/563,141 US8014065B2 (en) 2007-04-11 2009-09-20 Microscope apparatus with fluorescence cube for total-internal-reflection fluorescence microscopy

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JP2007104054 2007-04-11
JP2007-104054 2007-04-11
JP2007-329046 2007-12-20
JP2007329046A JP5286774B2 (ja) 2007-04-11 2007-12-20 顕微鏡装置と、これに用いられる蛍光キューブ

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2006162994A (ja) * 2004-12-07 2006-06-22 Olympus Corp 全反射蛍光顕微鏡
JP2007072391A (ja) * 2005-09-09 2007-03-22 Olympus Corp レーザ顕微鏡

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2006162994A (ja) * 2004-12-07 2006-06-22 Olympus Corp 全反射蛍光顕微鏡
JP2007072391A (ja) * 2005-09-09 2007-03-22 Olympus Corp レーザ顕微鏡

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