JPH11231227A - Stereomicroscope - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a stereomicroscope permitting to observe weak fluorescence with a sharp contrast. SOLUTION: A fluorescent image of an excited specimen 4 is observed in a predetermined range of wavelengths passing through an absorption filter 16 by arranging a lighting path 12 formed out of an optical path split by dichroic mirror 11 in the path 1 of the two observation optical paths 1, 2 characterized by a seteromicroscope, and exciting the specimen 4 with the exciting light from a light source 14 of this lighting path 12. Therefore, this arrangement permits to eliminate influences by scattering of the exciting light and self- fluorescence generated from optical members, etc., and observe weak and faint fluorescence with a sharp contrast.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a stereo microscope used for observing a sample that emits fluorescence.

[0002]

2. Description of the Related Art Conventionally, in fluorescence observation of a specimen, a specific site in a cell or a tissue is stained with a fluorescent substance, and a site which emits fluorescence by the staining of the fluorescent substance is observed using a microscope. In this case, since the fluorescence emitted from the specimen is extremely weak and is observed at the cell level, observation with a stereoscopic microscope has been considered impossible due to the brightness of the observation target.

[0003] As an example of slightly performing fluorescence observation using such a stereomicroscope, as disclosed in JP-A-6-63164, laser irradiation is performed to specify an affected part during surgery. Some observe the autofluorescence emitted by tumor cells and the like.

[0004] On the other hand, such a stereomicroscope has a brightness stop on each of two observation optical axes, as disclosed in Japanese Patent Publication No. 4-3294. Some are arranged at the pupil position of the system, and are opened and closed in conjunction with the left and right to enable adjustment of the depth of focus and brightness of the observed image. And such 2
In a stereoscopic microscope observation that performs fluorescence observation with two observation optical axes, a specimen having a thickness like Drosophila may be used as it is, and in the case of a specimen having such a thickness, a specimen having a deep depth of focus is used. An image may be required.

[0005]

SUMMARY OF THE INVENTION However, Japanese Patent Laid-Open No.
Japanese Patent No. 63164 discloses a technique for irradiating a diseased part with a powerful laser beam by a dichroic mirror to specify a relatively rough position of a tumor, and removes autofluorescence of an optical member and a substance other than a target substance. Can not do
Fine and weak fluorescence cannot be observed with good contrast. In addition, the illumination optical system uses a high-power laser to ensure the brightness of the observation site without considering the utilization efficiency and the like, and therefore is expensive in terms of price.

By the way, GFP (Green Fl) which is a glowing protein recently discovered from Oan jellyfish
uresent Protein) has attracted attention. Such GFP emits green fluorescence when irradiated with excitation light in the same manner as a normal fluorescent substance, and this fluorescent light is very bright and does not accompany fluorescence-specific fading, allowing GFP to be expressed in a living organism. The possibility of observing it is being considered.

On the other hand, in the device disclosed in Japanese Patent Publication No. 4-3294, an image with a large depth of focus can be obtained by narrowing the brightness of the optical path. At the same time, the brightness of the image decreases. For this reason, when trying to take a photograph with a large depth of focus by coaxial epifluorescence observation, when the aperture stop is stopped down, the illumination light is also stopped down at the same time, so the fluorescent image becomes very dark and the exposure time becomes long. , It is more susceptible to the effects of film reciprocity failure and background noise,
A good photograph cannot be taken, and in some cases, there is a problem that recording such as photographing becomes impossible due to insufficient brightness.

In view of this, a stereoscopic microscope has been proposed in which a stop is arranged in a lens barrel or the like to avoid reducing the illumination light. Generally, when the aperture is located at another position, it is not located near the original pupil position, so if you stop down more than a certain amount, vignetting will occur, the periphery of the field of view will be dark, observation and photography Inconvenience occurs in recording such as Also, it is conceivable to relay the image with a lens in the shooting optical path, create a pupil position, and place an aperture, but even if a relay is unnecessary, a lens for the relay is always required, and a new aperture is required. Since it becomes necessary, the number of parts increases, the configuration becomes complicated, and the cost increases.

The present invention has been made in view of the above circumstances, and has as its object to provide a stereo microscope capable of observing weak fluorescence with good contrast. Another object of the present invention is to provide a stereoscopic microscope capable of observing and recording an observation image having a large depth of focus while maintaining the brightness of the observation optical path.

[0010]

According to the first aspect of the present invention,
Two observation optical paths through which a fluorescence observation image of the specimen is guided, an optical path dividing unit that divides one of the two observation optical paths, and an optical path dividing unit that is formed by the optical path division by the optical path dividing unit. An illumination optical path having a light source that excites the fluorescence of the sample, and filter means arranged in the two observation optical paths and transmitting a fluorescence observation image of a predetermined wavelength region of the sample excited by the light source.

According to a second aspect of the present invention, in the first aspect, an optical path reflecting means for reflecting a fluorescence observation image of the other observation optical path of the two observation optical paths, and the light is reflected through the optical path reflecting means. An imaging means for imaging the fluorescence observation image.

According to a third aspect of the present invention, there are provided two observation optical paths through which an observation image of a sample is guided, an optical path dividing unit for dividing one of the two observation optical paths, and an optical path dividing unit using the optical path dividing unit. And an illumination optical path having a light source that excites the fluorescence of the sample via the optical path splitting means, and is arranged in the two observation optical paths and transmits a fluorescence observation image of a predetermined wavelength range of the sample excited by the light source. Filter means, and aperture means provided separately for the two observation light paths, respectively, so that the respective aperture means of the two observation light paths can be opened and closed independently.

As a result, according to the first aspect of the present invention,
Being able to remove reflections of auto-fluorescence and illumination light outside the predetermined wavelength range of the fluorescence of the specimen excited by the light source, for example, optical members due to illumination light that excites fluorescence, and to observe minute and weak fluorescence with good contrast. Can be.

According to the second aspect of the present invention, a photographing optical path is provided in another observation optical path which is not affected by auto-fluorescence at all, such as an optical member by illumination light for exciting fluorescence, so that a high-quality photograph record or the like can be obtained. It can be performed.

According to the third aspect of the present invention, the aperture of the observation optical path having the illumination optical path is kept open while the aperture of the other observation optical path is stopped down as much as necessary, so that the brightness of the illumination on the specimen is reduced. Observed image with a large depth of focus can be obtained without affecting the image quality.

[0016]

Embodiments of the present invention will be described below with reference to the drawings. (First Embodiment) FIG. 1 shows a schematic configuration of a stereo microscope to which the present invention is applied. Here, only the optical systems arranged in two observation optical paths 1 and 2 of the stereo microscope are shown. ing.

In FIG. 1, reference numeral 3 denotes an objective lens common to the observation optical paths 1 and 2, and a specimen 4 taken in from the objective lens 3.
Are guided to the observation optical paths 1 and 2, respectively. These two observation optical paths 1 and 2 respectively have zoom lens groups 5 and 6 for performing zooming with a knob (not shown), imaging lenses 7 and 8 and eyepieces 9 and 10, and the specimens are provided by eyepieces 9 and 10. The four surfaces can be observed. In this case, the brightness diaphragms 51 and 61 are arranged at or near the observation optical path pupil position in the zoom lens groups 5 and 6.

As shown in FIG. 2, the aperture diaphragms 51 and 61 are attached to an aperture ring 512 (612) provided at the tip of the operation rod 511 (611) and the aperture diaphragm main body 513 (613).
By turning the aperture ring 512 (612) by a predetermined angle by operating the operation rod 511 (611) in the direction of the arrow shown in the figure, a predetermined opening state can be obtained in the aperture stop body 513 (613). Like that.

A dichroic mirror 11 is arranged between the zoom lens group 5 and the imaging lens 7 on the observation optical path 1.
The dichroic mirror 11 reflects light of a certain wavelength or less and transmits light of a longer wavelength, and forms an optical path divided from the observation optical path 1 by the dichroic mirror 11 in the illumination optical path 12. .

The illumination optical path 12 is a pump filter 1 composed of a band-pass filter capable of selecting only a specific wavelength.
3, for example, a light source 14 composed of a mercury lamp,
An illumination optical system 15 for guiding the light emitted from 4 to the excitation filter 13 is provided.

A long path that transmits only light of a certain wavelength or more is provided between the dichroic mirror 11 on the observation optical path 1 and the imaging lens 7 and between the zoom lens 6 and the imaging lens 8 on the observation optical path 2. Absorption filters 16 and 17 composed of filters are arranged.

In this case, it is assumed that the specimen 4 expresses GFP, is excited by illumination light in a wavelength range around 490 nm, and emits fluorescence having an intensity peak around 510 nm. As a result, the excitation filter 13 of the illumination light path 12 has a band having a characteristic of selectively transmitting only a wavelength range around 490 nm necessary for exciting the specimen 4 in which GFP is expressed, as shown in FIG. As shown in FIG. 3B, the absorption filters 16 and 17 are configured to transmit light having a wavelength of 505 or more, which transmits fluorescence having a peak of intensity around 510 nm emitted by GFP of the sample 4 as shown in FIG. As shown in FIG. 3C, the dichroic mirror 11 reflects illumination light in a wavelength range around 490 nm required for exciting the sample 4 and emits GFP of the sample 4 as shown in FIG. It has the property of transmitting fluorescence near 510 nm, which has a longer wavelength than light.

Next, the operation of the embodiment configured as described above will be described. Now, when illumination light is emitted from the light source 14, it is guided to the excitation filter 13 via the illumination optical system 15. Then, the illumination light only in the wavelength region near 490 nm required for exciting the sample 4 by the excitation filter 13 is transmitted and guided to the dichroic mirror 11. The dichroic mirror 11 reflects illumination light in a wavelength range around 490 nm, and transmits the reflected light to the zoom lens group 5 (brightness stop 5).
1) The sample 4 is illuminated through the objective lens 3. As a result, the GFP of the specimen 4 is excited by the illumination light in the wavelength range around 490 nm, and 510n having a longer wavelength than the excitation light.
Fluorescence having an intensity peak near m is emitted.

The fluorescence emitted from the sample 4 is condensed by the objective lens 3 and guided to the observation optical paths 1 and 2 as an observation image of the sample 4. As a result, the fluorescent light guided to the observation optical path 1 is guided to the dichroic mirror 11 through the zoom lens group 5 (brightness diaphragm 51), transmitted therethrough, and further transmitted to the 510 by the absorption filter 16.
Fluorescence having an intensity peak near nm is transmitted, is imaged by the imaging lens 7, and is observed by the eyepiece 9. On the other hand, the fluorescent light guided to the observation optical path 2 also passes through the zoom lens group 6 (brightness stop 61) and is absorbed by the absorption filter 1
The fluorescence having an intensity peak near 510 nm is transmitted therethrough, is imaged by the imaging lens 8, and is observed by the eyepiece 10.

Here, the aperture diaphragms 51 and 61 are usually
Although used in the open state, if the user wants to observe an image with a large depth of focus, the image obtained through the eyepiece 10 by narrowing the required amount of the aperture stop 61 on the observation optical path 2 becomes Observed as a deep observation image. In this case, the brightness of the illumination light from the light source 12 to the sample 4 is not affected at all by keeping the aperture stop 51 of the observation optical path 1 open.

Therefore, according to this configuration, an illumination optical path 12 formed by optical path division by a dichroic mirror 11 is provided in one of the two observation optical paths 1 and 2 which is a feature of the stereomicroscope. The sample 4 is excited by the excitation light from the light source 14 in the optical path 12, and among the excited fluorescent images of the sample 4, those in a predetermined wavelength range passing through the absorption filter 16 are observed. It is possible to remove the influence of auto-fluorescence generated from the light source and optical members, and it is possible to observe minute and weak fluorescence with good contrast.

In the first embodiment, a long-pass filter that transmits light of 505 nm or more is used as the absorption filters 16 and 17, but if a band-pass filter that transmits only light near 505 nm is used, G
Since most of the fluorescence other than the fluorescence emitted from the FP can be removed, the fluorescence can be further emphasized. In the above description, GF
Although the wavelength range of the filter is limited for P, the specimen stained with other fluorescent reagents and the specimen emitting autofluorescence can be selected by selecting an appropriate filter.
Similar effects can be expected. Further, brightness apertures 51, 6 provided together with the zoom lenses 5, 6
1 can be omitted. (Second Embodiment) FIG. 4 shows a schematic configuration of a second embodiment of the present invention, and the same parts as those in FIG. 1 are denoted by the same reference numerals.

In this case, the absorption filter 17 in the observation optical path 2
A beam splitter 18 is inserted between the imaging lens 8 and the observation optical path 2 so that the beam splitter 18 can be inserted into and removed from the observation optical path 2, and an optical path split by the beam splitter 18 is formed in a photographing optical path 19. A relay lens 20,
A total reflection mirror 21 for deflecting the imaging optical path 19 and an imaging lens 23 for forming an image on the imaging surface 22 are arranged.

Also in this case, the fluorescent light emitted from the specimen 4 is condensed by the objective lens 3 and guided to the observation optical paths 1 and 2 as the observation image of the specimen 4, respectively. Is guided to the zoom lens group 6
The light having the intensity peak near 510 nm is transmitted through the (brightness stop 61) to the absorption filter 17, and guided to the beam splitter 18. Then, the beam splitter 18 splits the observation optical path 2 and the imaging optical path 19 at a fixed ratio, and the light guided to the imaging optical path 19 is formed through a relay lens 20 and a total reflection mirror 21 into an imaging lens. 23 forms an image on the imaging surface 22.

As a result, the film or CC is
If an image sensor such as D is arranged, a fluorescent image of the specimen 4 is captured and recorded. In this case, the aperture diaphragms 51 and 61 are normally used in an open state. However, when an image having a large depth of focus is required at the time of recording such as photographing, the observation optical path 2 is used.
By stopping down the upper aperture stop 61 by a necessary amount, the image taken on the image-capturing optical path 19 divided from the observation optical path 2 by the beam splitter 18 is recorded as an image with a large depth of focus. In this case, the brightness diaphragm 51 of the observation optical path 1
Is left open, so that the brightness of the illumination light from the light source 12 to the specimen 4 is not affected at all.

In other words, in order to obtain an image having a large depth of focus during recording such as photographing, the brightness stop 6 on the observation optical path 2
Open and close only 1 and use the aperture 5 for the illumination light path
Since the illumination light guided to the specimen 4 can be used without opening, the illumination light guided to the specimen 4 is not lost, whereby sufficient illumination light is applied to the specimen 4 to secure the brightness, and the fluorescence of the observed image is reduced. It is possible to increase the depth of focus while maintaining the strength, minimize the extension of the exposure time, and take good photos without being affected by the characteristics of the film or the background. it can.

On the other hand, if it is not necessary to record and record a fluorescent image, the beam splitter 18 can be removed from the observation optical path 2 and observation with the 100% light amount through the eyepiece 10 can be performed in the same manner as described in the first embodiment. It becomes possible.

In the observation optical path 1, weak auto-fluorescence is emitted on the optical path of the zoom lens 5 and the objective lens 3 by the illumination light reflected by the dichroic mirror 11 through the excitation filter 13. When such autofluorescence is emitted, light having a wavelength transmitted through the absorption filter 16 is guided to the eyepiece lens 9, and the contrast of the fluorescent image of the specimen 4 may be reduced. Although the degree of the decrease in contrast is not uniform, it may be fatal for recording a photograph or the like. However, shooting optical path 1
The observation optical path 2 having 9 is not affected by the above-mentioned auto-fluorescence at all, and therefore does not cause a decrease in contrast. In other words, of the two observation optical paths 1 and 2, which are the characteristics of the stereomicroscope, by avoiding the optical path 1 used for excitation illumination and providing the photographing optical path 19 in the other observation optical path 2, it is completely affected by autofluorescence. And a fluorescent image with high contrast can be recorded.

In the second embodiment described above, the beam splitter 18 splits the photographing optical path 19 at a fixed ratio, but this ratio can be set arbitrarily. In some cases, a total reflection prism that reflects 100% toward the imaging optical path 19 may be used. (Third Embodiment) FIG. 5 shows a schematic configuration of a third embodiment of the present invention, and the same parts as those in FIG. 1 are denoted by the same reference numerals.

In this case, the dichroic mirror 11 and the absorption filter 16 in the observation light path 1, the excitation filter 13 in the illumination light path 12, and the absorption filter 17 in the observation light path 2 are provided integrally with the slide member 31, respectively. The slide member 31 is movable in a direction perpendicular to the optical axes of the observation optical paths 1 and 2, that is, in a direction perpendicular to the optical axis (front-back direction), and moves the dichroic mirror 11 and the absorption filter 16 to the observation optical path 1. And the excitation filter 13 is connected to the illumination light path 1.
2, the absorption filter 17 can be inserted into and removed from the observation optical path 2 at the same time.

In this manner, when the specimen 4 is to be observed by an observation method other than fluorescence, for example, when it is desired to view the entire image, the illumination is performed by an external illumination such as a fiber illumination or a fluorescent lamp (not shown), and the slide member 31 is connected to the optical path. The dichroic mirror 11 and the absorption filter 1
6. The excitation filter 13 in the illumination light path 12 and the absorption filter 17 in the observation light path 2 can be removed, so that a whole image observation without a filter can be performed.

Accordingly, in addition to the effects described in the first embodiment, if the absorption filters 16 and 17 and the dichroic mirror 11 are arranged in the respective observation optical paths 1 and 2, the colored observation In some cases, the observation image may be darkened due to a decrease in transmittance. However, it is possible to easily switch to bright-field observation with only one operation of removing the slide member 31 from the optical path.

The switching direction of the slide member 31 is
Since the direction is perpendicular to the optical axis (front-back direction), the sliding member 31 is moved in the lateral direction with respect to the optical axis, and compared with the case where the moving amount must be increased to avoid the optical axis.
Can be minimized, and space restrictions can be eliminated. This makes it easy to switch the slide member 31 not only in two stages of insertion and removal but also in three or more stages.

Further, when the specimen 4 is stained with a fluorescent dye other than GFP or when stained with two kinds of fluorescent dyes, each fluorescent dye is provided at each of the plurality of switching positions of the slide member 31. By arranging dichroic mirrors, excitation filters, and absorption filters having different characteristics adapted to each other, each fluorescent dye can be observed with good contrast only by switching the slide member 31. In this case, since the dichroic mirror, the excitation filter, and the absorption filter are simultaneously switched, quick and easy switching is possible. (Fourth Embodiment) FIG. 6 shows a schematic configuration of a fourth embodiment of the present invention, and the same parts as those in FIG. 1 are denoted by the same reference numerals.

In this case, instead of the illumination optical system 15 disposed between the light source 14 of the illumination optical path 12 and the excitation filter 13, the light source 14 is located at the pupil position of the optical system composed of the objective lens 3 and the zoom lens group 5. An illumination optical system 32 capable of projecting at a variable magnification is arranged.

Incidentally, recent microscopes employ a Koehler illumination optical system in order to uniformly illuminate the specimen surface in the observation field. When observing the specimen 4, the magnification of the zoom lens groups 5 and 6 is changed by moving the lenses of the zoom lens groups 5 and 6. At this time, the pupils of the zoom lens groups 5 and 6 are positioned in the optical axis direction. , The size of which changes, for example, decreases as the magnification increases and the magnification increases. On the other hand, the brightness of the illumination light emitted from the objective lens 3 to the specimen 4 is determined by how large the light source image is projected on the pupil of the objective lens 3. Therefore, in order to illuminate the specimen 4 with bright brightness, it is necessary to fill the pupils of the zoom lens group 5 and the objective lens 3 without waste, and if the size of the light source 14 is If the image is projected larger than the size and protrudes, only the light amount is wasted. For this reason, when the lens system of the illumination optical path 12 is fixed, the position and size of the projected image of the light source 14 are changed to any one of the position in the optical axis direction of the pupil and the change in size due to the change in zoom magnification. Must be set in advance, and it is impossible to perform optimum satisfaction at all zoom magnifications in this relation.

Therefore, in the fourth embodiment, the illumination optical system 32 is arranged in the illumination optical path 12, and the illumination optical system 32 is adjusted in accordance with the change in the pupil position and the size due to the change in the magnification of the zoom lens group 5. By making the focal length of the lens inside the lens variable, the position and size of the projected image of the light source 14 in the optical axis direction are changed so that the projected image is projected to the same size as the pupil. At this time, the NA of the light beam incident on the pupils of the zoom lens group 5 and the objective lens 3 is set to a value that can illuminate the entire observation visual field at each magnification.

Thus, in general, the NA of the objective lens is smaller and darker in the actual microscope than in the optical microscope.
In particular, in the case of fluorescence observation, bright illumination is required,
Even in such a case, it is possible to realize a bright, uniform and optimal illumination that always satisfies the pupil at any magnification in accordance with the zoom magnification of the zoom lens group 5. (Fifth Embodiment) FIGS. 7A and 7B show a specific configuration of the illumination optical system 32 described in the fourth embodiment, and the same parts as those in FIG. Is attached.

In this case, the illumination optical system 32 has constituent lenses 32a, 32b, and 32c.
2a, 32b, and 32c are connected to lens frames 33a, 33, respectively.
b, 34. Among them, the lens frame 34 is
It is fixed to the microscope main body 30, and the lens frames 33a, 33b
Allows the internal fitting portion of the frame 35 fixed to the lens frame 34 to be movable in the optical axis 36 direction. Also, the lens frame 33
Guide pins 37a and 37b are provided on the a and 33b, respectively. The guide pins 37a and 37b are guided along a long groove guide hole 38 formed in the frame 35, and are fitted to the outer peripheral portion of the frame 35. The guide is also guided by a rotatable cam 39. The cam 39 is formed with two cam grooves (not shown), each of which has a guide pin 3.
7a and 37b are guided.

A gear 40 is fixed to the cam 39 by a screw 41 so as to rotate integrally. Gear 40
, Another gear 42 is engaged with the other gear 4.
Numeral 2 rotates around a shaft 43 fixed to the microscope main body 30 by a fixing method (not shown). The other gear 42 is meshed with a gear 44 rotatably held by the microscope main body 30. A knob 45 is fixed to the distal end of the gear 44, and a rotation operation by the knob 45 is enabled. A moving mechanism (not shown) of the above-described zoom lens groups 5 and 6 is connected to the shaft 46 of the gear 44, and the knob 45 controls the shaft 46 via the gear 44.
By rotating, the zoom lens group 5,
Numeral 6 changes the zoom magnification.

In such a configuration, when the knob 45 is rotated, the gear 44 is rotated, and this rotation is transmitted to the moving mechanism (not shown) of the zoom lens groups 5 and 6 via the shaft 46, and the zoom magnification is changed. Done. At the same time, this gear 44
Is transmitted to the gear 40 via the rotation of the gear 42. When the gear 40 is rotated, the cam 39 is rotated, and the guide pins 37a and 37b are guided by the two cam grooves of the cam 39 and the long groove guide hole 38 of the frame 35, thereby moving the lens frames 33a and 33b. It is moved in the direction of the optical axis 36. In this case, the two cam grooves of the cam 39 optimize the projection magnification and position of the light source 14 in accordance with the change in the position and size of the pupil due to the change in the zoom magnification described in the fourth embodiment. Lens 32a, 32b
FIG. 7B shows a state in which the zoom is in the low magnification state shown in FIG.
Is moved to the zoom high magnification state shown in FIG.

In this way, the effect described in the fourth embodiment can be obtained simultaneously with zooming and zooming, and the viewer can change the zooming without worrying about the illumination. Even if the doubling is performed, optimal fluorescent illumination can always be performed. (Sixth Embodiment) FIG. 8 shows a schematic configuration of a sixth embodiment of the present invention, and the same parts as those in FIG. 1 are denoted by the same reference numerals.

In this case, the light source 14 is separated, the light source 14 is constituted by a light source main body 141 and a collector lens 142, and such a light source 14 is connected to a microscope via an optical fiber 71.

In this way, the light from the light source body 141 is condensed by the collector lens 142 and enters the illumination optical system 15 of the illumination optical path 12 via the optical fiber 71. Since the heat generated from the light source body 141 can be prevented from being transmitted to the microscope body side,
The thermal deformation on the microscope body side and the thermal effect on the microscopic person can be eliminated, and the use of the optical fiber 71 allows the specimen 4 to be removed.
The thermal effects on the air can also be eliminated. (Seventh Embodiment) FIG. 9 shows a schematic configuration of a seventh embodiment of the present invention, and the same parts as those in FIG. 1 are denoted by the same reference numerals.

In this case, the dichroic mirrors 11 and 11 'are arranged on the observation optical paths 1 and 2, respectively.
Illumination optical paths 12, 12 'having light sources 14, 14' and excitation filters 13, 13 'illumination optical systems 15, 15' are provided.

In this way, since the brightness is doubled by the light sources 14 and 14 ', an observation image of the specimen 4 which is illuminated more brightly can be obtained. (Eighth Embodiment) FIGS. 10 (a) and 10 (b) show a schematic configuration of an eighth embodiment of the present invention, and the same parts as those in FIG. 2 are denoted by the same reference numerals. . That is, in the eighth embodiment, the brightness stop 5 described in the first embodiment is used.
1 and 61 show different examples.

In this case, the aperture diaphragms 51 and 61 are provided with aperture rings 512 (6) provided at the tips of the operation sticks 511 (611).
12) has a brightness stop main body 513 (613). Each of these operation rods 511 (611) has an interlocking pin 514 (614).
14 (614) is movably mounted on the common interlocking operation plate 500 along the linear grooves 500a and 500b formed at predetermined intervals, and from this state, the interlocking operation plate 5
00 is slid in the arrangement direction of the aperture diaphragms 51 and 61, the operation rod 511 (611) is linked, and the aperture ring 512 (612) is rotated by a predetermined angle.
The same aperture state can be obtained in the aperture stop main body 513 (613).

The aperture diaphragms 51, 61 thus configured
In FIG. 4 described in the second embodiment, the interlocking operation plate 500 is attached when the beam splitter 18 is removed from the optical path and observation is performed with both eyes. By sliding the interlocking operation plate 500 and operating the operation rod 511 (611) by the same amount in the same direction via the interlocking pin 514 (614), the aperture ring 512 (61) is operated.
2) is rotated by a predetermined angle, and the aperture stop body 513 (61) is turned.
The same aperture state is set in 3), and a similar depth of focus and brightness can be obtained in the observation optical paths 1 and 2.

On the other hand, when taking a photograph, the beam splitter 18 is used as shown in FIG. 4 described in the second embodiment.
Is returned to the optical path, and by removing the interlocking operation plate 500 of the brightness diaphragms 51 and 61, the same operation as described in the second embodiment can be achieved.

In this way, at the time of observation with both eyes,
By opening and closing the left and right brightness apertures 51 and 61 in conjunction with each other, an aperture effect can be obtained at the same time on the left and right, so that high-precision observation can be easily obtained, operability is improved, and at the time of recording such as photographing. The same effect as that described in the second embodiment can be expected.

The present invention also includes the following inventions. (1) In claim 1, further comprising an integral member on which the optical path dividing means and the filter means are mounted, wherein the optical path dividing means and the filter means can be simultaneously inserted into and detached from the corresponding optical paths by moving the integral member. I have to.

In this way, by simply operating the integral member and simultaneously removing the optical path dividing means and the filter means from the optical path, it is possible to easily switch to the whole observation in the bright field. (2) The illumination optical system according to (1), wherein zoom lens groups are further arranged on the two observation optical paths, and a focal length is made variable according to a change in a pupil accompanying a change in magnification of the zoom lens groups. The system is located in the illumination beam path.

In this manner, the light source image can be projected onto the pupil in an optimum state at any zoom magnification in accordance with the change in the position and the size of the pupil accompanying the change in the magnification of the zoom lens group. (3) In the description of (2), the operation of changing the magnification of the zoom lens group and the operation of changing the focal length of the illumination optical system are performed in conjunction with each other.

With this arrangement, the projection onto the pupil can be set to an optimum state in conjunction with the zoom operation of the zoom lens group. (4) In the third aspect, the opening / closing operation of the aperture means of the two observation optical paths can be switched to an interlocking operation or an independent operation.

In this way, there is a case where the aperture is simultaneously operated on the left and right as in normal bright-field observation, and a case where it is desired to separately operate the aperture in the illumination system and the observation system for observation of a fluorescent image. Both can be used freely according to the application. (5) In the third aspect, among the aperture means provided separately in the two observation optical paths, the aperture means of the observation optical path having the illumination optical path is kept open.

With this configuration, the aperture means for the other observation optical path can be narrowed down as much as necessary while the aperture means for the observation optical path having the illumination optical path is kept open. A high-quality observation image with a large depth of focus can be obtained without any influence.

[0062]

As described above, according to the present invention, it is possible to remove auto-fluorescence other than the predetermined wavelength range of the fluorescence of the sample excited by the light source, for example, the optical member by the illumination light for exciting the fluorescence. Even minute and weak fluorescence can be observed with good contrast.

Further, since a photographing optical path is provided in another observation optical path that is not affected by autofluorescence at all, such as an optical member by illumination light for exciting fluorescence, a photograph is taken using this photographing optical path. Good quality photographic records can be realized.

Further, since the stop means of the other observation light path can be stopped down as much as necessary while keeping the stop means of the observation light path having the illumination light path open, there is no influence on the brightness of the illumination to the specimen. And a high-quality observation image with a large depth of focus can be obtained.

[Brief description of the drawings]

FIG. 1 is a diagram showing a schematic configuration of a first embodiment of the present invention.

FIG. 2 is a diagram illustrating a schematic configuration of a brightness stop used in the first embodiment.

FIG. 3 is a diagram illustrating characteristics of a filter and a dichroic mirror used in the first embodiment.

FIG. 4 is a diagram showing a schematic configuration of a second embodiment of the present invention.

FIG. 5 is a diagram showing a schematic configuration of a third embodiment of the present invention.

FIG. 6 is a diagram showing a schematic configuration of a fourth embodiment of the present invention.

FIG. 7 is a diagram showing a specific configuration of an illumination optical system used in a fifth embodiment of the present invention.

FIG. 8 is a diagram showing a schematic configuration of a sixth embodiment of the present invention.

FIG. 9 is a diagram showing a schematic configuration of a seventh embodiment of the present invention.

FIG. 10 is a diagram showing a schematic configuration of a brightness stop used in an eighth embodiment of the present invention.

[Explanation of symbols]

 1, 2 ... observation optical path, 3 ... objective lens, 4 ... sample, 5, 6 ... zoom lens group, 51, 61 ... brightness stop, 500 ... interlocking operation plate, 511, 611 ... operation rod, 512, 612 ... stop Ring, 513, 613: aperture stop body, 514, 614: interlocking pin, 7, 8: imaging lens, 9, 10: eyepiece, 11: dichroic mirror, 12: illumination optical path, 13: excitation filter, 14 ... Light source, 15: illumination optical system, 16, 17: absorption filter, 18: beam splitter, 19: imaging optical path, 20: relay lens, 21: total reflection mirror, 22: imaging surface, 23: imaging lens, 31: slide 32, illumination optical system, 32a, 32b, 32c: lens, 33a, 33b, 34: lens frame, 35: frame, 36: optical axis, 37a, 37b: guide pin, 3 8 ... long groove guide hole, 39 ... cam, 40 ... gear, 41 ... screw, 42 ... gear, 43 ... shaft, 44 ... gear, 45 ... knob, 46 ... shaft, 71 ... optical fiber.

Claims (3)

[Claims] 1. An observation optical path through which a fluorescence observation image of a specimen is guided, an optical path dividing means for dividing one of the two observation optical paths, and an optical path formed by the optical path division by the optical path dividing means. An illumination optical path having a light source that excites the fluorescence of the sample via the dividing unit; and a filter unit that is arranged in the two observation optical paths and transmits a fluorescence observation image of a predetermined wavelength range of the sample excited by the light source. A stereo microscope characterized by comprising: 2. An image forming apparatus according to claim 1, further comprising: an optical path reflection unit configured to reflect a fluorescence observation image of the other observation optical path of the two observation optical paths; and an imaging unit configured to capture the fluorescence observation image captured via the optical path reflection unit. The stereomicroscope according to claim 1, further comprising a photographing optical path. 3. An optical path split formed by two optical paths for guiding an observation image of a specimen, two optical paths for splitting one of the two optical paths, and an optical path split by the optical path splitting means. An illumination optical path having a light source that excites the fluorescence of the sample via a unit; a filter unit that is disposed in the two observation optical paths and transmits a fluorescence observation image of a predetermined wavelength range of the sample that is excited by the light source; A stereomicroscope comprising: aperture means provided separately for two observation light paths; and each of the aperture means of the two observation light paths can be independently opened and closed.