JP4895519B2 - Microscope equipment - Google Patents

Description

  The present invention relates to a microscope apparatus that enables time-resolved spectroscopy in a minute region under microscope observation.

As a conventional technique for performing time-resolved spectroscopy using a microscope, for example, there is one described in Patent Document 1 below.
JP-A-5-72480

The technique described in Patent Document 1 condenses pulsed laser light on an object to be detected and detects fluorescence from a minute point near the condensing position. Then, based on the time-varying characteristics of the fluorescence, energy diffusion (energy transfer) between molecules is measured.
According to the technique described in Patent Document 1, it is possible to analyze the environment around the fluorescent molecule by utilizing that the lifetime of the fluorescent molecule contained in the sample changes depending on the distance between the fluorescent molecules.

However, the conventional techniques have the following problems.
In a conventional time-resolving apparatus, the time domain that is a measurement target is about the lifetime of fluorescence. However, in general, the fluorescence lifetime of the fluorescent material is around nanoseconds. In such a time region, the fluorescence lifetime can be easily measured by a measuring instrument using an electronic device.
However, many physical properties that occur within a substance, such as chemical changes, occur in a shorter time domain. It has been difficult to measure the physical property changes occurring in these time regions using a conventional technique as described in Patent Document 1.

As an observation technique for measuring physical property changes occurring in a short time region, for example, there is pump-probe spectroscopy using an ultrashort light pulse. In this method, the sample is irradiated with pump light to give a stimulus to the sample, and the sample stimulated by the pump light is irradiated with probe light with a time difference from the pump light to probe changes in the physical properties of the sample due to the pump light. Detect from changes in light characteristics.
However, the intensity of the pump light must be stronger than that of the probe light in order to stimulate the sample. For this reason, when the pump light is included in the detection position of the probe light, the detection accuracy by the probe light becomes low.

  The present invention has been made in view of such a situation, and at the same time as observing a very small region, it is highly accurate to detect a physical property phenomenon that occurs in a very short time region (for example, femtoseconds to several tens of picoseconds). It is an object of the present invention to provide a microscope apparatus that can be easily grasped.

In order to achieve the above object, a microscope apparatus according to the present invention includes an optical microscope, a time-resolved spectroscopic unit, a first light guide unit that guides light from the time-resolved spectroscopic unit to the inside of the optical microscope, and the optical microscope. A second light guide means for guiding light into the time-resolved spectroscopic unit; the time-resolved spectroscopic unit; an ultrashort light pulse light source that oscillates an ultrashort light pulse; and the ultrashort light pulse as a reference light. A branching unit that branches into pump light and probe light; a multiplexing unit that combines the probe light and the reference light guided by the second light guiding unit; and a probe light and a reference light that reach a multiplexing position; A probe light-reference light time difference adjusting means for adjusting the time difference between the probe light and the reference light, and an image pickup device for picking up an interference fringe formed by combining the probe light and the reference light. Between elements , The light from the optical microscope which is guided by said second guiding means, wavelength axis direction while constituting the two-dimensional coordinates are deployed, the two-dimensional light waves time is expanded in the other coordinate axis A microscope apparatus in which a two-dimensional light wave conversion optical system for conversion is arranged, wherein the branching means includes pump light-probe light spatial separation means for spatially separating pump light and probe light; Pump light for adjusting the time difference between the pump light reaching the sample and the probe light-probe light time difference adjusting means, and further, reference light branching means for branching the reference light and other light, and the pump light- Probe light extraction means for extracting only the probe light from the light that has been separated through the probe light space separation means and passed through the sample in the optical microscope is provided.

  In the microscope apparatus of the present invention, only the reference light is extracted from the light separated through the pump light-probe light space separating means and branched to the reference light side through the reference light branching means. It is preferable to have a reference light extraction means.

  Further, in the microscope apparatus of the present invention, the pump light-probe light space separating means includes a transflective element configured by providing a transflective surface on both surfaces of a transparent parallel plate having a predetermined thickness, and A reflective element having a reflective surface that reflects light from one semi-transmissive reflective surface of the semi-transmissive reflective element toward the one semi-transmissive reflective surface, and light from the other semi-transmissive reflective surface of the semi-transmissive reflective element It is preferable to have a reflective element having a reflective surface that reflects the light toward the other semi-transmissive reflective surface.

  In the microscope apparatus of the present invention, the pump light-probe light space separating means reflects the semi-transmissive reflective element and one of the light transmitted through the semi-transmissive reflective element and the reflected light. A reflection element having two reflection surfaces leading to a semi-transmission reflection element, and a reflection element having one reflection surface for reflecting the other light from the semi-transmission reflection element toward the semi-transmission reflection element It is preferable.

  In the microscope apparatus of the present invention, it is preferable that the reflecting element having the two reflecting surfaces is two mirrors.

  In the microscope apparatus of the present invention, it is preferable that the reflecting element having the two reflecting surfaces is a right-angle prism having reflecting surfaces on two surfaces that form a right angle.

  In the microscope apparatus of the present invention, the pump light / probe light space separating means reflects the semi-transmissive reflective element and one of the light transmitted through the semi-transmissive reflective element and the reflected light. , A reflective element having one reflective surface that reflects the other light from the transflective element toward the transflective element, and a reflective surface that reflects the light reflected by the two mirrors And a prism having a transmission surface that transmits light reflected by the reflection element having the one reflection surface, and an optical axis of light reflected by the reflection surface of the prism and light emitted from the transmission surface of the prism It is preferable that the optical axis is parallel to the optical axis.

  Further, in the microscope apparatus of the present invention, the pump light-probe light space separation means is arranged on the incident optical axis of one of the transflective element and the light transmitted through and reflected from the transflective element. The lens includes a lens that is shifted relative to the lens, a reflective element that has one reflective surface that reflects light from the lens, and a single reflective element that reflects the other light from the transflective element. Is preferred.

  In the microscope apparatus of the present invention, it is preferable that the transflective element includes a single transflective surface.

  In the microscope apparatus of the present invention, it is preferable that the transflective element includes a transflective surface on both surfaces of a transparent parallel plate having a predetermined thickness.

  In the microscope apparatus of the present invention, it is preferable that the pump light / probe light space separating means includes a space separation amount adjusting means capable of adjusting a spatial separation amount of the pump light and the probe light.

  Further, in the microscope apparatus of the present invention, the space separation amount adjusting means is constituted by a moving means capable of moving the reflecting element having the two reflecting surfaces in a direction perpendicular to the incident optical axis of the one light. It is preferable.

  In the microscope apparatus of the present invention, the space separation amount adjusting means is a moving means capable of moving one of the two mirrors along the incident optical axis of the light to the one mirror. Preferably, it is configured.

  Further, in the microscope apparatus of the present invention, the space separation amount adjusting means is constituted by a moving means capable of moving the prism along the optical axis incident on the prism via the two mirrors. preferable.

  In the microscope apparatus of the present invention, the space separation amount adjusting means moves the lens shifted with respect to the incident optical axis of the one light perpendicularly to the incident optical axis of the one light. It is preferably composed of possible moving means.

  In the microscope apparatus of the present invention, the pump light-probe light time difference adjusting means reflects the light from one semi-transmissive reflective surface of the semi-transmissive reflective element toward the one semi-transmissive reflective surface. A reflective element having a surface and a reflective element having a reflective surface that reflects light from the other semi-transmissive reflective surface of the semi-transmissive reflective element toward the other semi-transmissive reflective surface. It is preferable that the moving means is configured to be movable along the incident direction of light from the surface.

  Further, in the microscope apparatus of the present invention, the pump light-probe light time difference adjusting means includes a transflective element having one transflective surface, and light transmitted through and reflected by the transflective element. A reflecting element having two reflecting surfaces that reflect one light and guide it to the transflective element, and one reflecting surface that reflects the other light from the transflective element toward the transflective element It is preferable that any one of the reflective elements having the above is constituted by a moving means capable of moving along the incident direction of light from the transflective surface.

  Further, in the microscope apparatus of the present invention, the pump light-probe light time difference adjusting means includes a transflective element having one transflective surface, and light transmitted through and reflected by the transflective element. A reflective element having two mirrors that reflect one light and guide it to the transflective element, and one reflective surface that reflects the other light from the transflective element toward the transflective element It is preferable that any one of the above is constituted by moving means that can move along the incident optical axis of the light from the transflective element.

  In the microscope apparatus of the present invention, the pump light-probe light time difference adjusting means is arranged to be shifted with respect to the incident optical axis of one of the light transmitted through the transflective element and the reflected light. Any one of a reflection element having a lens and one reflection surface that reflects light from the lens, and a reflection element having one reflection surface that reflects the other light from the semi-transmission reflection element is reflected by the semi-transmission reflection. It is preferable that the moving means is configured to be movable along the incident direction of light from the surface.

  In the microscope apparatus of the present invention, it is preferable that the probe light extraction means is constituted by a light shielding member having an opening through which only the probe light passes.

  Moreover, in the microscope apparatus of the present invention, the probe light extraction unit is further provided between the transflective element and a reflective element provided in an optical path that transmits the transflective element, or the transflective element. And a λ / 4 plate disposed between a reflection element provided in an optical path for reflecting the transflective element and a polarization disposed on a light-shielding member having an opening through which only the probe light passes. And a plate.

  In the microscope apparatus of the present invention, the probe light extraction unit further reflects the semi-transmissive light by reflecting one of the semi-transmissive reflective element, the light transmitted through the semi-transmissive reflective element, and the reflected light. A λ / 4 plate is provided between the reflection element having two reflection surfaces leading to the reflection element, and a polarizing plate is provided on the passage side of the light shielding member having an opening through which only the probe light passes. Is preferred.

  Further, in the microscope apparatus of the present invention, the probe light extraction means further includes a second opening that allows only the probe light to pass on a passage side of a light shielding member that includes an opening that allows only the probe light to pass. It is preferable to include a beam expander configured with a light shielding member interposed therebetween.

  In the microscope apparatus of the present invention, it is preferable that the reference light extraction unit is configured by a light shielding member having an opening through which only the reference light passes.

  According to the microscope apparatus of the present invention, a microscope apparatus capable of performing time-resolved spectroscopic measurement of probe light subjected to modulation in a femto to picosecond area with high accuracy simultaneously with observation of a minute area.

As a measurement technique using the high time resolution of the ultrashort light pulse, there is a technique described in Japanese Patent No. 3018173. According to this technique, it is possible to observe a high-speed physical property phenomenon in a femtosecond to picosecond region such as a chemical reaction.
The microscope apparatus of the present invention has been conceived in order to further increase the detection accuracy using the probe light after paying attention to the technique for measuring the time resolution.

The measurement technique of the time resolution will be described with reference to FIG.
FIG. 32 is a perspective view showing a schematic configuration of a two-dimensional light wave conversion optical system according to the waveform measurement technique of ultrashort light pulses. This figure is also described in Japanese Patent No. 3018173.
The two-dimensional light wave conversion optical system includes a beam expander 300, a diffraction grating 500, a first cylindrical lens 600, a filter 700, and a second cylindrical lens 800. The diffractive optical element 500 is a transmissive diffractive optical element. The diffraction grating 500 is disposed on the front focal plane (front focal position) of the first cylindrical lens 600. The filter 700 is disposed on the rear focal plane (rear focal position) of the first cylindrical lens 600. The position of the filter 700 coincides with the rear focal plane of the second cylindrical lens 800. The rear focal plane of the second cylindrical lens 800 and the front focal plane of the first cylindrical lens 600 are conjugate with each other.

A measurement process of time-resolved spectroscopy of a modulated ultrashort light pulse using the configuration of FIG. 32 will be described.
First, the incident light beam is enlarged by the beam expander 300 and obliquely incident on the diffraction grating 500. When the light flux at this time is viewed for each light beam, each light beam obliquely incident on the diffraction grating 500 does not reach the incident surface of the diffraction grating 500 at the same time. That is, regarding the x-axis direction of the diffraction grating 500, one end of the diffraction grating 500 is close to the beam expander 300 and the other end is far away from the beam expander 300. Therefore, there is a time difference between the light beam reaching the one end and the light beam reaching the other end. In other words, the time required for the light rays to reach differs depending on the position of the diffraction grating 500 in the x-axis direction. Therefore, here, time-resolved spectroscopy of the light beam that has reached the position along the line segment PQ in the figure will be considered.

  The diffraction grating 500 has a grating shape that diffracts incident light in the x-axis direction for each wavelength. Therefore, each wavelength component included in the light reaching the line segment PQ is diffracted in the x-axis direction at a different angle. Then, the light is condensed on the rear focal plane of the first cylindrical lens 600. At this time, since only the x-axis direction is condensed, light beams (light rays) elongated in the y-axis direction are arranged along the x-axis direction for each wavelength.

However, as shown in FIG. 33, the filter 700 includes a light blocking area and a light transmitting area. Here, the light transmission region is an opening. The shape of the opening is such that the y-axis direction increases as the x-axis direction increases. Since the area other than the opening is a light shielding area, the light is shielded.
For this reason, the light transmitted through the filter 700 has different wavelengths distributed in the y-axis direction with a time difference.

  Further, on the rear focal plane of the second cylindrical lens 800, the wavelength distribution in the y-axis direction is conserved. The rear focal plane of the second cylindrical lens 800 is conjugate with the front focal plane of the first cylindrical lens 600. Therefore, the position of the line segment P′-Q ′ conjugate with the position of the line segment PQ is conjugate. For this reason, as shown in FIG. 34, different wavelengths are arranged along the position of the line segment P′-Q ′.

Furthermore, the arrival time of the light beam varies depending on the position on the diffraction grating 500. Therefore, on the rear focal plane of the second cylindrical lens 800, as shown in FIG. 34, the wavelength is distributed in the y-axis direction and the time is changed in the x-axis direction in the arrow direction (left side in the figure). The resulting spectrogram will be generated. Hereinafter, this spectrogram is referred to as a two-dimensional light wave.
However, since the time change of the two-dimensional light wave is very fast, the normal image pickup device cannot capture the time change.
For this reason, reference light called a gate pulse is simultaneously irradiated onto the rear focal plane of the second cylindrical lens 800. By doing so, the spectrogram is acquired as an interference fringe pattern.

This two-dimensional lightwave conversion optical system enables time-resolved spectroscopy of light that has undergone some modulation by a sample, particularly ultrashort light pulses.
However, the two-dimensional light wave conversion optical system is a technique mainly used in the fields of optical communication and physical measurement.
The present applicant noticed that the above-described two-dimensional light wave conversion optical system is effective even in observation / measurement of a minute region, and came to the idea of using the two-dimensional light wave conversion optical system in a microscope apparatus. By doing so, it is possible to simultaneously observe a very small area and time-resolved spectroscopic measurement in the microscope apparatus.

However, the Applicant irradiates the sample with pump light for stimulating the sample, irradiates the sample stimulated by the pump light with probe light with a time difference from the pump light, and applies reference light to the probe light. The use of the above-described two-dimensional light wave conversion optical system was examined in a microscope apparatus configured to obtain an interference fringe pattern by overlapping.
The pump light has a higher light intensity than the probe light. For this reason, even in a microscope using the two-dimensional light wave conversion optical system, there is a problem that detection accuracy is adversely affected if pump light is included in the interference fringe formation position.
Therefore, the present applicant has come up with a configuration for spatially separating the pump light and the probe light as in the present invention.
According to the present invention, since the pump light after modulating the sample does not enter the detection position of the probe light, observation of a very small region and time-resolved spectroscopic measurement can be simultaneously performed with high accuracy.

FIG. 1 is a block diagram showing a configuration common to the microscope apparatus according to each embodiment of the present invention, and FIG. 2 is a schematic configuration diagram showing a configuration example of a microscope used in the microscope apparatus of FIG.
As shown in FIG. 1, the microscope apparatus according to each embodiment of the present invention includes a microscope 70 and a time-resolved spectroscopic unit 80. In addition, a first light guide unit 91 and a second light guide unit 92 are provided. The first light guide 91 guides the light from the time-resolved spectroscopic unit 80 to the inside of the microscope 70. Further, the second light guide unit 92 guides light from the microscope 70 to the inside of the time-resolving spectroscopic unit 80. As described above, the microscope apparatus is configured to perform time-resolved spectroscopy with the time-resolved spectroscopic unit 80 on the sample to be observed while simultaneously observing the sample with the microscope 70.

The time-resolved spectroscopic unit 80 includes an ultrashort light pulse light source 5, a branching unit 6, a probe light-reference light time difference adjusting unit 10, a two-dimensional light wave converting unit 1, a relay lens 2, a combining unit 3, An imaging device 4 is included.
The ultrashort light pulse light source 5 is configured to oscillate an ultrashort light pulse.
The branching unit 6 is configured to branch the ultrashort light pulse into pump light, probe light, and reference light.
The probe light-reference light time difference adjusting means 10 is configured to be able to adjust the optical path length difference between the probe light and the reference light. That is, when the imaging device 4 is irradiated with the probe light and the reference light via the multiplexing unit 3, the interference fringes are generated without the time difference to reach the multiplexing position.
The 1st light guide 91 is comprised by the mirror, for example. This mirror is fixed to a stage whose angle or position can be adjusted. By adjusting the angle or position of the mirror, the light emitted from the time-resolved spectroscopic unit 80 can be incident on the microscope 70.
Similarly, the second light guide unit 92 is configured by a mirror, for example. This mirror is also fixed to a stage whose angle or position can be adjusted. By adjusting the angle or position of the mirror, the light emitted from the microscope 70 can be made incident inside the time-resolving spectroscopic unit 80.
The first light guiding unit 91 and the second light guiding unit 92 may be provided inside either the microscope 70 or the time-resolving spectroscopic unit 80, or independent of the microscope 70 and the time-resolving spectroscopic unit 80. However, the arrangement location is not limited.

The two-dimensional light wave converting means 1 is configured to convert the light guided by the second light guiding means 92 into a two-dimensional light wave.
The relay lens 2 is configured to form an image of a two-dimensional light wave on the image pickup surface of the image pickup device 4 via the combining unit 3.
Further, the multiplexing means 3 is configured to reflect the reference light and irradiate the reference light onto the imaging surface of the imaging device 4. As a result, an interference fringe pattern is formed on the imaging surface of the imaging element 4.

Next, the microscope 70 will be described.
The microscope 70 may be an optical microscope that can observe the sample. For example, any configuration capable of observing bright field observation, fluorescence observation, differential interference observation, or the like may be used.

  In the configuration example shown in FIG. 2, the microscope 70 includes a transmission illumination light source 71, a transmission illumination optical system 72, an observation optical system 73, an observation imaging device 74, a sample stage 75, and a fluorescence illumination light source 76. A fluorescent illumination optical system 77, a filter unit 772, a beam splitter 723, and a dichroic mirror 78.

  The transmitted illumination optical system 72 includes a collector lens 721, a condenser lens 722, and a beam splitter 723. The transmitted illumination optical system 72 can illuminate the sample S with illumination light emitted from the transmitted illumination light source 71 and irradiate the sample with pump light and probe light via the beam splitter 723. In the configuration example of FIG. 2, the transmission illumination optical system 72 further includes a polarizer 724 and a first DIC (Differential Interference Contrast) filter 725.

The observation optical system 73 includes an objective lens 731, a second DIC filter 732, an analyzer 733, an imaging lens 734, and a beam splitter 735.
A dichroic mirror 78 and a filter unit 772 are disposed between the analyzer 733 and the imaging lens 734.
The filter unit 772 includes an excitation filter 7721, a dichroic mirror 7722, and an absorption filter 7723. The excitation filter 7721 is used to select excitation light for irradiating the sample S. The dichroic mirror 7722 reflects the excitation light to the sample S side and transmits the fluorescence generated from the sample S. The absorption filter 7723 blocks excitation light reflected from the sample S and transmits fluorescence.
In addition, the sample stage 75 includes an xy stage 752. Therefore, the relative position of the sample S with respect to the lenses 722 and 731 can be freely adjusted.

Next, a configuration unique to the present invention will be described in each embodiment.
(First embodiment)
FIG. 3 is an explanatory diagram showing the configuration of the branching means in the microscope apparatus according to the first embodiment of the present invention. The basic configuration of the entire microscope apparatus according to the first embodiment and the configuration of the microscope are the same as those shown in FIGS.
In the microscope apparatus of the first embodiment, the branching unit 6 includes a beam splitter 62, a mirror 63, a light attenuating element 630, a mirror 64, a fine movement stage 65, a beam splitter 723, and a spatial filter as a light shielding member. 66 and a spatial filter 67 as a light shielding member. In FIG. 3, reference numerals 912 and 911 denote mirrors constituting the first light guide unit 91.
The beam splitter 62 includes half mirror surfaces 62b and 62c on both surfaces of a transparent parallel plate 62a having a predetermined thickness. By providing the beam splitter 62 with this predetermined thickness, the beam splitter 62 emits incident light with a predetermined amount shifted by a refracting action.
The mirrors 63 and 64 are arranged perpendicular to the respective incident optical axes.
The fine movement stage 65 is configured to be able to move the mirror 64 in a direction along the optical axis incident on the mirror 64.

The ultrashort light pulse oscillated from the ultrashort light source 5 is branched into two via the beam splitter 62. One of the two branched lights is pump light and the other is probe light. Here, the pump light is light of an ultrashort light pulse that is transmitted through the half mirror surfaces 62 b and 62 c of the beam splitter 62 and reflected by the mirror 64. On the other hand, the probe light is an ultrashort light pulse light reflected by the half mirror surface 62 b of the beam splitter 62 and reflected by the mirror 63. In the microscope apparatus of the first embodiment, these pump light and probe light are used.
The fine movement stage 65 functions as a pump light-probe light time difference adjusting means in the present invention. Here, by adjusting the optical path length difference between the pump light and the probe light, it is possible to give a time difference for each light to reach the sample S (generate a time delay). In the example of FIG. 3, the pump light reaches the sample S before the probe light.

Further, the beam splitter 62 and the mirrors 63 and 64 function as pump light-probe light space separating means in the present invention.
That is, the light emitted from the ultrashort optical pulse light source 5 is divided into two through the half mirror surface 62 b of the beam splitter 62 into transmitted light and reflected light.
The transmitted light (light transmitted through the half mirror surface 62b) is refracted by the transparent parallel plate 62a having a predetermined thickness and reaches the half mirror surface 62c. The light transmitted through the half mirror surface 62c enters the mirror 64, is reflected by the mirror 64, follows an optical path in the reverse direction, and enters the half mirror surface 62c. The light reflected by the half mirror surface 62c is used as pump light. The pump light follows an optical path indicated by a broken line in FIG.
On the other hand, the reflected light (light reflected by the half mirror surface 62b) enters the mirror 63, is reflected by the mirror 36, travels in the opposite optical path, and enters the half mirror surface 62b. The light transmitted through the half mirror surface 62b is refracted by the transparent parallel plate 62a having a predetermined thickness and reaches the half mirror surface 62c. The light transmitted through the half mirror surface 62c is used as probe light. The probe light follows an optical path indicated by a solid line in FIG.
As a result, the pump light and the probe light are spatially separated in parallel.

The spatial filter 66 functions as probe light extraction means in the present invention. In other words, the spatial filter 66 allows only the probe light to pass through the opening 66a among the incident light separated in parallel and shields the other light.
The spatial filter 67 functions as reference light extraction means in the present invention. That is, the spatial filter 67 allows only reference light (pump light here) to pass through the opening 67a out of the incident light separated in parallel, and blocks other light (probe light here).

As described above, in the pump-probe method, the pump light is usually used as light for stimulating the sample, and the probe light is used as light for detecting the reaction of the sample after being stimulated by the pump light. use. Usually, the intensity of the pump light is strong to stimulate the sample, and the intensity of the probe light needs to be weak so as not to stimulate the sample.
For this reason, the light attenuating element 630 is provided between the beam splitter 62 and the mirror 63. The light attenuating element 630 is used to weaken the intensity of the probe light for irradiating the sample S.

In the microscope apparatus of the first embodiment configured as described above, the light emitted from the ultrashort light pulse light source 5 is converted into pump light and probe light via the beam splitter 62, mirror 63, mirror 64, and fine movement stage 65. To be separated. At this time, the pump light and the probe light are separated temporally and spatially, and both enter the beam splitter 723.
Pump light and probe light reflected by the beam splitter 723 are incident on the entrance pupil of the condenser lens 722. At this time, the pump light and the probe light are separated temporally and spatially and follow optical paths parallel to each other. In the example of FIG. 3, the pump light is shifted from the optical axis of the condenser lens 722 and the probe light is incident so as to coincide with the optical axis of the condenser lens 722. The pump light and the probe light are irradiated to the same minute region of the sample S via the condenser lens 722 while having a time difference. Further, after passing through the sample S, the pump light and the probe light are again spatially separated. Next, the pump light and the probe light become optical paths parallel to each other via the objective lens 731, and reach the spatial filter 66 via the second DIC filter 732, the analyzer 733, and the dichroic mirror 78. The pump light is shielded through the spatial filter 66, and only the probe light passes through the opening 66a. As a result, only the probe light reaches the two-dimensional lightwave means 1 shown in FIG.
On the other hand, the light corresponding to the pump light and the probe light transmitted through the beam splitter 723 reaches the spatial filter 67. Light corresponding to the probe light is blocked through the spatial filter 67, and only the light corresponding to the pump light passes through the opening 67a and is used as the reference light.

(First modification)
The branching means 6 in the microscope apparatus according to the first embodiment may be configured as shown in FIG. FIG. 4 is an explanatory diagram showing the configuration of a modification of the branching means 6 shown in FIG.
In the branching unit 6 of this modification, in the configuration of the branching unit 6 shown in FIG. 3, the λ / 4 plate 791 is disposed between the beam splitter 62 and the mirror 64, and the polarizer 792 is the spatial filter 66. It is arranged in the passing side optical path. The polarizer 792 is configured to shield the linearly polarized component of the pump light and transmit the linearly polarized component orthogonal to the linearly polarized component of the pump light. Other configurations are the same as those in FIG.
In the microscope apparatus using the branching means 6 of this modification configured as described above, the λ / 4 plate 791 and the polarizer 792 function as probe light extraction means that complements the spatial filter 66. That is, light that is emitted from the ultrashort optical pulse light source 5, passes through the half mirror surfaces 62 b and 62 c of the beam splitter 62, is reflected by the mirror 64, and is incident on the beam splitter 62 again, passes through the λ / 4 plate 791 twice. pass. For this reason, the linearly polarized light component is rotated by 90 degrees. Accordingly, the pump light that follows the optical path indicated by the broken line in FIG. 4 and the probe light that follows the optical path indicated by the solid line pass through the sample S with the linearly polarized light components orthogonal to each other, reflected by the dichroic mirror 78, and spatial. Only the probe light passes through the filter 66, and other light (pump light) is shielded.

By the way, as described above, since the pump light gives a stimulus to the sample S, it is necessary to make the light intensity much higher than that of the probe light. Here, most of the pump light is shielded through the spatial filter 66. However, a small amount of pump light may pass through the opening 66a of the spatial filter 66. On the other hand, the intensity of the probe light is weaker than that of the pump light.
For this reason, a slight amount of pump light that has passed through the opening 66a may adversely affect the detection of the probe light. Therefore, in order to prevent this, in the present modification, the linearly polarized light components of the pump light and the probe light are made to be orthogonal to each other, and the light after passing through the spatial filter 66 is made incident on the polarizer 792. Therefore, it is possible to more reliably block the pump light out of the light incident on the polarizer 792 and transmit the probe light. As a result, the pump light that could not be sufficiently removed only by the spatial filter 66 is sufficiently removed.
When the branching unit 6 is configured as in this modification, a λ / 2 plate is disposed on the optical path of the reference light that has passed through the spatial filter 67, and the linearly polarized component of the reference light is rotated by 90 °. Let As a result, the linearly polarized light components of the reference light and the probe light are combined in the same direction through the multiplexing means 3, and interference fringes can be generated. Although not shown, the polarization direction can be rotated by two mirrors instead of using the λ / 2 plate. Therefore, two mirrors may be newly added.

(Second modification)
Further, the branching means 6 in the microscope apparatus according to the first embodiment may be configured as shown in FIG. FIG. 5 is an explanatory view showing the configuration of another modification of the branching means 6 shown in FIG.
In the branching means 6 of this modification, the beam expander 793 is arranged on the passage side (light emission side) of the spatial filter 66 in the configuration of the branching means 6 shown in FIG. The beam expander 793 includes a lens 7931 and a lens 7932 on the spatial filter 66 side. Further, the beam expander 793 includes a spatial filter 794 as a second light shielding member. In the beam expander 793, light is collected inside. The spatial filter 794 is disposed at a position closer to the lens 7931 than the condensing position. Other configurations are the same as those in FIG.
In the microscope apparatus using the branching unit 6 of this modification configured as described above, the beam expander 793 functions as a probe light extraction unit that complements the spatial filter 66. That is, most of the pump light is blocked through the spatial filter 66, and the probe light and very little pump light pass through the opening 66a. Next, the probe light and very little pump light are collected via the lens 7931 of the beam expander 793 and reach the spatial filter 794. At this time, most of the probe light is focused on the optical axis of the lens 7931 and therefore passes through the opening 794a of the spatial filter 794. On the other hand, since the pump light which is off-axis light is condensed at a position away from the optical axis, it is shielded through the spatial filter 794. As a result, the pump light that could not be sufficiently removed only by the spatial filter 66 is sufficiently removed.

(Third Modification)
Further, the branching means 6 in the microscope apparatus according to the first embodiment may be configured as shown in FIG.
6 is an explanatory view of a main part showing the configuration of still another modification of the branching means 6 shown in FIG. 3, FIG. 7 is an enlarged view of the beam splitter 621 shown in FIG. 6, and FIG. 8 is a mirror shown in FIG. FIG.
The branching unit 6 of the present modification includes a beam splitter 621, a mirror 63, a light attenuating element 630, mirrors 641 and 642, a fine movement stage 643, and an optical path from the light source 5 to the light guiding unit 91. A prism 622 is provided. Fine movement stage 643 corresponds to fine movement stage 65 in the configuration of FIG.
In the microscope apparatus of this modification configured as described above, the ultrashort light pulse oscillated from the ultrashort light pulse light source 5 is branched into two via the beam splitter 621. One of the two branched lights is pump light and the other is probe light. Here, the pump light is ultrashort light pulse light that is transmitted through the surfaces 621 b and 621 c of the beam splitter 621 and reflected by the mirrors 641 and 642 and the prism 622. On the other hand, the probe light is an ultrashort light pulse light reflected by the surface 621 b of the beam splitter 621 and reflected by the mirror 63. The microscope apparatus of this modification uses these pump light and probe light.

The beam splitter 621 is configured by forming an antireflection film on one surface 621c of a transparent substrate 621a. Note that an antireflection film is not formed on the surface 621b opposite to the surface 621c.
The prism 622 includes a transparent member 622a having a parallelogram shape in cross section along the optical axis. A reflective film is deposited only on one surface 622c of the transparent member 622a. The prism 622 functions as a mirror for the pump light and functions as a prism for the probe light.

The light transmitted through the surface 621b of the substrate 621a of the beam splitter 621 is refracted by the substrate 621a and reaches the surface 621c. The light transmitted through the surface 621c is reflected by the mirrors 641 and 642, reflected by the reflecting surface 622c of the prism 622, and used as pump light as shown in FIG.
On the other hand, the light reflected by the surface 621b of the substrate 621a of the beam splitter 621 enters the mirror 63, is reflected by the mirror 63, follows the reverse optical path, and enters the surface 621b. The light transmitted through the surface 621b is refracted by the substrate 621a and reaches the surface 621c. The light transmitted through the surface 621c enters the surface 622b of the prism 622, passes through the surface 622b, is refracted by the transparent member 622a, and reaches the surface 622d. The light transmitted through the surface 622d is used as probe light as shown in FIG.
At this time, an antireflection film is formed on the surface 621c of the beam splitter 621. Therefore, the pump light-probe light that has reached the surface 621c can be transmitted with reduced light loss due to reflection.

The surfaces 622b and 622d through which the probe light in the prism 622 transmits may be provided with an antireflection film. This is preferable because the light amount loss due to the reflection of the probe light can be further reduced.
Further, the inclination of the prism 622 may be adjusted so that the probe light is incident on the surface 622b at an incident angle satisfying the Brewster angle. By doing in this way, the light quantity loss by reflection can be reduced.

(Fourth modification)
Further, as a modified example of FIG. 6, a prism 623 as shown in FIG. 9 may be used instead of the prism 622.
As shown in FIG. 10, the prism 623 includes a transparent member 623a having a trapezoidal cross section. A reflective film for reflecting the pump light is formed on the slope 623c of the transparent member 623a.
The other configuration is almost the same as the configuration of FIG.

In the third modification and the fourth modification, the light reflected by the prism 622 or the prism 623 becomes pump light. On the other hand, the light transmitted through the prism 622 or the prism 623 becomes probe light. Then, fine movement stage 643 in these modified examples moves mirror 642 in a predetermined direction. By doing in this way, the amount of space separation of pump light and probe light can be adjusted.
In the third and fourth modified examples, a fine movement stage is provided so that the prism 622 or the prism 623 is moved along the incident optical axis on the mirror 642 side instead of moving the mirror 642 along the incident optical axis. Also good.

(Second Embodiment)
FIG. 11 is an explanatory diagram showing the configuration of the branching means in the microscope apparatus according to the second embodiment of the present invention. The basic configuration of the entire microscope apparatus according to the second embodiment and the basic configuration of the microscope are the same as the configurations shown in FIGS.
In the microscope apparatus of the second embodiment, the branching unit 6 includes a beam splitter 61, a beam splitter 62 ', a mirror 63, a light attenuating element 630, a mirror 641, a mirror 642, a fine movement stage 643, and a fine movement stage. 65 and a spatial filter 66. In FIG. 11, reference numerals 912 and 911 denote mirrors constituting the first light guide unit 91. Reference numeral 922 denotes a mirror constituting the second light guide means.
The beam splitter 62 'is constituted by a thin plate-like half mirror surface 62a'.
The mirror 63 is disposed perpendicular to the incident optical axis.
The mirrors 641 and 642 are inclined with respect to the optical axis of incidence on the mirror 641 and are reflected by the mirror 641, and the optical axis of the light reflected by the mirror 642 is relative to the optical axis of incidence on the mirror 641. Arranged to shift quantitatively.
Fine movement stage 643 is configured to be able to move mirror 641 and mirror 642 in a direction perpendicular to the optical axis incident on mirror 641.
The fine movement stage 65 is configured such that the fine movement stage 643 on which the mirrors 641 and 642 are placed can be moved in a direction along the optical axis incident on the mirror 641.

The ultrashort light pulse oscillated from the ultrashort light source 5 is branched into two via the beam splitter 61. In the microscope apparatus of the second embodiment, the ultrashort light pulse reflected by the beam splitter 61 is used as the reference light. The ultrashort light pulse transmitted through the beam splitter 61 is further branched into two via the beam splitter 62 '. One of the two branched lights is pump light and the other is probe light. Here, the pump light is ultrashort light pulse light that is transmitted through the half mirror surface 62a ′ of the beam splitter 62 ′ and reflected by the mirrors 641 and 642. On the other hand, the probe light is an ultrashort light pulse light reflected by the half mirror surface 62 a ′ of the beam splitter 62 ′ and reflected by the mirror 63. In the microscope apparatus of this embodiment, these pump light and probe light are used.
The fine movement stage 65 functions as a pump light-probe light time difference adjusting means in the present invention. Here, by adjusting the optical path length difference between the pump light and the probe light, it is possible to give a time difference for each light to reach the sample S (generate a time delay). In the example of FIG. 11, the pump light reaches the sample S before the probe light.

Further, the beam splitter 62 ′ and the mirrors 63, 641, and 642 function as pump light-probe light space separating means in the present invention.
In other words, the light emitted from the ultrashort light pulse light source 5 and transmitted through the beam splitter 61 is divided into two by the transmitted light and the reflected light through the half mirror surface 62a ′ of the beam splitter 62 ′.
The transmitted light (light transmitted through the half mirror surface 62 a ′) enters the mirror 641 and is reflected by the mirrors 641 and 642. As a result, the light reflected by the mirror 642 and the optical path shifted in parallel with the optical path of the incident light are traced and incident on the half mirror surface 62a ′. The light reflected by the half mirror surface 62a ′ is used as pump light that follows an optical path indicated by a broken line in FIG.
On the other hand, the reflected light (light reflected by the half mirror surface 62a ′) is reflected by the mirror 63, follows an optical path in the opposite direction, and enters the half mirror surface 62a ′. The light transmitted through the half mirror surface 62a ′ is used as probe light that follows the optical path shown by the solid line in FIG.
As a result, the pump light and the probe light are spatially separated in parallel.

The fine movement stage 643 functions as a space separation amount adjusting means in the present invention.
That is, the mirror 641 and the mirror 642 are moved in a direction perpendicular to the optical axis incident on the mirror 641 via the fine movement stage 643. In this way, when light enters the mirror 641, the incident position of the incident light on the mirror 641 changes. Along with this, the interval between the optical axis of the light incident on the mirror 641 and the optical axis of the light reflected by the mirror 642 changes. As a result, the interval between the optical axis of the light reflected by the mirror 642 and reflected by the half mirror surface 62a ′ and the optical axis of the light reflected by the mirror 63 and transmitted through the half mirror surface 62a ′ changes.
Therefore, the spatial separation amount of the pump light and the probe light can be adjusted by adjusting the movement amount of the mirror 641 and the mirror 642 by the fine movement stage 643.

  In the branching means 6 of the first embodiment, the beam splitter 62 is a transparent parallel flat plate 62a having a predetermined thickness, and has half mirror surfaces 62b and 62c on both surfaces thereof. In this configuration, the incident light is emitted by being shifted by a predetermined amount due to the refracting action, so that the pump light and the probe light can be separated in space and can be propagated in parallel. However, in the case of the branching means 6 of the first embodiment, the spatial separation amount depends on the thickness of the parallel plate 62 a of the beam splitter 62 and is fixed. On the other hand, in the branching unit 6 of the second embodiment, the mirrors 641 and 642 are arranged so as to be inclined with respect to the optical axis of incidence on the mirror 641, and the mirror 641 and the mirror 642 are provided via the fine movement stage 643. Is moved in a direction perpendicular to the optical axis incident on the mirror 641. Thereby, the light reflected by the mirror 642 is shifted by a predetermined amount with respect to the light incident on the mirror 641. Therefore, in this embodiment, the amount of spatial separation between the pump light and the probe light can be finely adjusted.

  The spatial filter 66 functions as a probe light extraction unit in the present invention. In other words, the spatial filter 66 allows only the probe light to pass through the opening 66a among the incident light separated in parallel and shields the other light.

As described above, in the pump-probe method, the pump light is usually used as light for stimulating the sample, and the probe light is used as light for detecting the reaction of the sample after being stimulated by the pump light. use. Usually, the intensity of the pump light is strong to stimulate the sample, and the intensity of the probe light needs to be weak so as not to stimulate the sample.
For this reason, the light attenuating element 630 is provided between the beam splitter 62 ′ and the mirror 63. The light attenuating element 630 is used to weaken the intensity of the probe light for irradiating the sample S.

In the microscope apparatus according to the second embodiment configured as described above, the light emitted from the ultrashort light source 5 is divided into two via the beam splitter 61. The light reflected by the beam splitter 61 is used as reference light.
The light transmitted through the beam splitter 61 is separated into pump light and probe light via the beam splitter 62 ′, mirror 63, mirror 641, mirror 642, fine movement stage 643 and fine movement stage 65. At this time, the pump light and the probe light are separated spatially and temporally, and both enter the mirror 723.
Pump light and probe light reflected by the beam splitter 723 are incident on the entrance pupil of the condenser lens 722. At this time, the pump light and the probe light are separated temporally and spatially and follow optical paths parallel to each other. In the example of FIG. 11, the pump light is shifted from the optical axis of the condenser lens 722 and the probe light is incident so as to coincide with the optical axis of the condenser lens 722. The pump light and the probe light are irradiated to the same minute region of the sample S via the condenser lens 722 while having a time difference. Furthermore, after passing through the sample S, it is again spatially separated. Next, the pump light and the probe light become optical paths parallel to each other via the objective lens 731, and reach the spatial filter 66 via the second DIC filter 732, the analyzer 733, and the dichroic mirror 78. The pump light is shielded through the spatial filter 66, and only the probe light passes through the opening 66a. As a result, only the probe light reaches the two-dimensional lightwave means 1 shown in FIG.

In the microscope apparatus of the second embodiment, the beam splitter 62 ′ configured by the thin plate-like half mirror surface 62a ′ is used. However, the beam splitter 62 in the microscope apparatus of the first embodiment is used instead of the beam splitter 62 ′. May be used.
Further, instead of the mirrors 641 and 642, right angle prisms each having a reflecting surface on two surfaces that form a right angle may be used.
Also in the microscope apparatus of the second embodiment, similarly to the first modification of the microscope apparatus of the first embodiment, as shown in FIG. 12, the λ / 4 plate 791 is replaced with a beam splitter 62 ′ and a mirror 64. The polarizer 792 may be disposed in the optical path on the passing side of the spatial filter 66 while being disposed therebetween. The polarizer 792 is an element having a function of shielding the linearly polarized light component of the pump light and transmitting the linearly polarized light component orthogonal to the linearly polarized light component of the pump light.
Similarly to the second modification of the first embodiment, as shown in FIG. 13, a beam expander 793 provided with a spatial filter 794 may be arranged on the passage side of the spatial filter 66. .

(Third embodiment)
FIG. 14 is an explanatory diagram showing the configuration of the branching means in the microscope apparatus according to the third embodiment of the present invention. The basic configuration of the entire microscope apparatus according to the third embodiment and the basic configuration of the microscope are the same as those shown in FIGS.
In the microscope apparatus according to the third embodiment, the branching unit 6 includes a beam splitter 61, a beam splitter 62 ', a mirror 63, a light attenuating element 630, a lens 644, a mirror 643, a fine movement stage 643, and a fine movement stage. 643 and a spatial filter 66. Reference numerals 912 and 911 are mirrors constituting the first light guide unit 91. Reference numeral 922 denotes a mirror constituting the second light guide means.
The beam splitter 62 'is constituted by a thin plate-like half mirror surface 62a'.
The mirrors 63 and 64 are arranged perpendicular to the respective incident optical axes.
The lens 644 is arranged with a predetermined shift with respect to the optical axis incident on the lens 644. The lens 644 is configured by a normal spherical lens. Note that the lens 644 can also be formed of a cylindrical lens.
The fine movement stage 643 is configured to be able to move the lens 644 in a direction perpendicular to the optical axis incident on the lens 644.
The fine movement stage 65 is configured so that the fine movement stage 643 on which the lens 644 is placed is movable in a direction along the optical axis of incidence on the lens 644.

  The ultrashort light pulse oscillated from the ultrashort light source 5 is branched into two via the beam splitter 61. In the microscope apparatus according to the third embodiment, the ultrashort light pulse reflected by the beam splitter 61 is used as the reference light. The ultrashort light pulse transmitted through the beam splitter 61 is further split into two via the beam splitter 62 '. One of the two branched lights is pump light and the other is probe light. Here, the pump light is ultrashort light pulse light that is transmitted through the half mirror surface 62 a ′ of the beam splitter 62 ′, reflected by the mirror 64 through the lens 644. On the other hand, the probe light is an ultrashort light pulse light reflected by the half mirror surface 62 a ′ of the beam splitter 62 ′ and reflected by the mirror 63. In the microscope apparatus of this embodiment, these pump light and probe light are used.

Further, the beam splitter 62 ′, the mirror 63, the lens 644, and the mirror 64 function as a pump light-probe light space separating means in the present invention.
In other words, the light emitted from the ultrashort light pulse light source 5 and transmitted through the beam splitter 61 is divided into two by the transmitted light and the reflected light through the half mirror surface 62a ′ of the beam splitter 62 ′.
The transmitted light (light transmitted through the half mirror surface 62 a ′) enters the mirror 64 through the lens 644. The light reflected by the mirror 64 passes through the lens 644, follows an optical path shifted parallel to the optical path of the incident light, and enters the half mirror surface 62a ′. The light reflected by the half mirror surface 62a ′ is used as pump light that follows an optical path indicated by a broken line in FIG.
On the other hand, the reflected light (light reflected by the half mirror surface 62a ′) is reflected by the mirror 63, follows an optical path in the opposite direction, and enters the half mirror surface 62a ′. The light transmitted through the half mirror surface 62a ′ is used as probe light that follows the optical path indicated by the solid line in FIG.
As a result, the pump light and the probe light are spatially separated in parallel.

The fine movement stage 643 functions as a space separation amount adjusting means in the present invention.
That is, the lens 644 is moved in a direction perpendicular to the optical axis incident on the lens 644 via the fine movement stage 643. In this way, when light enters the lens 644, the incident position of the incident light changes. Accordingly, the distance between the optical axis of the light incident on the lens 644 and the optical path of the light reflected by the mirror 64 and emitted from the lens 644 changes. As a result, the distance between the optical path of the light emitted from the lens 644 and reflected by the half mirror surface 62a ′ and the optical axis of the light reflected by the mirror 63 and transmitted through the half mirror surface 62a ′ changes.
Therefore, the spatial separation amount of the pump light and the probe light can be adjusted by adjusting the moving amount of the lens 644 by the fine movement stage 643.
Other configurations are almost the same as those of the microscope apparatus of the second embodiment.

In the microscope apparatus according to the third embodiment configured as described above, the light emitted from the ultrashort optical pulse light source 5 is divided into two through the beam splitter 61. The light reflected by the beam splitter 61 is used as reference light.
The light transmitted through the beam splitter 61 is separated into pump light and probe light via the beam splitter 62 ′, mirror 63, lens 644, mirror 64, fine movement stage 643 and fine movement stage 65. At this time, the pump light and the probe light are separated spatially and temporally, and both enter the beam splitter 723.
Other operations are almost the same as those of the microscope apparatus of the second embodiment.

  In the microscope apparatus of the third embodiment, the beam splitter 62 ′ configured by the thin plate-like half mirror surface 62a ′ is used. However, the beam splitter 62 in the microscope apparatus of the first embodiment is used instead of the beam splitter 62 ′. May be used.

In the microscope apparatus of the third embodiment, as in the first modification of the microscope apparatus of the first embodiment, as shown in FIG. 15, a λ / 4 plate 791 is replaced with a beam splitter 62 ′, a mirror 64, and the like. The polarizer 792 may be disposed in the optical path on the passing side of the spatial filter 66. The polarizer 792 is an element having a function of shielding the linearly polarized light component of the pump light and transmitting the linearly polarized light component orthogonal to the linearly polarized light component of the pump light.
Similarly to the second modification of the first embodiment, as shown in FIG. 16, a beam expander 793 provided with a spatial filter 794 inside may be arranged on the passage side of the spatial filter 66. .

The following combination of the two-dimensional light wave conversion means 1, the relay lens 2, the multiplexing means 3, and the probe light-reference light time difference adjusting means 10 is applied to the branching means 6 of the first to third embodiments. Thus, the time-resolved spectroscopic unit 80 of the present invention can be configured. An example of the configuration is shown in FIGS.
FIG. 17 shows an example of the configuration of the combination of the two-dimensional light wave converting means 1, the relay lens 2, the multiplexing means 3, and the probe light-reference light time difference adjusting means 10 applicable to the branching means 6 of the first to third embodiments. 18 is an explanatory view showing the distribution of the probe light on the filter 141 in the configuration example of FIG. 17, FIG. 19 is an explanatory view showing the opening of the filter 141 in the configuration example of FIG. 17, and FIG. FIG. 21 is an explanatory diagram showing the two-dimensional light wave distribution generated by the two-dimensional light wave converting means 1 in the configuration example of FIG. 17, and FIG. 22 is a diagram showing the wavelength distribution of the probe light passing through the filter 141 in FIG. FIG. 23 is an explanatory view showing the diffraction grating array 15 used in the two-dimensional light wave converting means 1 in the configuration example of FIG. 17, and FIG. 23 shows tilt angles θ, φ with respect to the optical axes of the surfaces F3 ′ and F4 ′ in the configuration example of FIG. It is explanatory drawing which shows these relationships.

  In the configuration example of FIG. 17, the two-dimensional light wave conversion means 1 includes a beam expander 11, a diffraction grating 12, a first cylindrical lens 131 having a positive refractive power, a filter 141, and a first power having a positive refractive power. A two-cylindrical lens 132 and the diffraction grating array 15 are included. In FIG. 17, reference numerals 34 and 115 denote light attenuating elements.

The beam expander 11 is composed of rotationally symmetric lenses 111 and 112, and is configured to expand the beam diameter of the probe light. The beam expander 11 is arranged so that light is incident on the diffraction grating 12 obliquely.
The diffraction grating 12 is a transmission type diffraction grating. However, a reflection type diffraction grating may be used. The diffraction grating 12 is disposed in the vicinity of the F1 ′ plane inclined by a predetermined angle θ (here, 45 °) with respect to the normal line of the front focal plane F1 of the first cylindrical lens 131, and sequentially irradiates probe light. Diffraction in the x-axis direction.
The first cylindrical lens 131 distributes the wavelength component of the probe light diffracted by the diffraction grating 12 on the rear focal plane F2. The distribution state is shown in FIG.

  The filter 141 is disposed in the vicinity of the rear focal plane F2 of the first cylindrical lens 131. And as shown in FIG. 19, it has the elongate opening formed toward diagonally upward right from diagonally lower left. When the probe light is obliquely incident on the diffraction grating 12, the reflection irradiation time is shifted according to the incident position in the x-axis direction. The probe light thus obtained is composed of a plurality of wavelength components. Therefore, the filter 141 is used to extract each wavelength component for each position on the y-axis (vertical axis perpendicular to the optical axis (z-axis)). As a result, as shown in FIG. 20, filtering is performed so that different wavelengths are distributed in the y-axis direction. The probe light transmitted through the opening of the filter 141 enters the second cylindrical lens 132. At that time, the probe light is in a state where each wavelength is parallel to the xz plane and aligned in the y-axis direction.

  The light of each wavelength incident on the second cylindrical lens 132 forms an image on a surface F3 'near the rear focal plane F3 of the second cylindrical lens 132. Note that the position of the surface F3 'and the position of the diffraction grating 12 have a conjugate positional relationship. As a result, as shown in FIG. 21, the light of each wavelength is converted into a state (two-dimensional light wave) in which the time is developed in the x-axis direction and the wavelength is developed in the y-axis direction. That is, the probe light has (1) a continuous time delay for the light at each position in the x-axis direction, and (2) each wavelength constituting the probe light for the light at each position in the y-axis direction. Light is decomposed and distributed. Here, the two-dimensional light wave of the probe light is a time delay in the probe light continuously multiplexed in the x-axis direction, and the light of each wavelength constituting the probe light is decomposed in the y-axis direction. It means that it is in a distributed state.

The diffraction grating array 15 is disposed on the surface F3 ′. As shown in FIG. 22, diffraction gratings 15A to 15K having different grating constants (periods) for each wavelength are formed side by side in the y-axis direction. On the other hand, as shown in FIG. 21, in the two-dimensional light wave on the surface F3 ′ in the vicinity of the rear focal plane F3 of the second cylindrical lens 132, the light of each wavelength is spatially separated in the y-axis direction. For this reason, the diffraction directions of the respective wavelengths of the probe light can be aligned via the diffraction grating array 15. The traveling direction of light of each wavelength has an angular distribution (varies) in the x-axis direction. Therefore, the angular distribution in the x direction can be reduced by the diffraction grating array 15. That is, as shown in FIG. 21, since the two-dimensional light wave is developed with the wavelength spatially separated in the y-axis direction on the F3 ′ plane, the diffraction grating array in which the period is changed with respect to each wavelength. Can be used to align the propagation direction after diffraction of each wavelength.
In the configuration example of FIG. 17, the angular distribution of each wavelength after exiting the F3 ′ plane is configured to be substantially zero via the diffraction grating array 15.

The relay lens 2 includes a lens 21 and a lens 22, and forms a two-dimensional light wave image S4 on an imaging surface F4 ′ inclined by an angle φ with respect to the optical axis.
Here, the relationship between the inclination angles θ and φ with respect to the optical axis of the surfaces F3 ′ and F4 ′ will be described with reference to FIG.
The focal lengths of the lenses 21 and 22 of the relay lens 2 are f21 and f22, respectively, the length of the two-dimensional lightwave image S3 on the surface F3 ′ is L3, and the length of the two-dimensional lightwave image S4 on the surface F4 ′ is L4. To do. When the surface F3 ′ and the surface F4 ′ are in an imaging relationship, a point U where the extension of the lines along the surfaces intersects with the optical axis passing through the focal position L existing between the lens 21 and the lens 22. Exists on a vertical imaginary line. Therefore, the following relational expression (1) is satisfied.
2f21 tan (π / 2−θ) = 2f22 tan (π / 2−φ) (1)

Accordingly, the inclination angle φ of the surface F4 ′ and the two-dimensional light wave image size L4 on the surface F4 ′ are obtained as the following equations (2) and (3), respectively.
tan φ = f22 / f21 tan θ = M tan θ (2)
M = f22 / f21 (3)
L4 = ML3 cos θ / cos θ (4)
Here, M represents the magnification of the relay lens 2.
From equation (2), the following (2-1) and (2-2) can be said.
When M> 1, φ> θ (2-1)
When M <1, φ <θ (2-2)
That is, if the relay lens 2 has a reduction magnification, the tilt angle φ of the F4 ′ surface with the two-dimensional light wave image S4 is smaller than the tilt angle 45 ° of the F3 ′ surface with the two-dimensional light wave image S3. It is possible to reduce the influence of shading in the image sensor 4 for decomposition spectroscopy.

When the two-dimensional light wave image S4 is formed on the F4 ′ plane, simultaneously, the reference light is irradiated onto the imaging plane F4 via the multiplexing unit 3. Then, interference by the two-dimensional light wave image S4 and the reference light occurs on the imaging surface F4, and the spectrogram of the probe light is generated as an interference fringe pattern. Thereby, the time change of the wavelength component contained in the probe light can be measured.
The probe light-reference light time difference adjusting means 10 includes mirrors 101, 102, and 103 and a fine movement stage 104. Then, the mirrors 101 and 102 fixed to the fine movement stage 104 can be moved in the direction of the incident optical axis of the mirror 101. By this movement, the optical path lengths of the probe light and the reference light can be adjusted to be substantially equal.
The multiplexing means 3 includes a beam expander 33, a beam splitter 31, and a lens 32. Then, the reference light branched by the branching unit 6 is irradiated in a parallel light state onto the time-resolved spectral imaging element 4 through the lens 22.

  FIG. 24 shows another configuration of the combination of the two-dimensional light wave converting means 1, the relay lens 2, the multiplexing means 3, and the probe light-reference light time difference adjusting means 10 applicable to the branching means 6 of the first to third embodiments. FIG. 25 is an explanatory view showing the distribution of the probe light on the first diffraction grating array 142 in the configuration example of FIG. 24, and FIG. 26 is used for the two-dimensional light wave converting means 1 in the configuration example of FIG. 27 is an explanatory view showing the first diffraction grating array 142, and FIG. 27 is an explanatory view showing probe light diffracted by the first diffraction grating array 142 in the configuration example of FIG.

  In the configuration example of FIG. 24, the two-dimensional light wave converting means 1 includes a cylindrical beam expander 11 ′, a diffraction grating 12, a first lens 133 having positive refractive power, a first diffraction grating array 142, a positive The second lens 134 having refractive power and the second diffraction grating array 15 are configured. In FIG. 24, 34 and 115 are light attenuating elements.

The cylindrical beam expander 11 ′ is composed of rotationally symmetric lenses 111 and 112 and cylindrical lenses 113 and 114, and is configured to expand the beam diameter of the probe light. The cylindrical beam expander 11 ′ is arranged so that light is incident obliquely on the diffraction grating 12.
The diffraction grating 12 is a transmission type diffraction grating. However, a reflection type diffraction grating may be used. The diffraction grating 12 is disposed in the vicinity of the F1 ′ plane inclined by a predetermined angle θ (here, 45 °) with respect to the normal line of the front focal plane F1 of the first lens 133, and sequentially irradiates the probe light with x Diffraction in the axial direction.
The first lens 133 distributes the wavelength component of the probe light diffracted by the diffraction grating 12 on the rear focal plane F2. The state of distribution is shown in FIG.

  The first diffraction grating array 142 is a filter, and is disposed in the vicinity of the rear focal plane F <b> 2 of the first lens 133. As shown in FIG. 26, diffraction gratings 142a to 142k having different lattice constants (periods) are formed side by side in the x-axis direction. Thereby, as shown in FIG. 27, each wavelength of the probe light is diffracted at different diffraction angles in the direction along the y-axis via the diffraction gratings 142a to 142k. The light of each wavelength diffracted by the first diffraction grating array 142 enters the second lens 134.

The light of each wavelength incident on the second lens 134 forms an image on a surface F3 ′ in the vicinity of the rear focal plane F3 of the second lens 134. The position of the surface F3 ′ and the position of the diffraction grating 12 have a conjugate positional relationship. As a result, as shown in FIG. 21, it is converted into a two-dimensional light wave in which time is developed in the x-axis direction and wavelength is developed in the y-axis direction.
That is, the two-dimensional light wave of the probe light here also has a time delay in the probe light continuously multiplexed in the x-axis direction, and light of each wavelength constituting the probe light in the y-axis direction. It means that it is in a state of being decomposed and distributed.

The second diffraction grating array 15 is disposed on the surface F3 ′. As shown in FIG. 22, diffraction gratings 15A to 15K having different grating constants (periods) for each wavelength are formed side by side in the y-axis direction. On the other hand, as shown in FIG. 21, in the two-dimensional light wave on the surface F3 ′ in the vicinity of the rear focal plane F3 of the second lens 134, the light of each wavelength is spatially separated in the y-axis direction. For this reason, the diffraction directions of the respective wavelengths of the probe light can be aligned via the second diffraction grating array 15. The traveling direction of light of each wavelength has an angular distribution (varies) in the x-axis direction. Thus, the second diffraction grating array 15 can reduce the angular distribution in the x direction. That is, as shown in FIG. 21, since the two-dimensional light wave is developed with the wavelength spatially separated in the y-axis direction on the F3 ′ plane, the diffraction grating array in which the period is changed with respect to each wavelength. Can be used to align the propagation direction after diffraction of each wavelength.
In the configuration example of FIG. 24, the angular distribution of each wavelength after exiting the F3 ′ plane is configured to be substantially zero via the second diffraction grating array 15.

  Other configurations and operations are the same as those of the configuration example of FIG. That is, even in this configuration example, the micro area of the sample S can be irradiated with the pump light and the probe light under the microscope observation, and the micro area is stimulated with the pump light. Then, only the probe light modulated by the minute region is converted into a spectrogram developed in time and wavelength. By doing so, even in this configuration example, time-resolved spectroscopy in a minute region is possible.

FIG. 28 is an explanatory diagram showing a modification of the two-dimensional light wave converting means 1 in the configuration example of FIG.
Also in the configuration example of FIG. 28, the two-dimensional light wave conversion means 1 includes the diffraction grating 12 and the diffraction grating arrays 142 and 15 in the configuration example of FIG. However, the diffraction grating 12 is provided with a parallel plate 121 on the surface on which the diffraction grating is formed. Similarly, the diffraction grating array 142 is provided with a parallel plate 1421 on the surface where the diffraction grating is formed. The diffraction grating array 15 is provided with a parallel plate 151 on the surface on which the diffraction grating is formed. Other configurations are substantially the same as the configuration example of FIG.
In general, when dust adheres to the diffraction grating, it is almost impossible to clean the lens like a lens, and there is no method other than removing the adhered dust using a wind blower. However, if configured as shown in FIG. 28, it is possible to prevent dust from adhering to the diffraction grating via the parallel plate.

  In addition, in the configuration example of FIG. 17, in the diffraction gratings 12 and 15, a parallel plate may be arranged on the surface on the side where the diffraction grating is formed. By doing so, the effect of preventing dust from adhering to the diffraction grating can be obtained as in the configuration example of FIG.

FIG. 29 is an explanatory diagram illustrating the overall configuration of the microscope apparatus according to the first embodiment of the present invention. The basic configuration is the same as the configuration in FIGS. 1 and 2, and the sample is observed through the microscope 70 and the time-resolved spectroscopic unit 80 performs time-resolved spectroscopy on a specific region in the sample. Has been.
And in the microscope apparatus of Example 1, the structure of 1st Embodiment shown in FIG. Further, the two-dimensional light wave converting means 1, the relay lens 2, the multiplexing means 3, and the probe light-reference light time difference adjusting means 10 employ the configurations shown in FIGS.

The ultrashort light pulse light source 5 is a femtosecond laser that oscillates ultrashort light having a center wavelength of 800 nm, a wavelength width of ± 5 nm, and a pulse width of 100 femtoseconds.
The branching unit 6 includes a beam splitter 62, a mirror 63, a light attenuating element 630, a mirror 64, a fine movement stage 65, a beam splitter 723, a spatial filter 66, and a spatial filter 67. ing.
Then, the light emitted from the ultrashort light pulse light source 5 is divided into a transmitted light and a reflected light through a beam splitter 62. The bisected light is reflected by the mirrors 63 and 64 and enters the beam splitter 62 again. The light reflected by the mirror 63 and transmitted through the beam splitter 62 and the light reflected by the mirror 64 and reflected by the beam splitter 62 are incident on the beam splitter 723 via the light guide unit 91. The light reflected by the beam splitter 723 irradiates the sample S. At this time, one of the lights divided by the beam splitter 62 (probe light) is given a time delay to the other (pump light) via the fine movement stage 65. For this reason, the probe light is modulated by the sample S stimulated by the pump light.

In this embodiment, the probe light is light that is reflected by the beam splitter 62, reflected by the mirror 63, and further transmitted through the beam splitter 62. On the other hand, the pump light is light that has been transmitted through the beam splitter 62, reflected by the mirror 64, and further reflected by the beam splitter 62. The pump light and the probe light are separated spatially in parallel by being refracted by the parallel plate 62 a when passing through the beam splitter 62.
The spatial filter 66 passes only the probe light out of the light separated in parallel after passing through the sample S, and blocks the pump light. As a result, only the probe light is incident on the two-dimensional light wave conversion means 1.
Further, the two lights separated in parallel spatially pass through the beam splitter 723. The spatial filter 67 allows only light corresponding to the pump light out of the transmitted light to pass therethrough and blocks light corresponding to the probe light. The light that has passed through the spatial filter 67 is used as reference light.

  The two-dimensional light wave conversion means 1 includes a beam expander 11, a Bragg diffraction grating 12, a cylindrical lens 131, a filter 141, a cylindrical lens 132, and a diffraction grating array 15. The beam expander 11 includes a lens 111 having a focal length of 10 mm and a lens 112 having a focal length of 100 mm. Cylindrical lenses 131 and 132 are lenses each having a focal length of 100 mm.

The flag type diffraction grating 12 is disposed on a surface F1 ′ obtained by rotating the front focal plane F1 of the cylindrical lens 131 shown in FIG. 17 by about 45 °, and the angle between the normal line and the optical axis AX is about 45. °. A plane conjugate with the plane F1 ′ with respect to the cylindrical lenses 131 and 132 is a plane F3 ′ when the rear focal plane F3 of the cylindrical lens 132 is rotated by 45 °. For this reason, the conjugate plane F3 ′ is also at an angle of approximately 45 ° with respect to the optical axis AX.
Here, the Bragg diffraction grating 12 has a lattice constant of 1767 lines / mm, and is selected so that the axis of the propagation direction of the central wavelength of the probe light incident at 45 ° substantially coincides with the optical axis AX.

  Further, in the two-dimensional light wave conversion means 1 of the first embodiment, the probe light is enlarged approximately 10 times via the beam expander 11. The Bragg diffraction grating 12 has a size of about 14.14 mm × 10 mm. Probe light is obliquely incident on such a Bragg diffraction grating 12. At this time, the probe light has a wavelength width of 800 ± 5 nm. Therefore, on the rear focal plane F2 of the cylindrical lens 131, the wavelength component contained in the probe light is distributed in the x-axis direction as shown in FIG. At this time, for example, wavelengths of 805 nm, 800 nm, and 795 nm are distributed at positions of +1.26 mm, 0 mm, and −1.24 mm, respectively, with respect to the optical axis AX.

  Since the filter 141 has an opening as shown in FIG. 19, the wavelength component of the probe light is distributed in the y-axis direction on the F3 ′ plane. When the probe light is sequentially irradiated onto the Bragg diffraction grating 12 on the F1 ′ plane, the wavelength distribution moves in the x-axis direction on the F3 ′ plane conjugate with the F1 ′ plane. As a result, a two-dimensional light wave S3 that changes with time in the x-axis direction and expands in wavelength on the vertical axis is generated on the F3 'plane (FIG. 21). The size of the two-dimensional light wave S3 on the F3 ′ plane is 14.14 × 10 mm.

  The diffraction grating array 15 corrects the angular distribution generated for each wavelength of light with respect to the two-dimensional light wave S3 incident on the surface F3 ′. The following table shows the specifications of the diffraction grating array 15. It is shown in 1. table. The incident angle of 1 is an angle with respect to the normal line of the diffraction grating array 15 when light of each wavelength enters the surface F3 ′. The diffraction gratings 15A to 15K constituting the diffraction grating array 15 are shown in FIG. With a periodic structure such as 1, the diffraction angle for light of each wavelength is 45 °. As a result, the angular distribution of light of each wavelength after exiting from the surface F3 ′ is almost zero.

table. 1 Specifications of diffraction grating array 15

  The relay lens 2 includes a lens 21 having a focal length f21 = 100 mm and a lens 22 having a focal length f22 = 25 mm. The two-dimensional light wave image S4 is distributed on the imaging surface by causing the imaging surface of the imaging device 4 for time-resolved spectroscopy to coincide with the surface F4 'conjugate with the surface F3'. Since the surface F3 ′ is inclined by 45 ° with respect to the optical axis AX and the magnification of the relay lens 2 is 0.25, the inclination angle of the surface F4 ′ conjugate to the surface F3 ′ with respect to the optical axis AX is obtained from the equation (2). The calculated value is 14.0 °. Further, the size of the two-dimensional light wave image S4 on the F4 ′ plane is 2.58 × 2.5 mm from the equation (3).

The multiplexing unit 3 includes a beam expander 33, a lens 32 having a focal length of 100 mm, and a beam splitter 31.
When the two-dimensional light wave image S4 is distributed on the time-resolved spectroscopic imaging device 4 and the reference light is irradiated through the multiplexing means 3, the two-dimensional light wave image S4 can be recorded as an interference fringe pattern.
A solid-state imaging device (CCD) 4 is used as the imaging device 4 for time-resolved spectroscopy. With this imaging element 4, an interference pattern of the two-dimensional light wave image S4 is obtained.

As described above, in this embodiment, an optical system having a reduction magnification of 0.25 is used as the relay lens 2. Therefore, the inclination angle of the surface F4 ′ conjugate with the surface F3 ′ inclined by 45 ° with respect to the optical axis AX can be reduced to 14.0 °. Therefore, the influence of shading in the image sensor 4 can be greatly reduced.
Furthermore, a microscope that enables observation and time-resolved spectroscopy at the same time for a very small region can be obtained.

According to the microscope apparatus of the first embodiment, it is possible to perform time-resolved spectroscopic measurement of probe light subjected to modulation in a femto to picosecond region simultaneously with observation of a very small region.
In this embodiment, beam splitters 62 each having half mirror surfaces 62b and 62c are provided on both surfaces of a transparent parallel flat plate 62a having a predetermined thickness. Since the pump light and the probe light are spatially separated via the beam splitter 62 and the mirrors 63 and 64, only the necessary probe light and reference light can be extracted. As a result, observation of a very small area and time-resolved spectroscopic measurement can be performed simultaneously and with high accuracy. Furthermore, by disposing the light blocking member at a predetermined position in the optical path, it becomes possible to more reliably block unnecessary pump light at the detection position.

  In the configuration example shown in FIG. 29, the configuration of the first embodiment shown in FIG. 3 is adopted for the branching means 6, but the configuration of the modification shown in FIGS. 4 to 10 may be used. .

FIG. 30 is an explanatory diagram illustrating the overall configuration of the microscope apparatus according to the second embodiment of the present invention. The basic configuration is the same as the configuration in FIGS. 1 and 2, and the sample is observed through the microscope 70 and the time-resolved spectroscopic unit 80 performs time-resolved spectroscopy on a specific region in the sample. Has been.
And in the microscope apparatus of Example 2, the structure of 2nd Embodiment shown in FIG. The two-dimensional light wave converting means 1 employs the configuration shown in FIGS.

The ultrashort light pulse light source 5 is a femtosecond laser that oscillates ultrashort light having a center wavelength of 800 nm, a wavelength width of ± 5 nm, and a pulse width of 100 femtoseconds.
The branching unit 6 includes a beam splitter 61, a beam splitter 62 ', a mirror 63, a light attenuating element 630, mirrors 641 and 642, a fine movement stage 643, a fine movement stage 65, and a spatial filter 66. It is configured.
The light emitted from the ultrashort optical pulse light source 5 is divided into two through the beam splitter 61. The light reflected by the beam splitter 61 is used as reference light. The light that has passed through the beam splitter 61 is divided into two through the beam splitter 62 ′ into transmitted light and reflected light. The bisected light is reflected by the mirrors 63, 641, and 642, and is incident on the beam splitter 62 ′ again. The light reflected by the mirror 63 and transmitted through the beam splitter 62 ′ and the light reflected by the mirrors 641 and 642 and reflected by the beam splitter 62 ′ enter the beam splitter 723 via the light guide unit 91. The light reflected by the beam splitter 723 irradiates the sample S. At that time, one of the lights (probe light) divided into two through the beam splitter 62 ′ is given a time delay with respect to the other (pump light) through the fine movement stage 65. For this reason, the probe light is modulated by the sample S stimulated by the pump light.

In this embodiment, the pump light is light that has been transmitted through the beam splitter 62 ′, reflected by the mirrors 641 and 642, and further reflected by the beam splitter 62 ′. On the other hand, the probe light is light that is reflected by the beam splitter 62 ', then reflected by the mirror 63, and further transmitted through the beam splitter 62'. In the optical path for generating the pump light, the light reflected by the mirror 642 follows an optical path shifted parallel to the optical path of the incident light, and enters the beam splitter 62 ′. Therefore, the probe light (the light reflected by the mirror 63 and then transmitted through the beam splitter 62 ′) and the pump light (the light reflected by the mirrors 641 and 642 and then reflected by the beam splitter 62 ′) are: Spatially separated in parallel.
Note that the amount of spatial separation between the pump light and the probe light is adjusted by adjusting the amount of movement of the mirror 641 and the mirror 642 by the fine movement stage 643.
The spatial filter 66 passes only the probe light out of the light separated in parallel after passing through the sample S, and blocks the pump light. As a result, only the probe light is incident on the two-dimensional light wave conversion means 1.

  The two-dimensional light wave converting means 1 includes a lens 111, a lens 112, a cylindrical beam expander 11 ', a Bragg diffraction grating 12, a first lens 133, a first diffraction grating array 142, a second lens 134, and a second diffraction grating array 15. It consists of and. Here, the focal length of the lens 111 is 100 mm, and the focal length of the lens 112 is 50 mm. The cylindrical beam expander 11 ′ is composed of cylindrical lenses 113 and 114 with a magnification of 10. The first lens 133 and the second lens 134 are lenses having a positive refractive power with a focal length f = 40 mm.

  As shown in FIG. 24, the Bragg diffraction grating 12 is disposed on a surface F1 ′ obtained by rotating the front focal plane F1 of the first lens 133 by approximately 45 °, and is formed between the normal line and the optical axis AX. The angle is approximately 45 °. A surface conjugate with the surface F1 ′ with respect to the lenses 133 and 134 is a surface F3 ′ when the rear focal plane F3 of the second lens 134 is rotated by 45 °. For this reason, the conjugate plane F3 ′ is also at an angle of approximately 45 ° with respect to the optical axis AX.

  Further, in the two-dimensional light wave converting means 1 according to the second embodiment, the probe light is magnified approximately 10 times via the cylindrical beam expander 11 '. The Bragg diffraction grating 12 has a size of about 14.14 mm × 1 mm. Probe light is obliquely incident on such a Bragg diffraction grating 12. At this time, the modulated probe light has a wavelength width of 800 ± 5 nm. Therefore, on the rear focal plane F2 of the first lens 133, as shown in FIG. 25, each wavelength component included in the probe light is distributed along the x-axis direction. At this time, for example, wavelengths of 805 nm, 800 nm, and 795 nm are distributed at positions of +0.5 mm, 0 mm, and −0.5 mm, respectively, with respect to the optical axis AX.

  The first diffraction grating array 142 functions as a filter. As shown in FIG. 26, the first diffraction grating array 142 includes diffraction gratings 142a to 142k having different lattice constants in the x-axis direction, and diffracts each wavelength component of the probe light at different angles. table. 2 shows the specifications of the first diffraction grating array 142. table. 2, the sign of the lattice constant is expressed as positive when diffracting in the + direction with respect to the y-axis on the surface F2, and negative when diffracting in the-direction. The y coordinate represents the height in the y-axis direction of each wavelength after exiting the second lens 134.

table. 2 Specifications of the first diffraction grating array 142

When the light of each wavelength diffracted by the first diffraction grating array 142 is emitted from the second lens 134, it enters the surface F3 ′ in parallel with the xz plane. At this time, the wavelength component of the probe light is distributed in the y-axis direction on the plane F3 ′. When the probe light is sequentially irradiated onto the Bragg type first diffraction grating 12 on the surface F1 ′, the wavelength distribution moves in the x-axis direction on the surface F3 ′ conjugate with the F1 ′ surface. Therefore, similarly to FIG. 21, a two-dimensional light wave S3 is generated in which time changes in the x-axis direction and the wavelength is developed on the vertical axis. The size of the two-dimensional light wave S3 on the surface F3 ′ is 14.14 × 10 mm.

  As shown in FIG. 22, the second diffraction grating array 15 includes diffraction gratings 15 </ b> A to 15 </ b> K having different periods in the y-axis direction. The diffraction grating array 15 corrects the angular distribution of each wavelength for the two-dimensional light wave S3 incident on the surface F3 ′. The specifications of the second diffraction grating array 15 are shown in the following table. 3 shows. table. The incident angle 3 is an angle with respect to the normal line of the second diffraction grating array 15 when light of each wavelength is incident on the surface F3 ′. The diffraction gratings 15A to 15K constituting the second diffraction grating array 15 are shown in FIG. If it has a periodic structure like 3, the diffraction angle in the light of each wavelength will be 45 degrees. As a result, the angular distribution of light of each wavelength after exiting from the surface F3 ′ is almost zero.

table. 3 Specifications of the second diffraction grating array 15

  The relay lens 2 includes a lens 21 having a focal length f21 = 100 mm and a lens 22 having a focal length f22 = 25 mm. Also in this embodiment, a solid-state imaging device (CCD) 4 is used as an imaging device for time-resolved spectroscopy. Therefore, the two-dimensional light wave image S4 is distributed on the image pickup surface by matching the image pickup surface of the image pickup element 4 with the surface F4 'conjugate with the surface F3'. Since the surface F3 ′ is inclined by 45 ° with respect to the optical axis AX and the magnification of the relay lens 2 is 0.25, the inclination angle of the surface F4 ′ conjugate to the surface F3 ′ with respect to the optical axis AX is obtained from the equation (2). The calculated value is 14.0 °. Further, the size of the two-dimensional light wave image S4 on the F4 ′ plane is 2.58 × 2.5 mm from the equation (3).

The multiplexing means 3 includes a beam expander 33, a lens 32 having a focal length of 100 mm, and a beam splitter 31.
When the two-dimensional light wave image S4 is distributed on the image sensor 4 and the reference light is irradiated through the multiplexing unit 3, the two-dimensional light wave image S4 can be recorded as an interference fringe pattern.
As described above, the image pickup device 4 uses the solid-state image pickup device (CCD) 4, and an interference pattern of the two-dimensional light wave image S4 is obtained.

As described above, in this embodiment, the relay lens 2 is an optical system having a reduction magnification of 0.25. Therefore, the inclination angle of the surface F4 ′ conjugate with the surface F3 ′ inclined by 45 ° with respect to the optical axis AX can be reduced to 14.0 °. As a result, the influence of shading in the image sensor 4 can be greatly reduced.
Furthermore, a microscope that enables observation and time-resolved spectroscopy at the same time for a very small region can be obtained.

According to the microscope apparatus of the second embodiment, time-resolved spectroscopic measurement of probe light that has been modulated in the femto to picosecond region can be performed simultaneously with observation of a very small region.
In the present embodiment, since the pump light and the probe light are spatially separated via the beam splitter 62 ′, the mirror 63, and the mirrors 641 and 642, only the necessary probe light can be extracted. As a result, observation of a very small area and time-resolved spectroscopic measurement can be performed simultaneously and with high accuracy. Furthermore, by disposing the light blocking member at a predetermined position in the optical path, it becomes possible to more reliably block unnecessary pump light at the detection position.

Instead of the mirrors 641 and 642 of the branching means 6, a right-angle prism provided with reflection surfaces on two surfaces that form a right angle may be used.
Further, in the configuration example shown in FIG. 30, the configuration of the second embodiment shown in FIG. 11 is adopted for the branching means 6, but the configuration of the modification shown in FIGS. 12 and 13 may be used. . Moreover, you may use the structure of 3rd Embodiment shown in FIGS. 14-16, and the structure of the modification.

  Furthermore, the microscope apparatus of the present invention can constitute a microscope apparatus by an observation method other than the general pump-probe method. Below, the example of a structure is demonstrated.

FIG. 31 is an explanatory diagram illustrating the configuration of the branching unit 6 in the microscope apparatus according to the third embodiment of the present invention. The configuration other than the branching unit 6 can be the same as the configurations shown in the first to fourth embodiments and the first and second embodiments. The description of that part is omitted.
The microscope apparatus of Example 3 is a microscope apparatus that observes coherent Raman scattering light by applying the configuration of the branching unit 6 in the microscope apparatus of the present invention. In this microscope apparatus, light having two different frequencies is regarded as pump light, generated coherent Raman scattered light is regarded as probe light, and the time difference adjusting means is used so as to eliminate the time difference between the two lights. Device configuration is applicable.

The microscope apparatus in FIG. 31 is an example in which measurement is performed by irradiating a sample with two lights having different frequencies. When the sample is irradiated with two lights having different frequencies, the molecules in the sample are excited. Then, coherent anti-Stokes Raman scattering light (CARS light) is generated by the excited molecular vibration. The microscope apparatus of FIG. 31 measures the coherent anti-Stokes Raman scattering light. For the sake of consistency with the expression used in Example 2 or earlier, for convenience, two light beams having different frequencies will be referred to as pump light, and CARS light will be referred to as probe light. (Pump light used in coherent anti-Stokes Raman scattering is here. This is called light of frequency ω ₁ ). Here, light of two frequencies is ω ₁ , ω ₂ (ω ₁ > ω ₂ ), and wave number vectors are k ₁ and k ₂ . Then, the frequency and vector of the generated probe light (CARS light) satisfy 2ω ₁ −ω ₂ and 2k ₁ −k ₂ .

In the microscope apparatus of the third embodiment, the branching unit 6 includes the beam splitter 62 ″, the mirrors 631 ′, 632 ′, 633 ′, 634 ′, 635 ′, the fine movement stage 636 ′, the mirror 64 ″, and the fine movement stage 65. ", A beam splitter 62"', a beam splitter 723, a spatial filter 66, and a spatial filter 67.
The beam splitter 62 ″ is configured to transmit light having a predetermined wavelength used as the first light and reflect light having the predetermined wavelength used as the second light out of the light from the ultrashort optical pulse light source 5. Yes.
The mirror 64 ″ is arranged to reflect the first light transmitted through the beam splitter 62 ″ toward the beam splitter 62 ″ ′.
The mirrors 631 ′ to 635 ′ are arranged so as to reflect the second light reflected by the beam splitter 62 ″ toward the beam splitter 62 ″ ′.

The beam splitter 62 ″ ′ is configured to transmit the first light and reflect the second light. With this, the two light beams of the light guide unit 91 are separated in a state where they are separated from each other in parallel. The light can enter the mirror 912.
The fine movement stage 65 ″ is configured to be able to move the mirror 64 ″ in the direction along the optical axis of incidence on the mirror 64 ″. The fine movement stage 65 ″ is coupled with the mirror 46 to the second light. A function of adjusting the spatial separation amount of the first light is provided.

  The fine movement stage 636 'is configured to be able to move the mirror 633' and the mirror 634 'in a direction along the incident optical axis of the mirror 632' and the outgoing optical axis of the mirror 635 '. Fine movement stage 636 ', in combination with mirror 633' and mirror 634 ', has a function as time difference adjusting means for adjusting the time difference between the second light and the first light. However, in the microscope apparatus according to the third embodiment, the fine movement stage 636 'allows the first light and the second light to pass through the respective optical paths at the same timing. That is, fine movement stage 636 'is used to eliminate the time difference between the first light and the second light.

The beam splitter 723 functions as a branching unit in this embodiment. The beam splitter 723 reflects part of the two lights that have passed through the first light guide unit 91 and transmits the rest. In this way, each of the two lights is branched into two directions, an optical path toward the sample S in the microscope 70 and an optical path not toward the sample S.
As a result, the light reflected by the beam splitter 723 passes through the optical path toward the sample S of the microscope 70. Both the first light and the second light passing through this optical path are irradiated to the same minute region of the sample S. Then, the molecular vibration in the sample is excited by the difference frequency ω ₁ −ω ₂ between the first light and the second light. Then, CARS light having a center frequency 2ω ₁ −ω ₂ is generated in the direction of the vector k ₁ −k ₂ . The CRAS light is incident on the two-dimensional light wave converting means 1 after passing through the spatial filter 66.
On the other hand, the light transmitted through the beam splitter 723 passes through an optical path not directed to the sample S. Of the first light and the second light passing through this optical path, the first light is used as reference light.
The spatial filter 66 functions as CARS light extraction means in the present invention. That is, only the CARS light out of the incident light separated in parallel is allowed to pass through the opening 66a and the other light is shielded.
The spatial filter 67 functions as reference light extraction means in the present invention. That is, only the reference light (here, the first light) out of the incident light separated in parallel is allowed to pass through the opening 67a, and the other light (here, the second light) is shielded.
In addition to the spatial filters 66 and 67, wavelength selection filters 66 ′ and 67 ′ may be used.

  As described above, the microscope apparatus of the present invention has the following features in addition to the invention described in the claims.

(1) The two-dimensional light wave conversion optical system includes a beam expander, a first diffraction grating, a first lens having a positive refractive power, a filter, a second lens having a positive refractive power, and a second lens. And a relay optical system having a reduction magnification, wherein the first diffraction grating is disposed in the vicinity of the front focal position of the first lens, and the filter includes a rear focal position of the first lens, and Arranged in the vicinity of the front focal position of the second lens, and the second diffraction grating is disposed in the vicinity of the rear focal position of the second lens and in the conjugate plane of the first diffraction grating to constitute incident light. The microscope apparatus according to any one of claims 1 to 24, wherein a diffraction angle generated by the first diffraction grating of each wavelength component is compensated and diffracted in the same direction.

(2) The tilt angle of the imaging surface of the image sensor is set to be the tilt angle of the first diffraction grating by the relay optical system in which the first diffraction grating is tilted with respect to the first lens and has a reduction magnification. The microscope apparatus according to (1), wherein the microscope apparatus is smaller.

(3) The beam expander is configured by a rotationally symmetric lens, and the first lens and the second lens are configured by a cylindrical lens, as described in (1) or (2) above Microscope device.

(4) In the above (1) or (2), the beam expander includes a cylindrical lens, and the first lens and the second lens are configured by rotationally symmetric lenses. The microscope apparatus described.

(5) When the axis parallel to the direction having refractive power in the cylindrical lens is the x axis and the axis parallel to the direction having no refractive power in the cylindrical lens is the y axis, the first diffraction grating and the second diffraction grating The diffraction grating has a grating shape that diffracts incident light only in the x-axis direction, the filter includes a light shielding region and an elongated light transmission region, and the light transmission region has the x-axis and the y-axis. The microscope apparatus according to (3), wherein the microscope apparatus is formed in an inclined direction.

(6) When the axis parallel to the direction having refractive power in the cylindrical lens is the x axis and the axis parallel to the direction having no refractive power in the cylindrical lens is the y axis, the first diffraction grating is the x axis The filter has a grating shape that diffracts incident light only in the axial direction, and the filter includes a plurality of diffraction regions formed in a direction along the x-axis, and each of the plurality of diffraction regions extends along the y-axis. A grating shape that diffracts incident light so that the diffraction angles in directions are different, and a grating shape that diffracts incident light so that the second diffraction grating is parallel to the optical axis of the relay optical system. The microscope apparatus according to (4) above, characterized by comprising:

(7) In any one of the above (1) to (6), a parallel plate is provided on the surface side of the first diffraction grating and the second diffraction grating on which the respective diffraction gratings are formed. The microscope apparatus described.

(8) The microscope apparatus according to (7), wherein a parallel plate is provided on a surface side on which the diffraction grating of the filter is formed.

  The microscope apparatus of the present invention is useful in the fields of medicine, pharmaceutics, and biology, in which it is required to measure physical property changes that occur in a very small time region and in a very short time region.

It is a block diagram which shows the structure common to the microscope apparatus concerning each embodiment of this invention. It is a schematic block diagram which shows the example of 1 structure of the microscope used for the microscope apparatus of FIG. It is explanatory drawing which shows the structure of the branch means in the microscope apparatus concerning 1st Embodiment of this invention. It is explanatory drawing which shows the structure of the modification of the branch means 6 shown in FIG. It is explanatory drawing which shows the structure of the other modification of the branch means 6 shown in FIG. It is principal part explanatory drawing which shows the structure of the further another modification of the branch means 6 shown in FIG. It is an enlarged view of the beam splitter 621 shown in FIG. It is an enlarged view of the mirror 622 shown in FIG. It is principal part explanatory drawing which shows the structure of the further another modification of the branch means 6 shown in FIG. FIG. 10 is an enlarged view of the prism 623 shown in FIG. 9. It is explanatory drawing which shows the structure of the branch means in the microscope apparatus concerning 2nd Embodiment of this invention. FIG. 12 is an explanatory diagram illustrating a configuration of a modification of the branching unit 6 illustrated in FIG. 11. It is explanatory drawing which shows the structure of the other modification of the branch means 6 shown in FIG. It is explanatory drawing which shows the structure of the branch means in the microscope apparatus concerning 3rd Embodiment of this invention. It is explanatory drawing which shows the structure of the modification of the branch means 6 shown in FIG. It is explanatory drawing which shows the structure of the other modification of the branch means 6 shown in FIG. The perspective view which shows one structural example about the combination of the two-dimensional light wave conversion means 1, the relay lens 2, the multiplexing means 3, and the probe light-reference light time difference adjustment means 10 applicable to the branch means 6 of 1st-3rd embodiment. FIG. FIG. 18 is an explanatory diagram showing the distribution of probe light on the filter 141 in the configuration example of FIG. 17. FIG. 18 is an explanatory diagram showing an opening of a filter 141 in the configuration example of FIG. 17. It is explanatory drawing which shows wavelength distribution of the probe light which passes the filter 141 in the structural example of FIG. It is explanatory drawing which shows the two-dimensional light wave distribution produced by the two-dimensional light wave conversion means 1 in the structural example of FIG. It is explanatory drawing which shows the diffraction grating array 15 used for the two-dimensional light wave conversion means 1 in the structural example of FIG. It is explanatory drawing which shows the relationship between inclination-angle (theta) and (phi) with respect to the optical axis of the surface F3 'and surface F4' in the structural example of FIG. Another configuration example of the combination of the two-dimensional light wave conversion unit 1, the relay lens 2, the multiplexing unit 3, and the probe light-reference light time difference adjusting unit 10 applicable to the branching unit 6 of the first to third embodiments is shown. It is a perspective view. FIG. 25 is an explanatory diagram showing a distribution of probe light on the first diffraction grating array 142 in the configuration example of FIG. 24. It is explanatory drawing which shows the 1st diffraction grating array 142 used for the two-dimensional light wave conversion means 1 in the structural example of FIG. FIG. 25 is an explanatory diagram showing probe light diffracted by the first diffraction grating array 142 in the configuration example of FIG. 24. It is explanatory drawing which shows the modification of the two-dimensional light wave conversion means 1 in the structural example of FIG. It is explanatory drawing which shows the whole structure of the microscope apparatus concerning Example 1 of this invention. It is explanatory drawing which shows the whole structure of the microscope apparatus concerning Example 2 of this invention. It is explanatory drawing which shows the structure of the branch means 6 in the microscope apparatus concerning Example 3 of this invention. It is a perspective view which shows schematic structure of the two-dimensional space conversion optical system concerning the waveform measurement technique of an ultrashort light pulse. It is explanatory drawing of the filter 700 used for the two-dimensional space conversion optical system of FIG. It is a graph which shows the wavelength distribution two-dimensionally space-transformed by the two-dimensional space-conversion optical system of FIG.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Two-dimensional light wave conversion means 2 Relay lens 3 Combining means 4 Imaging element, CCD
5 Ultrashort optical pulse light source 6 Branch means 10 Probe light-reference light time difference adjusting means 11, 33, 793 Beam expander 11 ′ Cylindrical beam expander 12 Bragg diffraction grating 15 (second) diffraction grating arrays 21, 22, 32 111, 112, 7931, 7932 Lens 31, 61, 62, 62 ', 62 ", 62"', 621, 735 Beam splitter 34, 115, 630 Light attenuating element 62a, 121, 1421, 151 Parallel flat plate 62b, 62c , 62a ′ Half mirror surface 63, 64, 64 ″, 101, 102, 103, 631 ′, 632 ′, 633 ′, 634 ′, 635 ′, 641, 642, 911, 912, 922 Mirror 65, 65 ″, 104 , 636 ', 643 Fine movement stage 66, 67, 794 Spatial filter 66a, 67a Opening 0 Microscope 71 Transmitted illumination light source 72 Transmitted illumination optical system 73 Observation optical system 74 Observation image sensor 75 Sample stage 76 Fluorescent illumination light source 77 Fluorescent illumination optical system 78 Dichroic mirror 80 Time-resolving spectroscopic unit 91 First light guide means 92 Second light guide means 113, 114 Cylindrical lens 131 First cylindrical lens 132 Second cylindrical lens 133 First lens 134 Second lens 141 Filter 142 First diffraction grating array 621a Substrate 621b, 621c, 622b, 622d Surfaces 622, 623 Prism 622a, 623a Transparent member 622c Reflective surface 623c Slope 721 Collector lens 722 Condenser lens 723 Beam splitter 724 Polarizer 725 First DIC filter 731 Objective lens 732 Second DIC filter 733 Child 734 imaging lens 752 x-y stage 772 filter unit 791 lambda / 4 plate 792 polarizer 7721 excitation filter 7722 dichroic mirror 7723 absorption filter

Claims (24)

An optical microscope, a time-resolving spectroscopic unit, a first light guiding means for guiding light from the time-resolving spectroscopic unit to the inside of the optical microscope, and a second guide for guiding light from the optical microscope to the inside of the time-resolving spectroscopic unit. An optical means, and the time-resolved spectroscopic unit includes an ultrashort light pulse light source that oscillates an ultrashort light pulse, a branching means that branches the ultrashort light pulse into reference light, pump light, and probe light, and A multiplexing unit that combines the probe light guided by the second light guiding unit and the reference light; a probe light-reference light time difference adjusting unit that adjusts a time difference between the probe light reaching the multiplexing position and the reference light; includes an imaging device for imaging the interference fringes formed by the reference beam and the probe light are multiplexed, between said second light guiding means and the imaging device, the light guide by the second light guiding means Optical microscope The light from the wavelength in the coordinate axis direction of the one constituting the two-dimensional coordinates are deployed, the microscope time in the other coordinate axis are arranged two-dimensional light waves conversion optical system for converting a two-dimensional optical wave deployment device Because
Pump light-probe light spatial separation means for spatially separating the pump light and probe light, and the pump light-probe for adjusting the time difference between the pump light and the probe light reaching the sample in the optical microscope. Optical time difference adjusting means, and
A reference light branching means for branching the reference light and other light, and
A microscope apparatus comprising probe light extraction means for extracting only probe light from light separated through the pump light-probe light spatial separation means and having passed through a sample in the optical microscope.   Reference light extraction means for extracting only reference light from the light separated through the pump light-probe light space separation means and branched to the reference light side through the reference light branching means, The microscope apparatus according to claim 1.   The pump light-probe light space separating means includes a transflective element having a transflective surface on both surfaces of a transparent parallel plate having a predetermined thickness, and one transflective reflection of the transflective element. A reflective element having a reflective surface for reflecting light from the surface toward the one semi-transmissive reflective surface, and directing light from the other semi-transmissive reflective surface of the semi-transmissive reflective element to the other semi-transmissive reflective surface The microscope apparatus according to claim 1, further comprising a reflective element having a reflective surface that reflects light.   The pump light-probe light space separating means reflects the semi-transmissive reflective element and one of the light transmitted through the semi-transmissive reflective element and the reflected light to guide the semi-transmissive reflective element to the semi-transmissive reflective element And a reflective element having one reflective surface for reflecting the other light from the transflective element toward the transflective element. The microscope apparatus according to 1 or 2.   The microscope apparatus according to claim 4, wherein the reflection element having the two reflection surfaces is two mirrors.   5. The microscope apparatus according to claim 4, wherein the reflection element having the two reflection surfaces is a right-angle prism having reflection surfaces on two surfaces forming a right angle.   The pump light-probe light space separating means includes a transflective element, two mirrors that reflect one of the light transmitted through the transflective element and the reflected light, and the transflective element. A reflection element having one reflection surface for reflecting the other light of the light toward the transflective element, a reflection surface for reflecting light reflected by the two mirrors, and a reflection element having the one reflection surface So that the optical axis of the light reflected by the reflecting surface of the prism and the optical axis of the light emitted from the transmitting surface of the prism are parallel to each other. The microscope apparatus according to claim 1, wherein the microscope apparatus is configured.   The pump light-probe light space separating means includes a transflective element, a lens that is shifted with respect to the incident optical axis of one of the light transmitted through and reflected by the transflective element, 3. A reflecting element having one reflecting surface for reflecting light from the lens and one reflecting element for reflecting the other light from the transflective element. The microscope apparatus described in 1.   The microscope apparatus according to any one of claims 4 to 8, wherein the transflective element includes one transflective surface.   The microscope apparatus according to any one of claims 4 to 8, wherein the transflective element includes a transflective surface on both surfaces of a transparent parallel plate having a predetermined thickness.   The said pump light-probe light space separation means is equipped with the space separation amount adjustment means which can adjust the spatial separation amount of pump light and probe light, The any one of Claims 4-10 characterized by the above-mentioned. The microscope apparatus described.   The space separation amount adjusting means comprises a moving means capable of moving a reflecting element having the two reflecting surfaces in a direction perpendicular to an incident optical axis of the one light. The microscope apparatus according to claim 11, depending on any one of 4 to 6.   The space separation amount adjusting means includes a moving means capable of moving one of the two mirrors along an incident optical axis of light to the one mirror. 12. The microscope apparatus according to claim 11, which is dependent on item 7.   The subordinate to claim 7, wherein the space separation amount adjusting means comprises moving means capable of moving the prism along an optical axis incident on the prism through the two mirrors. Item 12. The microscope apparatus according to Item 11.   The space separation amount adjusting means is constituted by a moving means capable of moving a lens shifted with respect to the incident optical axis of the one light perpendicularly to the incident optical axis of the one light. The microscope apparatus according to claim 11, which is dependent on claim 8.   The pump light-probe light time difference adjusting means includes a reflective element having a reflective surface that reflects light from one semi-transmissive reflective surface of the semi-transmissive reflective element toward the one semi-transmissive reflective surface, and the semi-transmissive Any one of the reflective elements having a reflective surface that reflects light from the other semi-transmissive reflective surface of the reflective element toward the other semi-transmissive reflective surface is arranged along the incident optical axis of the light from the semi-transmissive reflective surface. The microscope apparatus according to claim 3, wherein the microscope apparatus is configured to be movable.   The pump light-probe light time difference adjusting means reflects the semi-transmissive reflective element having one semi-transmissive reflective surface and one of the light transmitted through the semi-transmissive reflective element and the reflected light to reflect the semi-transmissive reflective element. One of a reflective element having two reflective surfaces for guiding to the transmissive reflective element and a reflective element having one reflective surface that reflects the other light from the semi-transmissive reflective element toward the semi-transmissive reflective element, It is comprised by the moving means which can move along the incident optical axis of the light from a transflective element, The subordinate to any one of Claims 4-6 and Claims 4-6 characterized by the above-mentioned. The microscope apparatus in any one of -12.   The pump light-probe light time difference adjusting means reflects the semi-transmissive reflective element having one semi-transmissive reflective surface and one of the light transmitted through the semi-transmissive reflective element and the reflected light to reflect the semi-transmissive reflective element. Either of the two mirrors for guiding to the transflective element and the reflective element having one reflecting surface for reflecting the other light from the transflective element toward the transflective element, the transflective element 15. It is comprised by the moving means which can move along the incident optical axis of the light from following, Claims 9-11, 13 and 14 which depend on Claim 7 characterized by the above-mentioned. Microscope equipment.   The pump light-probe light time difference adjusting unit reflects the light shifted from the incident optical axis of one of the light transmitted through the transflective element and the reflected light, and the light from the lens. Either a reflecting element having one reflecting surface or a reflecting element having one reflecting surface that reflects the other light from the semi-transmissive reflecting element is arranged along the incident optical axis of light from the semi-transmissive reflecting element. The microscope apparatus according to any one of claims 9 to 11 and 15, which is dependent on claim 8 and claim 8, characterized in that the microscope apparatus is configured to be movable.   The microscope apparatus according to any one of claims 1 to 19, wherein the probe light extraction unit is configured by a light shielding member having an opening through which only the probe light passes.   The probe light extraction unit further includes an optical path between the transflective element and a reflective element provided in an optical path that transmits the transflective element, or an optical path that reflects the transflective element and the transflective element. A λ / 4 plate disposed between the reflection element and a polarizing plate disposed on a light-shielding member having an opening through which only the probe light is transmitted. 21. The microscope apparatus according to claim 20, which is dependent on claim 3 or claim 8.   The probe light extraction unit further includes two reflecting surfaces that reflect one of the transflective element and the light transmitted through the transflective element and the reflected light and guide the light to the transflective element. Dependent on claim 4, comprising a λ / 4 plate between the reflection element and a polarizing plate on the passage side of a light shielding member having an opening through which only the probe light passes. The microscope apparatus according to claim 20.   The probe light extracting means further includes a beam formed by sandwiching a second light-shielding member having an opening for allowing only the probe light to pass through a passage side of the light-shielding member having an opening for allowing only the probe light to pass. The microscope apparatus according to claim 20, comprising an expander.   24. The reference light extraction unit according to claim 2, wherein the reference light extraction unit includes a light shielding member having an opening that allows only the reference light to pass therethrough. Microscope equipment.

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