JP2013174848A - Observation device with illuminating light intensity distribution measurement function and illuminating light intensity distribution measurement method for observation device and illuminating light illumination method for observation device - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an observation device with illuminating light intensity distribution measurement function capable of measuring two-dimensional illuminating light intensity distribution on an observation surface in a real time without fluctuating an observation environment during the observation of an observation object such as a sample, and an illuminating light intensity distribution measurement method of an observation device.SOLUTION: The observation device includes: an illuminating light source 10; an array-shaped detector 11 to be arranged on an observation surface 16 only in intensity distribution measurement at a point of time of the initial state of an illuminating light; and an array-shaped detector 12 arranged on an optical path branched from a predetermined position between the light source 10 on an illumination optical path and a lens 15 at the side of the light source 10 arranged at the position nearest to the observation surface 16. The observation device also includes an arithmetic processing unit 13 for calculating illuminating light intensity distribution on the observation surface 16 at an arbitrary point of time by using illuminating light intensity distribution on the observation surface 16 at a point of time of an initial state measured by the detector 11 and the illuminating light intensity distribution at the arrangement position of the detector 12 from the point of time of the initial state to the current point of time measured by the detector 12.

Description

  The present invention relates to an observation device that measures a two-dimensional intensity distribution of illumination light on an observation surface during observation of an observation target such as a specimen, an illumination light intensity distribution measurement method for the observation device, and an illumination light illumination method for the observation device. is there.

  During observation of weak light such as fluorescence emitted from an observation target such as a specimen using an observation apparatus such as a microscope, the intensity of the observed light may change. However, with regard to the causes of changes in the intensity of the weak light emitted from the observation target under observation, conventionally, this is due to a change in the state of the observation target and due to fluctuations in the optical system including the illumination light source that illuminates the observation target. There is a problem that it cannot be distinguished from each other. Regarding the cause of the change in the intensity of the weak light emitted from the observation target under observation, it is possible to identify the weak light observation such as fluorescence observation by identifying the cause caused by fluctuations in the optical system including the illumination light source that illuminates the observation target. This is very important for improving the reliability of analysis, quantification and diagnosis based on data.

  However, for example, in the following Patent Document 1, a detector is arranged in the vicinity of the observation surface to measure a change in excitation light intensity caused by fluctuations in the excitation light source and the optical element arranged on the optical path through which the excitation light passes. A method has been proposed.

FIG. 9 is an explanatory view showing the configuration of the main part of the microscope illumination intensity measuring device described in Patent Document 1. (a) is a diagram showing an example thereof, (b) is a diagram showing another example, and (c) is a further illustration. It is a figure which shows another example.
In the example of FIG. 9A, the stage 51 of the microscope apparatus is provided with the light receiving unit 52 of the microscope illumination intensity measuring apparatus. In the figure, 53 is a living cell, 54 is a container containing the living cell 53, 55 is an irradiation light that irradiates the living cell 53 with excitation light passing through the optical path of an epi-illumination optical system (not shown) and observes the living cell 53. The objective lens 56 is a converter that converts the signal output from the light receiving unit 52 into light intensity information, 57 is a wavelength selection unit that sets the wavelength of the light to be measured, and 58 is the light intensity information converted by the converter. It is a display part to display.
The stage 51 is fixed to a microscope main body (not shown), a movable stage 51b that can move in the B direction in the figure with respect to the fixed stage 51a, and a movable stage 51b that moves in the A direction in the figure. The movable stage 51c is comprised, The container 54 which put the living cell 53 in the upper surface of the movable stage 51c is hold | maintained. The stages 51a to 51c are formed with holes 51a1, 51b1, and 51c1 that allow light to pass through and avoid interference with the objective lens.
The light receiving unit 52 is configured to output an analog signal indicating the intensity of received light, and is fixed at a position where it can be disposed on the observation optical axis O on the upper surface of the movable stage 51c.
In the example of FIG. 9A configured as described above, the light receiving unit 52 is disposed at substantially the same height as the living cell 53 on the observation optical axis O by the movement of the movable stages 51b and 51c as necessary. It is possible to measure the light intensity information of the excitation light having substantially the same length as the optical path length of the excitation light applied to the living cells 53.

In the example of FIG. 9B, a detector 62 that is detachable similarly to the objective lens 55 is provided at the attachment portion of the revolver 61.
The detector 62 includes a light receiving portion 62b at the tip of a hole 62a for passing excitation light passing through the optical path of the epi-illumination optical system.
In the example of FIG. 9B configured as described above, the light receiving unit 62b may be disposed on the observation optical axis O in the vicinity of the living cell 53 by rotating the revolver 61 or the like as necessary. It is possible to measure the light intensity information of the excitation light having a length as close as possible to the optical path length of the excitation light irradiated to the living cells 53.

In the example of FIG. 9C, the light receiving unit 72 is provided in one holding unit of the turret 71b of the condenser unit 71 including the condenser lens 71a that irradiates the living cells 53 with excitation light passing through the optical path of a transmission illumination optical system (not shown). I have.
In the example of FIG. 9C configured as described above, the light receiving unit 72 can be arranged on the observation optical axis O by the rotation of the turret 71b of the condenser unit 71 as necessary. The light intensity information of the excitation light at the arranged position can be measured.

Japanese Patent No. 4689975

However, with the technique described in Patent Document 1, it is not possible to measure in real time the two-dimensional illumination light intensity distribution (illumination unevenness) and its fluctuation on the observation surface while observing the observation target.
That is, when measuring the illumination light intensity while observing the observation target using the technique shown in FIG. 9, the stage 51, the revolver 61, and the turret 71b of the condenser unit 71 are operated each time to observe the observation target (FIG. 9). Then, after the living cells 53), the objective lens 55, and the condenser lens 71a in the container 54 are removed from the observation optical axis O, it is necessary to operate so that the light receiving portions 52, 62b, and 72 are positioned on the observation optical axis O. is there. For this reason, the illumination light at the time when the light receiving units 52, 62b, and 72 receive light is delayed for a predetermined time with respect to the illumination light that is observing the observation target, and the illumination light measured by the light receiving units 52, 62b, and 72 The light intensity is not obtained by measuring the intensity of illumination light during observation of the observation object in real time.

  In addition, when measuring the illumination light intensity while observing the observation target using the technique described in FIG. 9, the observation target is irradiated with the illumination light from the illumination light source while the observation target is being observed. Therefore, when observing the state change of the observation target by irradiating the observation target with illumination light continuously, such as type-lapse observation, the observation target is irradiated during observation. The amount of illumination light cannot be kept constant, and is easily affected by the observation state of the observation target due to vibration during operation of the stage 51, the revolver 61, the turret 71b of the condenser unit 71, and the like.

  In addition, in the example of FIG. 9A, after measuring the light intensity information of the excitation light through the light receiving unit 52 arranged on the observation optical axis O, the stage 51 is operated to enter the sample (in FIG. 9, the container 54). It is difficult to accurately adjust the position of the living cells 53) to their original positions on the two-dimensional coordinates that intersect perpendicularly to the observation optical axis O.

  Further, in the example of FIG. 9B, in order to measure the light intensity information of the excitation light having a length as close as possible to the optical path length of the excitation light irradiated to the living cells 53, the light receiving unit 62b is attached to the container 54 as much as possible. It needs to be close. In addition, since the length of the lens barrel of the objective lens 55 attached to the attachment portion of the revolver 61 varies depending on the magnification, the tip of the lens barrel may collide with the container 54 when the revolver 61 is rotated depending on the magnification. There is. For this reason, in addition to rotating the revolver 61 every time the light intensity information of the excitation light is measured through the light receiving unit 62b arranged on the observation optical axis O, the stage 51 and the revolver on which the container 54 is placed are rotated. It is necessary to perform an operation of changing the relative position with respect to 61 to avoid collision between the container 54 or the stage 51 and the detector 62 or the objective lens 55. However, each time the revolver 61 is operated, the focus of the objective lens 55 on the living cell 53 is shifted. As a result, in the example of FIG. 9B, after measuring the light intensity information of the excitation light through the light receiving unit 61b arranged on the observation optical axis O, the revolver 61 is rotated and the objective lens 55 is moved to the observation light. Even if the original position on the two-dimensional coordinate perpendicular to the axis O is returned, the focusing is likely to be complicated.

  In the example of FIG. 9C, even if the turret 71b is operated to place the light receiving unit 72 on the observation optical axis O, the distance between the light receiving unit 72 and the living cell 53 on the observation optical axis O is the same. The optical path length of the excitation light applied to the living cells 53 is different from the optical path length of the excitation light applied to the light receiving unit 72. For this reason, the light intensity information of the excitation light measured by the light receiving unit 72 is significantly different from the light intensity information at the position where the living cells 53 are arranged.

For this reason, in the technique described in Patent Document 1 as shown in FIG. 9, even if the excitation light intensity is measured at the light receiving unit every predetermined time and the fluorescence intensity is corrected using the temporal change of the excitation light intensity. The time change of the fluorescence intensity emitted from the observation object cannot be measured accurately.
At present, weak light emitted from an observation target such as a specimen is used for diagnosis of the presence or absence of a lesion, a state, and the like. For this reason, if the accuracy of the correction value of the weak light intensity such as fluorescence obtained using the observed illumination light intensity is inferior, it may adversely affect the accurate diagnosis of a lesion or the like.

  In addition, although the light intensity and distribution of the illumination light source vary with time, as described above, the technique described in Patent Document 1 measures the two-dimensional illumination intensity distribution (illumination unevenness) at the same time as observation. Therefore, even if the light intensity distribution adjusting means is used, it is impossible to always illuminate with an arbitrary light intensity distribution in real time.

  The present invention has been made in view of such conventional problems, and the two-dimensional illumination light intensity distribution on the observation surface while observing an observation target such as a specimen, without changing the observation environment, An observation device with an illumination light intensity distribution measurement function that can measure in real time and can always illuminate with an arbitrary light intensity distribution in real time, an illumination light intensity distribution measuring method of the observation device, and illumination light of the observation device It aims to provide a lighting method.

  In order to achieve the above object, an observation apparatus with an illumination light intensity distribution measurement function according to the present invention is disposed on an observation surface only when measuring an illumination light source and an intensity distribution at the time of an initial state of illumination light emitted from the illumination light source. And a predetermined position between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface in the illumination light path of the illumination light source. A second array of detectors arranged on the branched optical path, the observation device having the initial state measured on the observation surface measured by the first array detector Using the illumination light intensity distribution and the illumination light intensity distribution at the placement position of the second array detector from the time of the initial state to the present time measured by the second array detector. At the time of It is characterized by an arithmetic processing unit for calculating an illumination light intensity distribution on the observation surface.

  In the observation apparatus with an illumination light intensity distribution measurement function according to the present invention, the arithmetic processing unit is configured to measure the second array from the time of the initial state to the present time measured by the second array detector. An intensity distribution for detecting a temporal change in the illumination light intensity distribution at the arrangement position of the second array detector from the time of the initial state to the present time using the illumination light intensity distribution at the arrangement position of the shape detector A time change detection step, an illumination light intensity distribution on the observation surface at the time of the initial state measured by the first array detector, and the initial value detected in the intensity distribution time change detection step. The illumination light intensity distribution on the observation surface at an arbitrary time point is calculated using the temporal change of the illumination light intensity distribution at the position of the second array detector from the time point of the state to the present time point. 1's A degree distribution calculating step is preferably performed.

  Moreover, in the observation apparatus with an illumination light intensity distribution measurement function according to the present invention, the arithmetic processing unit is the illumination light on the observation surface at the time of the initial state, measured by the first array detector. Using the intensity distribution and the illumination light intensity distribution at the position of the second array detector at the time of the initial state measured by the second array detector, the time of the initial state An intensity distribution ratio calculating step of calculating a ratio of the illumination light intensity distribution on the observation surface to the illumination light intensity distribution at the position where the second array detector is disposed, and the second array detector The measured illumination light intensity distribution at the current arrangement position of the second array detector and the second array detector at the time of the initial state calculated in the intensity distribution ratio calculation step. At the location Performing a second intensity distribution calculating step of calculating the illumination light intensity distribution on the observation surface at an arbitrary time point using the ratio of the illumination light intensity distribution on the observation surface to the bright light intensity distribution. Is preferred.

  In the observation apparatus with an illumination light intensity distribution measurement function of the present invention, further, from the illumination light source to the lens on the illumination light source side disposed at the position closest to the observation surface in the illumination light path of the illumination light source A field stop at a predetermined position between the illumination light source side and the second array detector is disposed at a position closest to the field stop and the observation plane in the illumination light path of the illumination light source It is preferable that the lens is disposed on an optical path branched from a predetermined position between the lens and the lens.

  In the observation apparatus with an illumination light intensity distribution measurement function of the present invention, it is preferable that the second array detector is disposed at a position conjugate with the observation surface.

  In the observation apparatus with an illumination light intensity distribution measurement function according to the present invention, an optical path in which the first array detector is disposed from the illumination light source and the second array in the illumination light path of the illumination light source are further provided. A spatial light modulator arranged between the optical path where the state detector is arranged and a branch position, and the spatial light modulator is the observation surface at an arbitrary time point calculated by the arithmetic processing unit. It is preferable that the illumination light intensity distribution above can be adjusted so as to be in a desired distribution state.

  Moreover, in the observation apparatus with an illumination light intensity distribution measurement function of the present invention, the illumination light source further disposed at the position closest to the observation surface from the illumination light source in the illumination light path of the imaging element and the illumination light source A dichroic mirror provided at a predetermined position between the lens on the side and guiding the illumination light emitted from the illumination light source to the observation surface, and guiding light of a predetermined wavelength from the observation surface to the imaging device, A predetermined position between the illumination light source side lens disposed at a position closest to the observation surface in the illumination light path of the illumination light source; It is preferable to arrange on the optical path branched from

  In the observation apparatus with an illumination light intensity distribution measurement function according to the present invention, it is preferable that the first and second array detectors are two-dimensional spectrophotometers.

  An illumination light intensity distribution measuring method for an observation apparatus according to the present invention is a method for measuring an illumination light intensity distribution on an observation surface during observation of an observation target in an observation apparatus having an illumination light source, The first array detector is disposed on the observation surface, the illumination light intensity distribution on the observation surface at the initial state is measured by the first array detector, and the first array detector is measured after the measurement. Before the measurement of the illumination light intensity distribution on the observation surface at the time of the initial state by the first array detector. A second array detector is disposed on the optical path branched from a predetermined position between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface in the illumination optical path. And the second array detector A second step of measuring an illumination light intensity distribution at an arrangement position of the second array detector from the time of the initial state to the present time; and a time of the initial state measured in the first step. The illumination light intensity distribution on the observation surface and the time change of the illumination light intensity distribution at the arrangement position of the second array-shaped detector from the time of the initial state to the present time measured in the second step. And a third step of calculating an illumination light intensity distribution on the observation surface at an arbitrary time point.

  Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, in the second step, the second step from the time of the initial state to the present time measured by the second array detector is further performed. The time variation of the illumination light intensity distribution at the position of the second array detector from the initial state to the present time is detected using the illumination light intensity distribution at the position of the array detector. In the third step, the illumination light intensity distribution on the observation surface at the time of the initial state measured in the first step and the time of the initial state detected in the second step It is preferable to calculate the illumination light intensity distribution on the observation surface at an arbitrary time point using the temporal change of the illumination light intensity distribution at the position where the second array detector is disposed up to the present time.

  In the illumination light intensity distribution measuring method of the observation apparatus of the present invention, in the second step, the second array shape at the time of the initial state, which is further measured by the second array detector. Using the illumination light intensity distribution at the detector arrangement position and the illumination light intensity distribution on the observation surface at the initial state measured in the first step, the initial state time The ratio of the illumination light intensity distribution on the observation surface to the illumination light intensity distribution at the arrangement position of the second array detector is calculated, and in the third step, the current time measured in the second step is calculated. Illumination light intensity distribution at the arrangement position of the second array detector, and illumination light at the arrangement position of the second array detector at the time of the initial state calculated in the second step Against intensity distribution Using the ratio of the illumination light intensity distribution on the serial observation plane, it is preferable to calculate the illumination light intensity distribution on the observation surface at any time.

  In the illumination light intensity distribution measuring method of the observation device according to the present invention, the observation device is further arranged in the illumination light path of the illumination light source at a position closest to the observation surface from the illumination light source. A field stop at a predetermined position between the lens on the light source side, and the position of the second array detector in the second step is the field stop and the observation in the illumination optical path of the illumination light source It is preferable that the light path be branched from a predetermined position between the illumination light source side lens disposed at a position closest to the surface.

  Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, it is preferable that the arrangement position of the second array detector in the second step is a position conjugate with the observation surface.

  Further, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, the observation apparatus is further disposed at a position closest to the observation surface from the illumination light source in the illumination light path of the imaging element and the illumination light source. The illumination light source is provided at a predetermined position between the illumination light source side lens and guides illumination light emitted from the illumination light source to the observation surface and guides light of a predetermined wavelength from the observation surface to the imaging element. A dichroic mirror, and an arrangement position of the second array detector in the second step is arranged at a position closest to the dichroic mirror and the observation surface in an illumination optical path of the illumination light source. It is preferable that the optical path is branched from a predetermined position between the illumination light source side lens.

  Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, it is preferable that the first and second array detectors are two-dimensional spectrophotometers.

  Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, it is preferable that the first and second array detectors are shared by one array detector.

  Moreover, the illumination light illumination method of the observation apparatus using the illumination light intensity distribution measuring method of the observation apparatus according to the present invention is such that the first array detector is arranged from the illumination light source in the illumination light path of the illumination light source. A spatial light modulation means is disposed at a predetermined position between the optical path of the second array detector and the optical path where the second array detector is arranged, and the spatial light modulation means is used to The method further includes a step of adjusting the illumination light intensity distribution on the observation surface at an arbitrary time calculated in step 3 so as to be in a desired distribution state.

  In the illumination light illumination method for an observation apparatus according to the present invention, before the measurement of the illumination light intensity distribution on the observation surface at the time of the initial state by the first array detector, the spatial light modulation means It is preferable to set the modulation state so that the modulation state is uniform within the irradiation surface.

  According to the present invention, a two-dimensional illumination light intensity distribution on an observation surface during observation of an observation target such as a specimen is measured in real time without changing the observation environment. An observation apparatus with an illumination light intensity distribution measurement function that enables illumination with a light intensity distribution, an illumination light intensity distribution measurement method for the observation apparatus, and an illumination light illumination method for the observation apparatus are obtained.

It is explanatory drawing which shows schematic structure of the observation apparatus with an illumination light intensity distribution measurement function concerning 1st Embodiment of this invention. It is a flowchart which shows an example of the process sequence of the illumination light intensity distribution measuring method using the observation apparatus in 1st Embodiment. It is a flowchart which shows the other example of the process sequence of the illumination light intensity distribution measuring method using the observation apparatus in 1st Embodiment. It is explanatory drawing which shows schematic structure of the observation apparatus with an illumination light intensity distribution measurement function concerning the modification of FIG. It is explanatory drawing which shows schematic structure of the observation apparatus with an illumination light intensity distribution measurement function concerning 2nd Embodiment of this invention. It is explanatory drawing which shows schematic structure of the observation apparatus with an illumination light intensity distribution measurement function concerning 3rd Embodiment of this invention. It is a flowchart which shows an example of the process sequence of the illumination light illumination method of the observation apparatus in 3rd Embodiment. It is a flowchart which shows an example of the process sequence of the illumination light illumination method of the observation apparatus concerning 4th Embodiment of this invention. It is explanatory drawing which shows the principal part structure of the microscope illumination intensity measuring apparatus of patent document 1, (a) is a figure which shows the example, (b) is a figure which shows another example, (c) is another example. FIG.

Prior to the description of the embodiment, the function and effect of the present invention will be described.
An observation apparatus with an illumination light intensity distribution measurement function according to the present invention is a first array for disposing on an observation surface only when measuring an illumination light source and an intensity distribution at the time of an initial state of illumination light emitted from the illumination light source. And an optical path branched from a predetermined position between the state detector and the illumination light path of the illumination light source between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface. A second array detector, and an illumination light intensity distribution on the observation surface at the time of the initial state measured by the first array detector, Using the illumination light intensity distribution at the arrangement position of the second array-shaped detector from the time of the initial state to the present time, measured by the second array-shaped detector, on the observation surface at any time Illumination light intensity for An arithmetic processing unit for calculating a.

The first array detector is arranged on the observation surface only when measuring the intensity distribution at the time of the initial state of the illumination light emitted from the illumination light source, and the second array detector is illuminated by the illumination light source. An observation target such as a sample is being observed if it is placed on the optical path branched from a predetermined position in the optical path between the illumination light source and the lens on the illumination light source side, which is located closest to the observation surface. The two-dimensional illumination light intensity distribution from the illumination light source can be measured in real time without changing the observation environment.
Further, the illumination light intensity distribution on the observation surface at the time of the initial state measured by the first array detector, and the initial state measured by the second array detector. Provided with an arithmetic processing unit that calculates the illumination light intensity distribution on the observation surface at an arbitrary time point using the illumination light intensity distribution at the arrangement position of the second array detector from the time point to the present time point The real-time two-dimensional illumination light intensity distribution on the observation surface can be measured from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array detector not arranged on the observation surface.
As a result, regarding the cause of the change in the intensity of the weak light emitted from the observation target being observed, it is caused by a change in the state of the observation target, and by the fluctuation of the optical system including the illumination light source that illuminates the observation target. Thus, the fluorescence intensity of the observation target at the observation position can be measured very accurately, and a lesion or the like can be accurately diagnosed.

In this case, the arithmetic processing unit uses the illumination light intensity distribution at the arrangement position of the second array detector from the time of the initial state to the present time measured by the second array detector. An intensity distribution time change detecting step for detecting a time change of the illumination light intensity distribution at the arrangement position of the second array detector from the time of the initial state to the present time; and the first array detector. The intensity distribution of the illumination light on the observation surface at the time of the initial state and the second array detection from the time of the initial state to the present time detected in the intensity distribution time change detection step It is preferable to perform a first intensity distribution calculation step of calculating the illumination light intensity distribution on the observation surface at an arbitrary time point using the temporal change of the illumination light intensity distribution at the placement position of the vessel.
In this way, real-time two-dimensional illumination light intensity distribution on the observation surface can be measured from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array detector that is not arranged on the observation surface. it can.

Alternatively, the arithmetic processing unit measures the illumination light intensity distribution on the observation surface at the time of the initial state measured by the first array detector and the second array detector. Further, using the illumination light intensity distribution at the arrangement position of the second array detector at the time of the initial state, illumination at the arrangement position of the second array detector at the time of the initial state An intensity distribution ratio calculating step for calculating a ratio of the illumination light intensity distribution on the observation surface to the light intensity distribution; and the current second array detector measured by the second array detector. The illumination light intensity distribution at the arrangement position and the illumination light intensity distribution at the arrangement position of the second array detector at the time of the initial state calculated in the intensity distribution ratio calculation step on the observation surface Illumination light intensity distribution By using the ratio, and the second intensity distribution calculating step of calculating an illumination light intensity distribution on the observation surface at any time, preferably performed.
Even in this way, real-time two-dimensional illumination light intensity distribution on the observation surface can be detected from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array detector that is not arranged on the observation surface. it can.

  In addition, the observation object in this application includes the living cell etc. which were cultured in transparent containers, such as a microplate and a petri dish, besides a preparation specimen.

In the observation apparatus with an illumination light intensity distribution measurement function of the present invention, further, from the illumination light source to the lens on the illumination light source side disposed at the position closest to the observation surface in the illumination light path of the illumination light source A field stop at a predetermined position between the illumination light source side and the second array detector is disposed at a position closest to the field stop and the observation plane in the illumination light path of the illumination light source It is preferable that the lens is disposed on an optical path branched from a predetermined position between the lens and the lens.
By arranging the second array-shaped detector in this way, the illumination light traveling toward the observation surface and the illumination light traveling toward the second array-shaped detector do not cause a difference in light quantity due to the field stop. In other words, the real-time two-dimensional illumination light intensity distribution on the observation surface can be measured with higher accuracy.

In the observation apparatus with an illumination light intensity distribution measurement function of the present invention, it is preferable that the second array detector is disposed at a position conjugate with the observation surface.
By arranging the second array detector in this way, the real-time two-dimensional illumination light intensity distribution on the observation surface can be measured with higher accuracy.

In the observation apparatus with an illumination light intensity distribution measurement function according to the present invention, an optical path in which the first array detector is disposed from the illumination light source and the second array in the illumination light path of the illumination light source are further provided. A spatial light modulator arranged between the optical path where the state detector is arranged and a branch position, and the spatial light modulator is the observation surface at an arbitrary time point calculated by the arithmetic processing unit. It is preferable that the illumination light intensity distribution above can be adjusted so as to be in a desired distribution state.
In this way, it is possible to always illuminate with light having a desired light intensity distribution in real time.

Moreover, in the observation apparatus with an illumination light intensity distribution measurement function of the present invention, the illumination light source further disposed at the position closest to the observation surface from the illumination light source in the illumination light path of the imaging element and the illumination light source A dichroic mirror provided at a predetermined position between the lens on the side and guiding the illumination light emitted from the illumination light source to the observation surface, and guiding light of a predetermined wavelength from the observation surface to the imaging device, A predetermined position between the illumination light source side lens disposed at a position closest to the observation surface in the illumination light path of the illumination light source; It is preferable to arrange on the optical path branched from
If the second array detector is arranged in this manner, when the excitation light filter is provided in the dichroic mirror, the wavelengths incident on the first array detector and the second array detector are the same. Therefore, it is not necessary to consider the wavelength characteristics of each detector, and the real-time two-dimensional illumination light intensity distribution on the observation surface can be measured with higher accuracy.

In the observation apparatus with an illumination light intensity distribution measurement function according to the present invention, it is preferable that the first and second array detectors are two-dimensional spectrophotometers.
If the first and second array detectors are configured with a two-dimensional spectrophotometer, it is possible to accurately measure the intensity distribution for each wavelength.

  Further, the illumination light intensity distribution measuring method of the observation device of the present invention is a method for measuring the illumination light intensity distribution on the observation surface during observation of the observation target in the observation device having the illumination light source, The first array detector is disposed on the observation surface, the illumination light intensity distribution on the observation surface at the initial state is measured by the first array detector, and the first array detector is measured after the measurement. Before the measurement of the illumination light intensity distribution on the observation surface at the time of the initial state by the first array detector. A second array detector is disposed on the optical path branched from a predetermined position between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface in the illumination optical path. And the second array detector A second step of measuring an illumination light intensity distribution at an arrangement position of the second array-like detector from the time of the initial state to the present time, and the time of the initial state measured in the first step The illumination light intensity distribution on the observation surface, and the time change of the illumination light intensity distribution at the position where the second array-shaped detector is measured from the time of the initial state to the present time, measured in the second step. And a third step of calculating the illumination light intensity distribution on the observation surface at an arbitrary time.

In the first step, the first array detector is arranged on the observation surface, and the illumination light intensity distribution on the observation surface at the time of the initial state is measured by the first array detector, and the measurement is performed. Later, the first array detector is retracted to a position off the observation surface, and in the second step, the intensity of illumination light on the observation surface at the time of the initial state by the first array detector Before the distribution is measured, the second light beam on the optical path branched from the predetermined position in the illumination light path of the illumination light source from the illumination light source to the lens on the illumination light source side disposed at the position closest to the observation surface. The two array detectors are arranged, and the illumination intensity distribution at the position where the second array detector is disposed from the time of the initial state to the present time is measured by the second array detector. If you are observing the observation object such as a sample The 2-dimensional illumination intensity distribution from the illumination light source, without changing the viewing environment can be measured in real time.
In the third step, the illumination light intensity distribution on the observation surface at the time of the initial state measured in the first step and the time of the initial state measured in the second step If the illumination light intensity distribution on the observation surface at an arbitrary time point is calculated using the temporal change of the illumination light intensity distribution at the position where the second array detector is located up to the present time, The real-time two-dimensional illumination light intensity distribution on the observation surface can be measured from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array-shaped detector that is not arranged on the observation surface.
As a result, regarding the cause of the change in the intensity of the weak light emitted from the observation target being observed, it is caused by a change in the state of the observation target, and by the fluctuation of the optical system including the illumination light source that illuminates the observation target And the fluorescence intensity of the observation target on the observation surface can be measured very accurately, and a lesion or the like can be accurately diagnosed.

In this case, in the second step, the illumination light intensity distribution at the arrangement position of the second array detector from the initial state to the present time, which is measured by the second array detector. Is used to detect a temporal change in the illumination light intensity distribution at the arrangement position of the second array detector from the time of the initial state to the present time. In the third step, in the first step, The measured illumination light intensity distribution on the observation surface at the time of the initial state and the arrangement position of the second array-shaped detector detected from the time of the initial state to the present time detected in the second step. It is preferable to calculate the illumination light intensity distribution on the observation surface at an arbitrary time point using the time variation of the illumination light intensity distribution at.
In this way, real-time two-dimensional illumination light intensity distribution on the observation surface can be measured from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array detector that is not arranged on the observation surface. it can.

Alternatively, in the second step, the illumination light intensity distribution at the arrangement position of the second array detector at the time of the initial state, measured by the second array detector, and the second array detector, Using the illumination light intensity distribution on the observation surface at the time of the initial state, measured in step 1, the illumination light intensity at the arrangement position of the second array detector at the time of the initial state The ratio of the illumination light intensity distribution on the observation surface to the distribution is calculated, and in the third step, the illumination at the current arrangement position of the second array-shaped detector measured in the second step is calculated. The ratio of the light intensity distribution on the observation surface to the light intensity distribution and the illumination light intensity distribution at the position of the second array detector at the time of the initial state calculated in the second step. And use Preferably, for calculating the illumination light intensity distribution on the observation surface at the time of.
Even in this way, real-time two-dimensional illumination light intensity distribution on the observation surface can be measured from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array detector not disposed on the observation surface. it can.

In the illumination light intensity distribution measuring method of the observation device according to the present invention, the observation device is further arranged in the illumination light path of the illumination light source at a position closest to the observation surface from the illumination light source. A field stop at a predetermined position between the lens on the light source side, and the position of the second array detector in the second step is the field stop and the observation in the illumination optical path of the illumination light source It is preferable that the light path be branched from a predetermined position between the illumination light source side lens disposed at a position closest to the surface.
By arranging the second array-shaped detector in this way, the illumination light traveling toward the observation surface and the illumination light traveling toward the second array-shaped detector do not cause a difference in light quantity due to the field stop. In other words, the real-time two-dimensional illumination light intensity distribution on the observation surface can be measured with higher accuracy.

Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, it is preferable that the arrangement position of the second array detector in the second step is a position conjugate with the observation surface.
By arranging the second array detector in this way, the real-time two-dimensional illumination light intensity distribution on the observation surface can be measured with higher accuracy.

Further, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, the observation apparatus is further disposed at a position closest to the observation surface from the illumination light source in the illumination light path of the imaging element and the illumination light source. The illumination light source is provided at a predetermined position between the illumination light source side lens and guides illumination light emitted from the illumination light source to the observation surface and guides light of a predetermined wavelength from the observation surface to the imaging element. A dichroic mirror, and an arrangement position of the second array detector in the second step is arranged at a position closest to the dichroic mirror and the observation surface in an illumination optical path of the illumination light source. It is preferable that the optical path is branched from a predetermined position between the illumination light source side lens.
By arranging the second array detector in this way, when the dichroic mirror is provided with the excitation light filter, the wavelengths incident on the first array detector and the second array detector are the same. Therefore, it is not necessary to consider the wavelength characteristics of each detector, and the real-time two-dimensional illumination light intensity distribution on the observation surface can be measured with higher accuracy.

Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, it is preferable that the first and second array detectors are two-dimensional spectrophotometers.
If a two-dimensional spectrophotometer is used as the first and second array detectors, it can be accurately measured in consideration of the intensity distribution for each wavelength.

Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of the present invention, it is preferable that the first and second array detectors are shared by one array detector.
If the first and second array detectors are shared by one array detector, the cost can be reduced accordingly.
When the first, first, and second array detectors are shared by one array detector, the initial state when the illumination light intensity distribution on the observation surface is measured and the second array detector The time point of the initial state when the illumination light intensity distribution at the position where the detector is arranged does not completely match, and the timing is shifted. However, immediately after measuring the illumination light intensity distribution on the observation surface with the shared array detector, measure the illumination light intensity distribution by placing the shared array detector at the position of the second array detector. As a result, almost the same simultaneity is maintained, so that there is very little possibility that the accuracy of the measurement value will be affected by such a shift in the measurement timing.

Moreover, the illumination light illumination method of the observation apparatus using the illumination light intensity distribution measuring method of the observation apparatus according to the present invention is such that the first array detector is arranged from the illumination light source in the illumination light path of the illumination light source. A spatial light modulation means is disposed at a predetermined position between the optical path of the second array detector and the optical path where the second array detector is arranged, and the spatial light modulation means is used to The method further includes a step of adjusting the illumination light intensity distribution on the observation surface at an arbitrary time calculated in step 3 so as to be in a desired distribution state.
In this way, it is possible to always illuminate with light having a desired light intensity distribution in real time.

In the illumination light illumination method of the observation apparatus using the illumination light intensity distribution measuring method of the observation apparatus of the present invention, the illumination light on the observation surface at the time of the initial state by the first array detector. Prior to the measurement of the intensity distribution, the modulation state of the spatial light modulation means is set to be a uniform modulation state within the irradiation surface.
When the illumination light intensity distribution on the observation surface in the initial state is measured using the first array detector in the configuration including the spatial light modulation unit, for example, the light amount is partially reduced to zero. If the modulation state of the light modulation means is not uniform, the accurate illumination light intensity distribution in the initial state cannot be measured. As a result, even if the spatial light modulation means is modulated into an arbitrary pattern, the observation surface cannot be illuminated with high accuracy with the illumination light intensity distribution based on the desired modulation pattern.
However, as in the illumination light illumination method of the observation apparatus of the present invention, the spatial light modulation means before the measurement of the illumination light intensity distribution on the observation surface at the time of the initial state by the first array detector. If the modulation state is set to be a uniform modulation state within the irradiation surface, the accurate illumination light intensity distribution in the initial state can be measured, and the illumination light intensity distribution based on the desired modulation pattern can be measured on the observation surface. It can be illuminated with high accuracy.

Hereinafter, embodiments of the present invention will be described with reference to the drawings.
First Embodiment FIG. 1 is an explanatory diagram showing a schematic configuration of an observation apparatus with an illumination light intensity distribution measuring function according to a first embodiment of the present invention. FIG. 2 is a flowchart illustrating an example of a processing procedure of an illumination light intensity distribution measuring method using the observation apparatus according to the first embodiment. FIG. 3 is a flowchart showing another example of the processing procedure of the illumination light intensity distribution measuring method using the observation apparatus in the first embodiment. FIG. 4 is an explanatory diagram showing a schematic configuration of an observation apparatus with an illumination light intensity distribution measuring function according to a modification of FIG.

The observation apparatus with an illumination light intensity distribution measuring function of the present embodiment includes an illumination light source 10, a first array detector 11, a second array detector 12, and an arithmetic processing device 13. In FIG. 1, 14 is a field stop, 15 is an objective lens, 16 is a specimen surface, 17 is an image sensor, 18 is a dichroic mirror, 19 is an imaging lens, and 20 is an optical path branching member.
The illumination light source 10 emits illumination light for irradiating the sample surface 16. Here, for convenience of explanation, it is assumed that the illumination light source 10 as a whole emits excitation light.
The first array detector 11 is composed of a CCD, a beam profiler, a PD array or the like, and is arranged on the sample surface 16 only when measuring the intensity distribution at the time of the initial state of the illumination light emitted from the illumination light source 10. Be prepared to be.
The second array detector 12 is a predetermined position in the illumination light path of the illumination light source 10 between the field stop 14 and the lens (objective lens 15) on the illumination light source side disposed at the position closest to the sample surface 16. The optical path branching member 20 disposed on the optical path branches on the optical path branched in a direction different from the specimen surface 16 and is disposed at a position conjugate with the specimen surface 16.
The optical path branching member 20 is composed of a half mirror.
The arithmetic processing unit 13 is configured by using a central processing unit such as a personal computer, for example, and the illumination light on the sample surface 16 at the time of the initial state measured by the first array detector 11. Using the intensity distribution and the illumination light intensity distribution at the arrangement position of the second array detector 12 from the time of the initial state to the present time measured by the second array detector 12 at any time. The illumination light intensity distribution on the sample surface 16 is calculated.

More specifically, the arithmetic unit 13 is configured as in the following first example or second example.
The arithmetic processing device 13 of the first example is configured to perform an intensity distribution time change detection step and a first intensity distribution calculation step.
In the intensity distribution time change detection step, the arithmetic processing unit 13 of the first example arranges the second array detector 12 from the time of the initial state to the present time measured by the second array detector 12. Using the illumination light intensity distribution at the position, a temporal change in the illumination light intensity distribution at the position where the second array-shaped detector 12 is arranged from the time of the initial state to the present time is detected.
In the first intensity distribution calculating step, the arithmetic processing device 13 of the first example measures the illumination light intensity distribution on the sample surface 16 at the time of the initial state, measured by the first array detector 11, Using the time change of the illumination light intensity distribution at the arrangement position of the second array-like detector 12 from the time point of the initial state to the current time point detected in the intensity distribution time change detection step, the sample surface 16 at an arbitrary time point is used. The illumination light intensity distribution above is calculated.

The arithmetic processing device 13 of the second example is configured to perform an intensity distribution ratio calculation step and a second intensity distribution calculation step.
In the intensity distribution ratio calculating step, the arithmetic processing unit 13 of the second example includes the illumination light intensity distribution on the sample surface 16 at the time of the initial state, measured by the first array detector 11, and the second The second array detector at the time of the initial state using the illumination light intensity distribution at the position where the second array detector 12 at the time of the initial state measured by the array detector 12 of the initial state is used. The ratio of the illumination light intensity distribution on the sample surface 16 to the illumination light intensity distribution at the 12 arrangement positions is calculated.
In the second intensity distribution calculating step, the arithmetic processing unit 13 of the second example is the illumination light measured by the second array detector 12 at the current arrangement position of the second array detector 12. The intensity distribution and the ratio of the illumination light intensity distribution on the sample surface 16 to the illumination light intensity distribution at the arrangement position of the second array detector 12 at the time of the initial state calculated in the intensity distribution ratio calculation step. The illumination light intensity distribution on the sample surface 16 at an arbitrary time is calculated.

  The first array-shaped detector 11 is arranged on the sample surface 16 only at the time of measuring the illumination light intensity distribution in the initial state, and is retracted to a position off the sample surface 16 at other times of observation. Is done. The arrangement of the first array detector 11 on the sample surface 16 and the retraction to the position off the sample surface 16 during sample observation at the time of measuring the intensity distribution in the initial state may be performed manually. However, the arrangement of the first array-shaped detector 11 on the sample surface 16 and the position away from the sample surface 16 via a transfer mechanism such as a movable stage and a robot arm and a control means for controlling the transfer mechanism. You may comprise so that evacuation may be performed automatically.

  The dichroic mirror 18 is provided at a predetermined position in the illumination light path of the illumination light source 10 between the field stop 14 and the lens (objective lens 15) on the illumination light source 10 side that is disposed at a position closest to the sample surface 16. The illumination light emitted from the illumination light source 10 is reflected and guided to the sample surface 16, and light having a predetermined wavelength from the sample surface 16 is transmitted and guided to the image sensor 17.

The processing procedure of the illumination light intensity distribution measuring method using the observation apparatus of the first embodiment configured as described above will be described.
First, the first array detector 11 is arranged on the sample surface 16, and the illumination light intensity distribution on the sample surface 16 at the time of the initial state is measured by the first array detector 11, and after the measurement, the first array detector 11 is measured. One array detector 11 is retracted to a position away from the specimen surface 16 (step S1).
In addition, the illumination intensity distribution at the position where the second array detector 12 is arranged from the initial state to the present time is measured by the second array detector 12 (step S2).
Further, the illumination light intensity distribution on the specimen surface 16 at the time of the initial state measured in step S1, and the arrangement position of the second array detector 12 from the time of the initial state to the present time measured in step S2. Is used to calculate the illumination light intensity distribution on the sample surface 16 at an arbitrary time point (step S3).

Here, a more detailed processing procedure of step S2 and step S3 will be described for each case where the arithmetic processing unit 13 is configured in the first example and the second example.
When the arithmetic processing unit 13 is configured in the first example, as shown in FIG. 2, in step S <b> 2, the measurement is further performed from the time point of the initial state measured by the second array detector 12. By using the illumination light intensity distribution at the arrangement position of the second array detector 12 up to the time of the illumination light intensity distribution at the arrangement position of the second array detector 12 from the time of the initial state to the present time. Change is detected as the rate of change. In step S3, the illumination light intensity distribution on the sample surface 16 at the time of the initial state measured in step S1 and the second array detector from the time of the initial state to the present time detected in step S2. The illumination light intensity distribution on the sample surface 16 at an arbitrary time point is calculated by multiplying the change rate of the illumination light intensity distribution at the 12 arrangement positions.

  When the arithmetic processing unit 13 is configured in the second example, as shown in FIG. 3, in step S <b> 2, the second array detector 12 further measures the first state at the time of the initial state. The illumination light intensity distribution at the arrangement position of the two array detectors 12 and the illumination light intensity distribution on the sample surface 16 at the time of the initial state measured in step S1 are used to obtain the first light intensity distribution at the time of the initial state. The ratio of the illumination light intensity distribution on the sample surface 16 to the illumination light intensity distribution at the arrangement position of the two array detectors 12 is calculated. In step S3, the illumination light intensity distribution at the current arrangement position of the second array detector 12 measured in step S2 and the second array detection in the initial state calculated in step S2. The illumination light intensity distribution on the sample surface 16 at an arbitrary time point is calculated using the ratio of the illumination light intensity distribution on the sample surface 16 to the illumination light intensity distribution at the arrangement position of the device 12.

According to the observation apparatus with the illumination light intensity distribution measuring function of the present embodiment, the first array detector 11 is used only when measuring the intensity distribution at the time of the initial state of the illumination light emitted from the illumination light source 10. A lens (objective lens 15) on the side of the illumination light source 10 that is disposed above and the second array detector 12 is disposed in the illumination light path of the illumination light source 10 at a position closest to the sample surface 16 from the illumination light source 10 Therefore, the two-dimensional illumination light intensity distribution from the illumination light source 10 during sample observation can be measured in real time without changing the observation environment.
In addition, the arithmetic processing unit 13 measures the illumination light intensity distribution on the sample surface 16 at the time of the initial state, which is measured by the first array detector 11, and is measured by the second array detector 12. Since the illumination light intensity distribution on the sample surface 16 at an arbitrary time point is calculated using the illumination light intensity distribution at the arrangement position of the second array detector 12 from the time point of the initial state to the present time point, The real-time two-dimensional illumination light intensity distribution on the sample surface 16 can be measured from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array detector 12 that is not arranged on the surface 16.
As a result, regarding the cause of the change in the intensity of the weak light emitted from the observation target such as the sample under observation, this is caused by the change in the state of the sample and the fluctuation of the optical system including the illumination light source that illuminates the observation target. The fluorescence intensity of the observation object at the observation position can be measured with high accuracy by distinguishing the object from the object, and the lesion or the like can be accurately diagnosed.

  In this case, the arithmetic processing unit 13 uses the illumination light intensity distribution at the arrangement position of the second array detector 12 from the time of the initial state to the present time measured by the second array detector 12. An intensity distribution time change detecting step for detecting a time change of the illumination light intensity distribution at the arrangement position of the second array detector 12 from the time of the initial state to the present time, and measurement by the first array detector 11 The illumination light intensity distribution on the sample surface 16 at the time of the initial state and the arrangement position of the second array-like detector 12 from the time of the initial state to the present time detected in the intensity distribution time change detection step. And the first intensity distribution calculating step for calculating the illumination light intensity distribution on the sample surface 16 at an arbitrary time point using the temporal change of the illumination light intensity distribution of Can be realized measurement of the two-dimensional illumination intensity distribution of real-time on the sample surface 16 from the two-dimensional illumination intensity distribution of real-time at the position of the second array detector 12 is not disposed.

  The arithmetic processing unit 13 measures the illumination light intensity distribution on the sample surface 16 at the time of the initial state, measured by the first array detector 11, and measured by the second array detector 12. The illumination light intensity distribution at the arrangement position of the second array-like detector 12 at the time of the initial state is used by using the illumination light intensity distribution at the arrangement position of the second array-like detector 12 at the time of the initial state. The intensity distribution ratio calculating step for calculating the ratio of the illumination light intensity distribution on the sample surface 16 with respect to the sample surface 16, and the current arrangement position of the second array detector 12 measured by the second array detector 12. Of the illumination light intensity distribution on the sample surface 16 with respect to the illumination light intensity distribution at the arrangement position of the second array-like detector 12 at the time of the initial state calculated in the intensity distribution ratio calculation step. Ratio and And a second intensity distribution calculating step for calculating an illumination light intensity distribution on the sample surface 16 at an arbitrary time point, and a second array detector not arranged on the sample surface 16 The real-time two-dimensional illumination light intensity distribution on the sample surface 16 can be measured from the real-time two-dimensional illumination light intensity distribution at the 12 arrangement positions.

  Further, in the observation apparatus with an illumination light intensity distribution measurement function of the present embodiment, the lens on the illumination light source 10 side that is disposed at the position closest to the sample surface 16 from the illumination light source 10 in the illumination light path of the illumination light source 10 is further provided. A field stop 14 is provided at a predetermined position between the objective lens 15 and the second array detector 12 is disposed at a position closest to the field stop 14 and the sample surface 16 in the illumination optical path of the illumination light source 10. Since it is arranged on the optical path branched from the predetermined position between the lens on the illumination light source 10 side (objective lens 15), the illumination light directed to the sample surface 16 and the illumination directed to the second array detector 12 It is not necessary to cause a difference in light quantity due to the field stop 14 in the light, and the real-time two-dimensional illumination light intensity distribution on the sample surface 16 can be measured with higher accuracy.

  Further, according to the observation apparatus with the illumination light intensity distribution measurement function of the present embodiment, since the second array detector 12 is arranged at a position conjugate with the sample surface 16, the real-time 2 on the sample surface 16. Dimensional illumination light intensity distribution can be measured with higher accuracy.

  Further, according to the observation device with the illumination light intensity distribution measurement function of the present embodiment, the imaging device 17 and the illumination light path of the illumination light source 10 are further arranged at a position closest to the sample surface 16 from the field stop 14. Provided at a predetermined position between the illumination light source 10 side lens (objective lens 15), guides the illumination light emitted from the illumination light source 10 to the sample surface 16, and transmits light of a predetermined wavelength from the sample surface 16 to the imaging device. And the second array detector 12 is arranged on the illumination light path of the illumination light source 10 at the position closest to the dichroic mirror 18 and the specimen surface 16. When the dichroic mirror 18 is provided with an excitation light filter, it is arranged on the optical path branched from a predetermined position between the lens (objective lens 15) and Since the wavelengths incident on the first array detector 11 and the second array detector 12 are the same, it is not necessary to consider the wavelength characteristics of the respective detectors. The illumination light intensity distribution can be measured with higher accuracy.

  In the observation apparatus with the illumination light intensity distribution measuring function of the present embodiment, the first and second array detectors 11 and 12 may be configured by a two-dimensional spectrophotometer. If the detectors 11 and 12 are constituted by a two-dimensional spectrophotometer, it is possible to accurately measure in consideration of the intensity distribution for each wavelength.

  Moreover, although the observation apparatus with an illumination light intensity distribution measurement function in FIG. 1 is configured to include an epi-illumination optical system, the observation apparatus with an illumination light intensity distribution measurement function according to the present embodiment includes an epi-illumination optical system. It is not limited to the configuration. For example, as shown in FIG. 4, the structure provided with the transmission illumination optical system may be sufficient. In that case, the lens on the side of the illumination light source 10 arranged at the position closest to the sample surface 16 is not the objective lens 15 but the condenser lens 21. In FIG. 4, 22 is a mirror.

In the observation apparatus with the illumination light intensity distribution measurement function of FIG. 1, the optical path branching member 20 is configured with a half mirror, but may be configured with a mirror provided with a flip-up mechanism instead of the half mirror.
In this way, the sample surface 16 can be irradiated on the specimen surface without reducing the amount of illumination light from the illumination light source during the initial state and during specimen observation by flipping the mirror and retracting it from the optical path. During the specimen observation, the second array detector 12 is used to change the amount of illumination light from the illumination light source while maintaining almost the same time by performing time-division insertion and withdrawal from the optical path during specimen observation. The illumination light intensity distribution at the arrangement position of the second array detector 12 can be measured substantially in real time without being reduced.

Further, according to the illumination light intensity distribution measuring method of the observation apparatus of the present embodiment, in the first step (step S1), the first array-like detector 11 is arranged on the sample surface 16, and the first array. The illumination light intensity distribution on the sample surface 16 at the time of the initial state is measured by the shape detector 11, and after the measurement, the first array detector 11 is retracted to a position off the sample surface 16, and the second step In (step S2), before measuring the illumination light intensity distribution on the sample surface 16 at the time of the initial state by the first array detector 11, the sample surface 16 from the illumination light source 10 in the illumination optical path of the illumination light source 10 is measured. On the optical path branched from a predetermined position between the lens on the side of the illumination light source 10 (the objective lens 15 in the example of FIG. 1 and the condenser lens 21 in the example of FIG. 4) disposed at a position closest to Array detection 12 and the illumination light intensity distribution at the arrangement position of the second array detector 12 from the time of the initial state to the present time is measured by the second array detector 12 so that the specimen is being observed. The two-dimensional illumination light intensity distribution from the illumination light source 10 can be measured in real time without changing the observation environment.
In the third step (step S3), the illumination light intensity distribution on the sample surface 16 at the time of the initial state measured in the first step and the time of the initial state measured in the second step. Since the illumination light intensity distribution on the sample surface 16 at an arbitrary time point is calculated using the time variation of the illumination light intensity distribution at the position where the second array-shaped detector 12 is arranged up to the present time, the sample surface 16 The real-time two-dimensional illumination light intensity distribution on the sample surface 16 can be measured from the real-time two-dimensional illumination light intensity distribution at the arrangement position of the second array-shaped detector 12 not arranged on the top.
As a result, regarding the cause of the change in the intensity of the weak light emitted from the observation target such as the sample under observation, this is caused by the change in the state of the sample and the fluctuation of the optical system including the illumination light source that illuminates the observation target. The fluorescence intensity of the observation object on the observation surface can be measured with high accuracy by distinguishing from the object, and the lesion or the like can be accurately diagnosed.

  In this case, when the arithmetic processing unit 13 is configured in the first example, in the second step, further, the time from the initial state to the present time measured by the second array detector 12 is further measured. Using the illumination light intensity distribution at the arrangement position of the second array-shaped detector 12, the time change of the illumination light intensity distribution at the arrangement position of the second array-shaped detector 12 from the time of the initial state to the present time is calculated. In the third step, the illumination light intensity distribution on the sample surface 16 at the time of the initial state measured in the first step and the time from the time of the initial state detected in the second step to the present time are detected. By calculating the illumination light intensity distribution on the sample surface 16 at an arbitrary time point using the temporal change of the illumination light intensity distribution at the position where the second array detector 12 is disposed, 2nd array not arranged It can be realized measurement of the two-dimensional illumination intensity distribution of real-time on the sample surface 16 from the two-dimensional illumination intensity distribution of real-time at the location of Jo detector 12.

  Further, when the arithmetic processing unit 13 is configured in the second example, in the second step, the second array at the time of the initial state measured by the second array detector 12 is further included. The illumination light intensity distribution at the arrangement position of the shape detector 12 and the illumination light intensity distribution on the sample surface 16 at the time of the initial state measured in the first step are used to obtain the second light at the time of the initial state. The ratio of the illumination light intensity distribution on the sample surface 16 with respect to the illumination light intensity distribution at the arrangement position of the array detector 12 is calculated, and in the third step, the current second time measured in the second step is calculated. The sample surface 16 with respect to the illumination light intensity distribution at the arrangement position of the array-shaped detector 12 and the illumination light intensity distribution at the arrangement position of the second array-shaped detector at the time of the initial state calculated in the second step. Lighting light on By calculating the illumination light intensity distribution on the sample surface 16 at an arbitrary time point using the ratio of the degree distribution, the real time at the arrangement position of the second array detector 12 not arranged on the sample surface 16 is obtained. The real-time measurement of the two-dimensional illumination light intensity distribution on the sample surface 16 can be realized from the two-dimensional illumination light intensity distribution.

  Moreover, according to the illumination light intensity distribution measuring method of the observation device of the present embodiment, the observation device is further arranged in the illumination light path of the illumination light source 10 at the position closest to the sample surface 16 from the illumination light source 10. A field stop 14 is provided at a predetermined position between the lens on the 10th side (the objective lens 15 in the example of FIG. 1 and the condenser lens 21 in the example of FIG. 4), and the second array detector 12 in the second step. Is arranged at the position closest to the field stop 14 and the sample surface 16 in the illumination light path of the illumination light source 10 (the objective lens 15 in the example of FIG. 1 and the lens 15 in the example of FIG. 4). Since the optical path is branched from a predetermined position between the condenser lens 21) and the illumination light traveling toward the sample surface 16 and the illumination light traveling toward the second array-shaped detector 12, the field stop 14 is the original. It finished without causing the quantity difference that a can be measured two-dimensional illumination intensity distribution of real-time on the sample surface 16 with higher accuracy.

  Further, according to the illumination light intensity distribution measuring method of the observation apparatus of the present embodiment, since the arrangement position of the second array detector 12 in the second step is a conjugate position with the sample surface 16, the sample surface 16 The real-time two-dimensional illumination light intensity distribution can be measured with higher accuracy.

  Moreover, according to the illumination light intensity distribution measuring method of the observation apparatus of the present embodiment, the observation apparatus further moves the imaging element 17 and the illumination light path of the illumination light source 10 to the position closest to the sample surface 16 from the illumination light source 10. The illumination light emitted from the illumination light source 10 is guided to the specimen surface 16 and is provided at a predetermined position between the arranged lenses on the illumination light source 10 side (the objective lens 15 in the example of FIG. 1). A dichroic mirror 18 that guides light having a predetermined wavelength from the image sensor 17 to the arrangement position of the second array-like detector 12 in the second step, and the dichroic mirror 18 in the illumination optical path of the illumination light source 10. On the optical path branched from the predetermined position between the lens and the lens on the illumination light source 10 side (objective lens 15 in the example of FIG. 1) arranged closest to the sample surface 16. When the Loic mirror 18 is provided with an excitation light filter, the wavelengths incident on the first array detector 11 and the second array detector 12 are the same, so the wavelength characteristics of each detector are considered. Therefore, the real-time two-dimensional illumination light intensity distribution on the sample surface 16 can be measured with higher accuracy.

Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of this embodiment, it is preferable that the first and second array detectors 11 and 12 are two-dimensional spectrophotometers.
If a two-dimensional spectrophotometer is used as the first and second array detectors 11 and 12, accurate measurement can be performed in consideration of the intensity distribution for each wavelength.

Moreover, in the illumination light intensity distribution measuring method of the observation apparatus of this embodiment, it is preferable that the first and second array detectors 11 and 12 are shared by one array detector.
If the first and second array detectors 11 and 12 are shared by one array detector, the cost can be reduced accordingly.
When the first, first, and second array detectors 11 and 12 are shared by one array detector, the initial time point when the illumination light intensity distribution on the sample surface 16 is measured, The time point of the initial state when the illumination light intensity distribution at the position where the two array detectors 12 are arranged is not completely coincident, and the timing is shifted. However, immediately after measuring the illumination light intensity distribution on the sample surface 16 with the shared array detector, the shared array detector is disposed at the position where the second array detector 12 is disposed, and the illumination light intensity distribution is thus obtained. By measuring this, almost simultaneousness is maintained, and therefore there is very little possibility that the accuracy of the measurement value will be affected by such a deviation in measurement timing.

  In the observation apparatus with the illumination light intensity distribution measurement function of the first embodiment, the second array-shaped detector 12 in the illumination light path of the illumination light source 10 is disposed at a position closest to the sample surface 16 from the field stop 14. The optical path branching member 20 arranged at a predetermined position between the illumination light source side lens (objective lens 15) is arranged on the optical path branched in a direction different from the specimen surface 16, but the specimen from the illumination light source 10 On the optical path branched in a direction different from the sample surface 16 by the optical path branching member 20 disposed at a predetermined position between the illumination light source side lens (objective lens 15) disposed at the position closest to the surface 16 For example, the light is branched in a direction different from the specimen surface 16 by the optical path branching member 20 disposed at a predetermined position between the illumination light source 10 and the field stop 14. Above, it may be a second array detector 12 is arranged. Further, in the observation apparatus with the illumination light intensity distribution measuring function of the example of FIG. 1, the lens (objective lens 15) on the illumination light source 10 side where the dichroic mirror 18 is disposed at the position closest to the sample surface 16 from the field stop 14. However, if it is a predetermined position between the illumination light source 10 and the lens (objective lens 15) on the illumination light source side disposed at the position closest to the sample surface 16, it is arranged anywhere. May be.

Second Embodiment FIG. 5 is an explanatory diagram showing a schematic configuration of an observation apparatus with an illumination light intensity distribution measuring function according to a second embodiment of the present invention.
In the observation apparatus with the illumination light intensity distribution measurement function of the present embodiment, the second array detector 12 is disposed at a position closest to the dichroic mirror 18 and the sample surface 16 in the illumination optical path of the illumination light source 10. It is arranged on an optical path branched in a direction different from the sample surface 16 by an optical path branching member 20 arranged at a predetermined position between the lens (objective lens 15) on the light source 10 side.
Other configurations are substantially the same as those of the observation apparatus with the illumination light intensity distribution measurement function of the first embodiment.

According to the observation device with the illumination light intensity distribution measurement function and the illumination light intensity distribution measurement method of the observation device of the present embodiment, the second array detector 12 is connected to the dichroic mirror 18 and the sample in the illumination light path of the illumination light source 10. Since it is arranged on the optical path branched from a predetermined position between the lens (objective lens 15) on the side of the illumination light source 10 arranged closest to the surface, the dichroic mirror 18 is provided with an excitation light filter. In this case, since the wavelengths incident on the first array detector 11 and the second array detector 12 are the same, it is not necessary to consider the wavelength characteristics of each detector, and the real time on the sample surface 16 is eliminated. The two-dimensional illumination light intensity distribution can be measured with higher accuracy.
Other functions and effects are substantially the same as those of the observation apparatus with the illumination light intensity distribution measurement function and the illumination light intensity distribution measurement method of the observation apparatus according to the first embodiment.

Third Embodiment FIG. 6 is an explanatory diagram showing a schematic configuration of an observation apparatus with an illumination light intensity distribution measuring function according to a third embodiment of the present invention. FIG. 7 is a flowchart illustrating an example of a processing procedure of the illumination light illumination method of the observation apparatus according to the third embodiment.
The observation apparatus with the illumination light intensity distribution measurement function of the present embodiment is an illumination light source in the illumination optical path of the illumination light source 10 in addition to the configuration of the observation apparatus with the illumination light intensity distribution measurement function of the first embodiment shown in FIG. To the branching position between the optical path where the first array detector 11 is arranged and the optical path where the second array detector 12 is arranged (in the example of FIG. 6, the field stop 14 and the optical path branching member 20). Spatial light modulation means 23 disposed between the two.
The spatial light modulator 23 is configured to be adjustable so that the illumination light intensity distribution on the sample surface 16 at an arbitrary time point calculated by the arithmetic processing device 13 is in a desired distribution state.
More specifically, as shown in FIG. 7, the arithmetic processing unit 13 calculates the modulation amount of the spatial light modulator 23 in addition to the intensity distribution time change detection step and the first intensity distribution calculation step shown in FIG. It is configured to perform steps.
In the modulation amount calculation step of the spatial light modulation unit 23, the arithmetic processing unit 13 uses the illumination light intensity distribution on the sample surface 16 at an arbitrary time point calculated in the intensity distribution calculation step, in the spatial light modulation unit 23. A modulation amount necessary for obtaining a desired distribution state is calculated.
The desired distribution state in the present application is the same distribution state as the illumination light intensity distribution on the sample surface 16 in the initial state as a specified value. The observation apparatus according to the present embodiment further includes a modulation mode selection unit 24 that can select a modulation mode. The modulation mode selection unit 24 includes a keyboard, a touch panel, and the like connected to the arithmetic processing unit 13, and as a modulation mode other than the specified value, for example, a specified value on the sample surface 16 with a predetermined light intensity according to the observation application A plurality of types of modulation modes for irradiating illumination light in a desired distribution state other than the above can be selected and input.
Other configurations are substantially the same as those of the observation apparatus with the illumination light intensity distribution measurement function of the first embodiment.

A processing procedure of the illumination light illumination method of the observation apparatus using the illumination light intensity distribution measurement method using the observation apparatus of the third embodiment configured as described above will be described.
The procedure from the first step (step S1) to the third step (step S3) is substantially the same as the procedure shown in FIG.
After step S3, the arithmetic processing unit 13 is necessary to obtain a desired distribution state in the spatial light modulator 23 using the illumination light intensity distribution on the sample surface 16 at an arbitrary time point calculated in step S3. A simple modulation amount is calculated (step S4).
Next, the spatial modulation means 23 modulates the light emitted from the illumination light source 10 based on the calculated modulation amount (step S5).
And while observing a sample, the processing of these steps S2 to S5 is repeated.
According to the observation device and the illumination light illumination method of the observation device of the present embodiment, the illumination light intensity distribution on the sample surface 16 at any time calculated by the arithmetic processing device 13 can be adjusted to a desired distribution state. Since the spatial light modulation means 23 configured as described above is provided, it is possible to always perform illumination with a desired light intensity distribution in real time at an arbitrary time. In addition, since the modulation mode selection means 24 is provided, the specimen surface 16 can be illuminated with light having a desired light intensity distribution in real time at any time according to the observation application.
The spatial light modulation means 23 is an optical path of the illumination light source 10 between the optical path where the first array detector 11 is arranged from the illumination light source and the optical path where the second array detector 12 is arranged. It may be arranged anywhere as long as it is between the branch positions. However, in order to perform the light modulation action by the spatial light modulation means 23 with high density, the area of the irradiation surface incident on the spatial light modulation means 23 such as the vicinity of the field stop 14 is large as shown in FIG. It is preferable to arrange in the vicinity of the pupil position where the light beams intersect.

Fourth Embodiment FIG. 8 is a flowchart showing an example of the processing procedure of the illumination light illumination method of the observation apparatus according to the fourth embodiment of the present invention.
The illumination light illumination method of the observation apparatus of the present embodiment is a measurement of the illumination light intensity distribution on the observation surface 16 at the time of the initial state by the first array detector 11 in addition to the illumination method of the third embodiment. Before, the modulation state of the spatial light modulation means 23 is set to be a uniform modulation state within the irradiation surface (step S0).
The configuration of the observation apparatus, the illumination light intensity distribution measuring method using the observation apparatus, and the illumination light illumination methods of other observation apparatuses are substantially the same as those in the third embodiment.
When the illumination light intensity distribution on the observation surface in the initial state is measured using the first array detector in the configuration including the spatial light modulation means 23, for example, the amount of light is partially reduced to zero. If the modulation state of the light modulator 23 is not uniform, the accurate illumination light intensity distribution in the initial state cannot be measured. As a result, even if the spatial light modulator 23 is modulated into an arbitrary pattern, the sample surface 16 cannot be illuminated with high accuracy with the illumination light intensity distribution based on the desired modulation pattern.
However, like the illumination light illumination method of the observation apparatus of the present embodiment, the spatial light modulation means before the measurement of the illumination light intensity distribution on the sample surface 16 at the time of the initial state by the first array detector 11 is performed. If the 23 modulation states are set to be a uniform modulation state within the irradiation surface, the sample surface 16 can be illuminated with high accuracy with an illumination light intensity distribution based on a desired modulation pattern.
Other functions and effects are substantially the same as those of the illumination light illumination method of the third embodiment.

  In the third and fourth embodiments of FIGS. 6 to 8, the observation device of FIG. 1 includes the spatial light modulator 23. However, the spatial light modulator 23 is added to the observation device of FIGS. 4 and 5. You may comprise. Further, in the illumination light illumination method of the observation device of the third and fourth embodiments, the modulation amount calculation step (step S4) of the spatial light modulation means is added to the illumination light intensity distribution measurement procedure of the observation device shown in FIG. The modulation step by the light modulation means (step S5) and the setting step (step S0) of the modulation state of the spatial light modulation means before the initial state measurement are added. These steps are the same as those of the observation apparatus shown in FIG. You may comprise in addition to the illumination light intensity distribution measurement procedure.

  The observation apparatus with the illumination light intensity distribution measurement function, the illumination light intensity distribution measurement method of the observation apparatus, and the illumination light illumination method of the observation apparatus of the present invention, for example, increase weak light such as fluorescence emitted from an observation target such as a specimen. It is useful in all fields that require accurate observation.

DESCRIPTION OF SYMBOLS 10 Illumination light source 11 1st array detector 12 2nd array detector 13 Arithmetic processor 14 Field stop 15 Objective lens 16 Sample surface (observation surface)
17 Imaging device 18 Dichroic mirror 19 Imaging lens 20 Optical path branching member 21 Condenser lens 22 Mirror 23 Spatial light modulation device 24 Modulation mode selection means 51 Stage 51a Fixed stage 51b, 51c Movable stage 51a1, 51b1, 51c1 Hole 52 Light receiving portion 53 Cell 54 Container 55 Objective lens 56 Converter 57 Wavelength selection unit 58 Display unit 61 Revolver 61b Light receiving unit 71 Condenser unit 71a Condenser lens 71b Turret 72 Light receiving unit

Claims (20)

An illumination light source;
A first array detector for placing on the observation surface only when measuring the intensity distribution at the time of the initial state of the illumination light emitted from the illumination light source;
A second array arranged on an optical path branched from a predetermined position in the illumination light path of the illumination light source from the illumination light source to the lens on the illumination light source side disposed at a position closest to the observation surface. A state detector;
An observation device comprising:
The illumination light intensity distribution on the observation surface at the time of the initial state measured by the first array detector, and the time of the initial state measured by the second array detector. It has an arithmetic processing unit for calculating the illumination light intensity distribution on the observation surface at an arbitrary time point using the illumination light intensity distribution at the arrangement position of the second array detector up to the present time. An observation device with an illumination light intensity distribution measuring function. The arithmetic processing unit includes:
Using the illumination light intensity distribution at the arrangement position of the second array-shaped detector from the time of the initial state to the current time, measured by the second array-shaped detector, from the time of the initial state to the current time An intensity distribution time change detecting step of detecting a time change of the illumination light intensity distribution at the arrangement position of the second array detector up to
The illumination light intensity distribution on the observation surface at the time of the initial state, measured by the first array detector, and the time from the initial state to the current time detected in the intensity distribution time change detection step. The first intensity distribution calculating step of calculating the illumination light intensity distribution on the observation surface at an arbitrary time point using the time variation of the illumination light intensity distribution at the arrangement position of the second array detector. When,
The observation apparatus with illumination light intensity distribution measuring function according to claim 1, wherein: The arithmetic processing unit includes:
The illumination light intensity distribution on the observation surface at the time of the initial state measured by the first array detector, and the time of the initial state measured by the second array detector. The illumination light intensity distribution at the arrangement position of the second array detector is used on the observation surface with respect to the illumination light intensity distribution at the arrangement position of the second array detector at the time of the initial state. An intensity distribution ratio calculating step of calculating a ratio of the illumination light intensity distribution at
Illumination light intensity distribution at the current arrangement position of the second array-shaped detector measured by the second array-shaped detector, and calculated at the intensity distribution ratio calculating step, at the time of the initial state Using the ratio of the illumination light intensity distribution on the observation surface to the illumination light intensity distribution at the arrangement position of the second array detector, the illumination light intensity distribution on the observation surface at an arbitrary time point A second intensity distribution calculating step for calculating;
The observation apparatus with illumination light intensity distribution measuring function according to claim 1, wherein: Furthermore, in the illumination light path of the illumination light source, a field stop is provided at a predetermined position between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface,
The second array detector is branched from a predetermined position between the field stop and the lens on the side of the illumination light source disposed at a position closest to the observation surface in the illumination light path of the illumination light source. The observation device with an illumination light intensity distribution measurement function according to claim 1, wherein the observation device is disposed on an optical path.   5. The observation apparatus with an illumination light intensity distribution measuring function according to claim 4, wherein the second array detector is disposed at a position conjugate with the observation surface. Further, in the illumination light path of the illumination light source, between the illumination light source and a branch position between the light path where the first array detector is disposed and the light path where the second array detector is disposed. Having spatial light modulation means arranged;
The spatial light modulation means is configured to be adjustable so that an illumination light intensity distribution on the observation surface at an arbitrary time point calculated by the arithmetic processing device is in a desired distribution state. The observation apparatus with an illumination light intensity distribution measurement function according to claim 1. Furthermore, an image sensor,
Illumination light emitted from the illumination light source is provided at a predetermined position in the illumination light path of the illumination light source between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface. A dichroic mirror that guides the light of a predetermined wavelength from the observation surface to the imaging device, and guides the light to the observation surface;
Have
The second array detector is branched from a predetermined position between the dichroic mirror and the lens on the side of the illumination light source disposed at a position closest to the observation surface in the illumination light path of the illumination light source. The observation apparatus with an illumination light intensity distribution measurement function according to claim 1, wherein the observation apparatus is disposed on an optical path.   The observation apparatus with an illumination light intensity distribution measuring function according to claim 1, wherein the first and second array detectors are two-dimensional spectrophotometers. A method of measuring an illumination light intensity distribution on an observation surface during observation of an observation target in an observation apparatus having an illumination light source,
A first array detector is arranged on the observation surface, and an illumination light intensity distribution on the observation surface at the time of an initial state is measured by the first array detector, and after the measurement, the first array detector is measured. A first step of retracting the array detector to a position off the viewing surface;
Before the measurement of the illumination light intensity distribution on the observation surface at the time of the initial state by the first array detector, it is arranged at the position closest to the observation surface from the illumination light source in the illumination light path of the illumination light source A second array detector is disposed on an optical path branched from a predetermined position between the illumination light source side lens and the time point of the initial state by the second array detector. A second step of measuring an illumination light intensity distribution at an arrangement position of the second array-like detector from to the present time;
Illumination light intensity distribution on the observation surface at the time of the initial state measured in the first step, and the second array from the time of the initial state to the present time measured in the second step A third step of calculating the illumination light intensity distribution on the observation surface at an arbitrary time point using the illumination light intensity distribution at the arrangement position of the shape detector;
An illumination light intensity distribution measuring method for an observation apparatus characterized by comprising: In the second step, the illumination light intensity distribution at the arrangement position of the second array detector from the time of the initial state to the present time measured by the second array detector is further used. , Detecting a temporal change in the illumination light intensity distribution at the arrangement position of the second array detector from the time of the initial state to the present time,
In the third step, the illumination light intensity distribution on the observation surface at the time of the initial state measured in the first step and the current time from the time of the initial state detected in the second step The illumination light intensity distribution on the observation surface at an arbitrary time point is calculated using the time variation of the illumination light intensity distribution at the arrangement position of the second array detector up to. Item 10. A method for measuring illumination light intensity distribution of an observation device according to Item 9. In the second step, the illumination light intensity distribution at the arrangement position of the second array detector at the time of the initial state, measured by the second array detector, and the first array detector, The illumination light intensity distribution on the observation surface at the time of the initial state measured in the step is used for the illumination light intensity distribution at the arrangement position of the second array detector at the time of the initial state. Calculate the ratio of the illumination light intensity distribution on the observation surface,
In the third step, the illumination light intensity distribution at the current arrangement position of the second array-shaped detector measured in the second step, and the initial state calculated in the second step Using the ratio of the illumination light intensity distribution on the observation surface to the illumination light intensity distribution at the arrangement position of the second array detector at the time point, the illumination light intensity on the observation surface at an arbitrary time point The illumination light intensity distribution measuring method for an observation apparatus according to claim 9, wherein the distribution is calculated. The observation device further includes a field stop at a predetermined position in the illumination light path of the illumination light source between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface,
The arrangement position of the second array detector in the second step is the illumination light source side lens arranged at the position closest to the field stop and the observation surface in the illumination light path of the illumination light source. The illumination light intensity distribution measuring method for an observation apparatus according to claim 9, wherein the illumination light intensity distribution is measured on a light path branched from a predetermined position.   The method for measuring an illumination light intensity distribution of an observation apparatus according to claim 12, wherein an arrangement position of the second array detector in the second step is a position conjugate with the observation surface.   A predetermined position in the illumination light path of the illumination light source between the illumination light source and a branch position between the light path where the first array detector is disposed and the light path where the second array detector is disposed The illumination light intensity distribution on the observation surface at an arbitrary time calculated in the third step becomes a desired distribution state by using a spatial light modulation means. An illumination light illumination method for an observation apparatus using the observation apparatus illumination light intensity distribution measuring method according to any one of claims 9 to 13, further comprising a step of adjusting as described above.   Before the measurement of the illumination light intensity distribution on the observation surface at the time of the initial state by the first array detector, the modulation state of the spatial light modulator is made uniform in the irradiation surface. The illumination light illumination method for an observation apparatus according to claim 14, wherein The observation apparatus further includes:
An image sensor;
Illumination light emitted from the illumination light source is provided at a predetermined position in the illumination light path of the illumination light source between the illumination light source and the lens on the illumination light source side disposed at a position closest to the observation surface. A dichroic mirror that guides the light of a predetermined wavelength from the observation surface to the imaging device, and guides the light to the observation surface;
Have
In the second step, the arrangement position of the second array detector is an illumination light source side lens arranged at a position closest to the dichroic mirror and the observation surface in an illumination optical path of the illumination light source. The illumination light intensity distribution measuring method for an observation apparatus according to claim 9, wherein the illumination light intensity distribution is measured on a light path branched from a predetermined position between the observation device and the light path.   The illumination light intensity distribution measuring method for an observation apparatus according to any one of claims 9 to 13, wherein the first and second array detectors are two-dimensional spectrophotometers. The illumination light intensity distribution measuring method for an observation apparatus according to claim 9, wherein the first and second array detectors are shared by one array detector. .   A predetermined position in the illumination light path of the illumination light source between the illumination light source and a branch position between the light path where the first array detector is disposed and the light path where the second array detector is disposed The illumination light intensity distribution on the observation surface at an arbitrary time calculated in the third step becomes a desired distribution state by using a spatial light modulation means. The illumination light illumination method for an observation device using the illumination light intensity distribution measuring method for the observation device according to any one of claims 16 to 18, further comprising a step of adjusting as described above.   Before the measurement of the illumination light intensity distribution on the observation surface at the time of the initial state by the first array detector, the modulation state of the spatial light modulator is made uniform in the irradiation surface. The illumination light illumination method for an observation apparatus according to claim 19, wherein the illumination light illumination method is used.

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