JP2003195172A - Scanning laser microscope - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a scanning laser microscope with which data with desired resolution, quantitative data and reproducible can be obtained. <P>SOLUTION: In the scanning laser microscope, a sample 5 is irradiated with laser beams from a laser beam source 1 via an objective lens 4 while being scanned and data corresponding to fluorescent light or reflected light from the sample 5 are obtained. A spot diameter of the laser beam on the sample 5 and intensity of the laser beam irradiating the sample 5 can be set in order to settle conditions for obtaining the data. <P>COPYRIGHT: (C)2003,JPO

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a scanning laser microscope for irradiating a sample with laser light while scanning the sample and detecting reflection or fluorescence from the sample.

[0002]

2. Description of the Related Art As is well known, a scanning laser microscope irradiates a sample with a point light source such as a laser beam while scanning the sample in the X- and Y-axis directions through an objective lens to emit fluorescence or reflected light from the sample. The observation light is detected again by the photodetector through the objective lens and the optical system, and two-dimensional information is obtained, and the result is displayed on a monitor screen such as a CRT so that it can be observed as image information. Is.

In this case, usually, as the condition setting for acquiring the image of the sample, the selection of the objective lens, the setting of the display image size (the number of vertical and horizontal pixels displayed on the screen), and the scanning speed at the time of displaying on the monitor are performed. Settings, laser light intensity settings
(Relative value to the intensity of the laser light emitted from the laser light source) and the like are performed. In other words, when acquiring the image of the sample, by setting these conditions, the scanning speed and deflection angle of the galvano scanner that scans the laser beam in the XY axis directions, the fluorescence emitted from the sample, and the sampling that detects the reflected light The frequency and the like are determined, and the resulting image signal is, for example, a CRT.
It is designed to be displayed on a monitor such as.

[0004]

However, in the conventional scanning laser microscope, the fluorescence (or reflection) image of the sample can be observed by the confocal optical system by measuring the observation light emitted from the sample. It is the main purpose, and when detecting the observation light emitted from these samples, the time to irradiate one point on the sample with laser light, the distance between the points to be sampled on the sample, or the sample is given. Various types of data are not acquired by actively setting the magnitude of stimulation (excitation). Therefore, there is a problem that data acquisition with a desired resolution, quantitative data acquisition, and reproducible data acquisition cannot be performed.

The present invention has been made in view of the above circumstances, and provides a scanning laser microscope that positively enables data acquisition at a desired resolution, quantitative data acquisition, and reproducible data acquisition. The purpose is to provide.

[0006]

The invention according to claim 1 is
A plurality of objective lenses having different magnifications, an objective lens switching means for selectively inserting one of the plurality of objective lenses into an observation optical axis, a laser light source, and adjusting the intensity of laser light emitted from the laser light source. Intensity adjusting means, scanning means for two-dimensionally scanning the laser light on the sample, pinholes arranged at a position conjugate with the focal plane of the objective lens inserted in the observation optical axis, and passing through the pinholes A photodetector for photoelectrically converting observation light from the sample, an A / D converter for converting an electric signal output from the photodetector into a digital signal, an operation unit for inputting conditions for acquiring an image of the sample, and Based on the image acquisition conditions input to the operating means, the objective lens switching means, the intensity adjusting means, the scanning means, and the A
A microscope control device for controlling an A / D converter and constructing an observation image of the sample from the digital signal;
In a scanning laser microscope equipped with a monitor configured to display an image constructed by the microscope control device, the image acquisition condition input to the operating unit is an irradiation spot diameter of the laser beam with which the sample is irradiated. And the irradiation intensity.

According to a second aspect of the present invention, in the first aspect of the present invention, the image acquisition conditions input to the operating means further include a scanning range of the laser beam, and the laser beam irradiates one point of the sample. And the sampling interval of the observation light.

The invention according to claim 3 is the invention according to claim 1 or 2.
In the invention described above, further comprising a zoom optical system for adjusting the beam diameter of the laser light incident on the objective lens, the microscope control device, the irradiation spot diameter condition of the laser light input to the operation means The zoom optical system is controlled so as to satisfy the condition.

The invention according to claim 4 is the invention according to claim 1 or 2.
In the invention described above, further comprising a laser power monitor means for detecting the intensity of the laser light, the microscope control device, of the laser light input to the operating means based on the detection result of the laser power monitor means. It is characterized in that the intensity adjusting means is controlled so that the irradiation intensity condition is satisfied.

The invention according to claim 5 is the invention according to claim 1 or 2.
In the invention described above, the microscope control device controls the objective lens switching means so as to satisfy a condition of an irradiation spot diameter of the laser light input to the operation means, and the laser light input to the operation means. The intensity adjusting means is controlled to satisfy the irradiation intensity condition, and the scanning width of the scanning means is controlled to satisfy the scanning range of the irradiation spot of the laser light input to the operating means. The scanning speed of the scanning means is controlled so that the irradiation time of the laser light input to the sample at one point on the operating means is satisfied, and the resolution of the image input to the operating means is satisfied. The sampling interval of the A / D converter is controlled.

As a result, according to the present invention, the spot diameter of the laser beam on the sample, the irradiation intensity of the laser beam, the irradiation time of the laser beam per point, the optical data acquisition pitch, the optical data acquisition range, etc. can be arbitrarily set. By setting to, it is possible to acquire various data of resolution according to these condition settings, so that the stimulation (excitation) applied to the sample can be controlled, and the fluorescence or reflected light emitted from the sample can be detected quantitatively. It is possible to perform quantitative measurement and reproducible measurement.

According to the present invention, the spot diameter of the laser beam on the sample is finely controlled by providing a zoom optical system and controlling the numerical aperture (NA) of the objective lens in a stepwise or stepless manner. You can

Further, according to the present invention, by providing a laser power monitor, the intensity of the laser beam irradiated on the sample can be controlled more accurately, and highly accurate quantitative measurement and reproducible measurement can be performed. It can be carried out.

[0014]

BEST MODE FOR CARRYING OUT THE INVENTION Embodiments of the present invention will be described below with reference to the drawings.

(First Embodiment) FIG. 1 shows a schematic structure of a scanning laser microscope to which the present invention is applied.

Reference numeral 1 is a laser light source, and this laser light source 1
The laser light emitted from the laser beam is irradiated onto the sample 5 in a spot shape through the laser power control means 2 as the intensity adjusting means, the dichroic mirror 3, and the objective lens 4. In this case, the laser power control means 2 adjusts the irradiation intensity of the laser light emitted from the laser light source 1. Specifically, the laser power control means 2 inserts into or removes from an optical path of an ND filter (not shown) or an AOTF (acousto-optic variable wavelength filter). ), The irradiation intensity of the laser light is adjusted.
A plurality of objective lenses 4 having different magnifications are provided in a revolver 4a as an objective lens switching means, and one of the plurality of objective lenses 4 is selectively inserted into the observation optical axis by the revolver 4a. It has become so.

The scanning means 6 scans the sample 5 with the spot-shaped laser light. In this case, the scanning means 6
According to the operation control signal from the microscope control device 7, the laser beam is two-dimensionally scanned on the sample 5 in the XY axis directions. At this time, the laser beam is stopped at a certain point on the sample 5. Alternatively, it can be moved at a desired speed.

Observation light of fluorescence or reflected light emitted from the sample 5 irradiated with the laser light is arranged at a position conjugate with the objective lens 4, the scanning means 6, the dichroic mirror 3, and the focal plane of the objective lens 4. It is projected on the photodetector 9 through the confocal pinhole 8.

The photodetector 9 photoelectrically converts the observation light from the sample 5 that has passed through the confocal pinhole 8, and the electric signal photoelectrically converted by the photodetector 9 is converted into a sampling pulse emitted from the microscope controller 7. The signals are synchronously converted into digital signals by the A / D converter 10 and given to the microscope control device 7.

The microscope control device 7 constructs an observation image of the sample 5 from the received signal and displays it on the monitor 11, and also generates light amount data to form a graph or a table on the monitor 1.
It is supposed to be displayed in 1.

On the other hand, the operating means 12 is used by a microscope observer to set conditions for acquiring an image (light intensity data). Here, (a) the laser wavelength for irradiating the sample 5, (b) The spot diameter of the laser beam formed on the sample 5, (c) the illuminance intensity of the laser beam irradiating the sample 5 (d) the time for irradiating one point on the sample 5, or the sample 5
The spot moving speed of the laser light scanning above (e) the interval for sampling the signal of the fluorescence or reflected light emitted from the sample 5 (f) the scanning range for acquiring the image (light intensity data) on the sample 5, etc. Conditions can be set.

Next, the processing for such condition setting will be described.

First, when the laser wavelength for irradiating the sample 5 and the spot diameter of the laser light formed on the sample 5 are set, the spot diameter of the laser light on the sample 5 here is
Since it is determined by the numerical aperture (NA) of the objective lens 4 and the laser wavelength to be used, the objective lens 4 to be used is determined from these laser wavelength and spot diameter. That is, by setting the laser wavelength with which the sample 5 is irradiated and the spot diameter of the laser beam formed on the sample 5 in the operating means 12, the microscope control device 7 determines the objective lens 4 suitable for the laser wavelength. In this case, the determined objective lens 4 is automatically inserted on the observation optical axis by the revolver 4a, or the type of the objective lens 4 to be switched to the optical path is displayed on the monitor 11 and displayed to the observer. The user is informed that the revolver 4a is manually inserted into the observation optical axis.

Next, when the irradiation intensity of the laser beam for irradiating the sample 5 is set, based on this setting, the microscope control device 7 causes the laser power control means 2 of the laser power control means 2 not shown in FIG.
The irradiation intensity of the laser light can be adjusted by inserting / removing the D filter into / from the optical path or by AOTF (acousto-optic variable wavelength filter).

Up to this point, the spot diameter of the laser light formed on the sample 5 and the irradiation intensity of the laser light are determined,
The irradiation intensity of the laser light with which one point on the sample 5 is irradiated can be set.

Next, by setting the time of the laser beam applied to one point on the sample, the moving speed of the spot of the laser beam on the sample 5 is determined. In this case, the time for which the laser light is irradiated to one point on the sample 5 corresponds to the time for which the spot of the laser light moves by the diameter of the spot,
It can be obtained from the value of the spot diameter and the time of irradiation with laser light. Instead of setting the time for irradiating one point on the sample 5, the moving speed of the spot of the laser beam may be set directly to obtain the time for irradiating one point on the sample 5. Then, the spot of the laser beam is the sample 5
When the moving speed for moving above is determined, a control signal is output from the microscope control device 7 to the scanning means 6 such as a galvanometer, and the spot of the laser light is changed to the sample 5.
It is possible to control to stop at a certain point above or move at a certain speed.

Next, when an interval for sampling the fluorescence or reflected light emitted from the sample 5 is set, at what density (resolution) the fluorescence or reflected light emitted from the sample 5 is detected by this sampling interval. Is determined. For example, when the distance between the points to be sampled on the sample 5 is set, the time interval for sampling is determined based on the set distance and the moving speed at which the spot of the laser light moves. Therefore, the microscope is based on this time interval. The resolution of the acquired image is determined by controlling the frequency of the sampling pulse generated from the control device 7 to the A / D converter 10. In this case, the frequency of the sampling pulse generated by the microscope controller 7 to the A / D converter 10 may be controlled by directly setting the time interval for detecting the fluorescence or the reflected light.

When the range for acquiring the image (light intensity data) on the sample 5 is set, the phase relationship with the scanning control signal issued to the scanning means 6 by the microscope control device 7 is controlled according to this setting. Sampling pulses are output to the A / D converter 10 by the set number of samplings, and only light intensity data in the set scanning range is acquired.

Thereafter, under the image acquisition conditions set in this way, the laser light from the laser light source 1 is passed through the objective lens 4 to the sample 5 by the scanning means 6 in the X and Y axis directions as described above. Irradiation is performed while scanning, and the observation light from the sample 5 is detected by the photodetector 9 via the objective lens 4. In this case, the information acquired from the photo detector 9 includes the spot diameter of the laser light on the sample 5, the irradiation intensity of the laser light, the irradiation time of the laser light per point, the optical data acquisition pitch, the optical data acquisition range, and the like. By arbitrarily setting, various data with a resolution according to each condition setting can be acquired. This makes it possible to quantitatively control the stimulation (excitation) applied to the sample 5 and detect the observation light such as fluorescence or reflected light emitted from the sample 5,
It enables quantitative and reproducible measurements.

In the above, the image acquisition conditions such as the spot diameter of the laser light on the sample 5, the irradiation intensity of the laser light, the irradiation time of the laser light per point, the optical data acquisition pitch, and the optical data acquisition range are set. Although an example of setting all is described, at least the spot diameter of the laser beam on the sample 5,
Quantitative and reproducible measurements are possible by setting the irradiation intensity of laser light.

(Second Embodiment) FIG. 2 shows a second embodiment of the present invention.
1 shows the schematic configuration of the embodiment, and the same portions as those in FIG. 1 are denoted by the same reference numerals.

In this case, the zoom optical system 1 is provided in the optical path between the laser power control means 2 and the dichroic mirror 3.
3 is provided. The zoom optical system 13 controls the beam diameter of the laser light incident on the objective lens 4 in a minute step or stepless manner by the zoom operation, and the objective lens 4
Effective Numerical Aperture (NA) of Irradiated Light to Numerical Aperture (NA)
Are trying to control.

With this arrangement, by providing the zoom optical system 13, the numerical aperture (NA) of the objective lens 4 can be narrowed down in minute steps or steplessly.

Usually, the numerical aperture (NA) of the objective lens 4 is
It is determined by the type (including magnification) of the objective lens 4, and for example, even if the spot diameter of the laser beam is set on the sample 5 by the configuration of the first embodiment, the spot diameter of the desired laser beam is not always required. Is not always possible. In view of this, in the second embodiment, the zoom optical system 13 is provided to control the beam diameter of the laser light incident on the objective lens 4 in minute steps or in a stepless manner so that the numerical aperture (NA) of the objective lens 4 is adjusted. Effective numerical aperture of irradiation light (N
By controlling A), the spot diameter of the laser beam on the sample 5 can be finely controlled. As a result, the degree of freedom in setting the irradiation of the laser light on the sample 5 can be increased.

As a modification of the second embodiment,
By providing a diaphragm having a variable opening instead of the zoom optical system 13, it is possible to obtain the same function and effect with similar control.

(Third Embodiment) FIG. 3 shows a third embodiment of the present invention.
1 shows the schematic configuration of the embodiment, and the same portions as those in FIG. 1 are denoted by the same reference numerals.

In this case, the laser power monitor means 14 is provided in the optical path between the scanning means 6 and the objective lens 4. The laser power monitor means 14 is composed of a beam splitter 141, a diaphragm 142, and a photodetector 143. Of these, the beam splitter 14 is included.
1 is inserted in the optical path of the laser light that is incident on the objective lens 4 from the laser light source 1, and the beam splitter 1
A part of the laser beam introduced to the objective lens 4 is returned by 41, and the photodetector 14 passes through the diaphragm 142.
It receives light at 3. Further, the light intensity detected by the photo detector 143 is converted into an electric signal and transmitted to the microscope control device 7. The aperture diameter of the diaphragm 142 can be controlled by the microscope control device 7.
In this case, the aperture diameter of the diaphragm 142 is controlled according to the pupil diameter of the objective lens 4 in order to more accurately monitor the irradiation intensity of the laser light emitted from the objective lens 4. In other words, when the beam diameter of the laser light is larger than the pupil diameter of the objective lens 4, the laser light cuts a part of the light larger than the pupil diameter of the objective lens 4, but it corresponds to the size of the pupil diameter of the objective lens 4. By providing the stop 142, the amount of light irradiated on the sample 5 can be accurately monitored even if the objective lens 4 is switched.

In this way, the laser power monitor means 14 is provided, and the irradiation intensity of the laser light actually emitted from the objective lens 4 can be monitored, and the monitoring result can be fed back. If the laser power control means 2 is controlled according to the comparison result with the value of the intensity of the laser light set by, even if the irradiation intensity of the laser light emitted from the laser light source 1 changes, the sample It is possible to make the intensity of the laser beam applied to the laser beam 5 always match the set value. With this, the irradiation intensity of the laser light with which the sample 5 is irradiated can be controlled more accurately, so that it is possible to perform more accurate quantitative measurement and reproducible measurement.

The laser power monitor means 14
May be constantly inserted in the optical path between the scanning means 6 and the objective lens 4 or may be removable. For example, the irradiation intensity of the laser light emitted from the laser light source 1 is small. In such a case or when the intensity of fluorescence or reflected light emitted from the sample 5 is weak, it may be removed from the optical path except when it is necessary to monitor the intensity of the laser light in order to prevent its attenuation.

It is also possible to add the zoom optical system described in the above-mentioned second embodiment to the structure of the above-mentioned third embodiment. The effect of this form can also be expected.

(Fourth Embodiment) By the way, as a method of using the scanning laser microscope, a method of observing mainly the morphology of the sample as an image is used as in the case of visually observing the fluorescence image of the sample. , There is a method of applying a certain stimulus to a sample and measuring a change caused by the stimulus, for example, a time change of fluorescence intensity.

Therefore, in the fourth embodiment, two setting modes are prepared for the condition setting method in consideration of various usages, and these setting modes can be selectively switched. Here, as the first setting mode,
The image observation mode by the conventional condition setting method and the measurement mode by the condition setting method described in each of the above-described embodiments are used as the second setting mode.

The fourth embodiment employs any one of the configurations of the above-described first to third embodiments. Here, the configuration of the first to third embodiments is adopted. The drawings shall be incorporated.

First, in the case of the image observation mode for observing the form of the sample 5 in the first setting mode as an image, the conventional condition setting, that is, the selection of the objective lens 4 is performed as the condition setting for acquiring the image. Set the display image size (number of pixels), the scanning speed for displaying on the monitor (several steps), the irradiation intensity of laser light (relative value), etc. Then, based on these condition settings, the scanning speed and deflection angle of the galvano scanner that scans the laser light in the XY axis directions, the fluorescence emitted from the sample 5, the sampling frequency that detects the reflected light, etc. are determined. The observation image of the sample 5 obtained as is displayed on the monitor 11.

On the other hand, in the case of the measurement mode in which a stimulus is given to the sample 5 and the change caused by the stimulus is measured, the first to third
1. The condition setting described in the embodiment, that is, the setting of the irradiation intensity of the laser beam applied to the sample 5, the setting of the spot diameter of the laser beam formed on the sample 5, and the setting of the spot of the laser beam scanning the sample 5. The movement continuity of the sample 5, the interval of sampling the signal of the fluorescence or the reflected light emitted from the sample 5, the range of acquiring the image (light intensity data) on the sample 5, and the like are set. Then, various data of resolution based on these condition settings are acquired, and quantitative measurement and reproducible measurement can be performed.

Therefore, in this way, by preparing two setting modes, it is possible to set conditions suitable for various uses, and it is possible to realize desired image observation and light intensity measurement.

In addition, the present invention is not limited to the above-described embodiment, and can be variously modified at the stage of carrying out the invention without changing the gist thereof.

Furthermore, the above-described embodiments include inventions at various stages, and various inventions can be extracted by appropriately combining a plurality of disclosed constituent elements. For example, even if some constituent elements are deleted from all the constituent elements shown in the embodiment, the problem described in the section of the problem to be solved by the invention can be solved, and it is described in the section of the effect of the invention. In the case where the effect described above is obtained, a configuration in which this constituent element is deleted can be extracted as an invention.

The following inventions are also included in the embodiments of the present invention.

(1) A plurality of objective lenses having different magnifications,
Objective lens switching means for selectively inserting one of the plurality of objective lenses into the observation optical axis, laser light source, intensity adjusting means for adjusting the intensity of laser light emitted from the laser light source, and the laser light A scanning means for two-dimensionally scanning the sample, a pinhole arranged at a position conjugate with the focal plane of the objective lens inserted in the observation optical axis, and the observation light from the sample passing through the pinhole is photoelectrically converted. A photodetector for conversion, an A / D converter for converting an electric signal output from the photodetector into a digital signal, an operation unit for inputting conditions for image acquisition of the sample, and the image acquisition input by the operation unit The objective lens switching means, the intensity adjusting means, the scanning means, and the A / D converter are controlled based on the conditions, and the digital signal is controlled. In the scanning laser microscope including a microscope controller that constructs an observation image of the sample, and a monitor that displays an image constructed by the microscope controller, the image acquisition condition input to the operating means is An image observation mode for setting a display image size and a scanning speed at the time of monitor display, an irradiation spot diameter of the laser beam irradiated to the sample, an irradiation intensity and a scanning range, and the laser beam irradiating a point on the sample. The scanning laser microscope is characterized in that it can be set by switching a measurement mode for setting the time to be observed and the sampling interval of the observation light.

In this way, by preparing two setting modes and selectively applying them, it is possible to set conditions suitable for various purposes, and as a result, perform desired image observation and light intensity measurement. be able to.

[0052]

As described above, according to the present invention, there is provided a scanning laser microscope which positively enables data acquisition at a desired resolution, quantitative data acquisition, and reproducible data acquisition. it can.

[Brief description of drawings]

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

FIG. 2 is a diagram showing a schematic configuration of a second embodiment of the present invention.

FIG. 3 is a diagram showing a schematic configuration of a third embodiment of the present invention.

[Explanation of symbols]

1 ... Laser light source 2 ... Laser power control means 3 ... Dichroic mirror 4 ... Objective lens 4a ... Revolver 5 ... Sample 6 ... Scanning means 7 ... Microscope control device 8 ... Confocal pinhole 9 ... Photo detector 10 ... A / D converter 11 ... Monitor 12 ... Operating means 13 ... Zoom optical system 14 ... Laser power monitor means 141 ... Beam splitter 142 ... Aperture 143 ... Photo detector

Continued front page    F term (reference) 2F065 AA51 AA54 BB02 BB05 BB30                       DD03 EE05 FF23 FF41 FF67                       GG04 HH04 JJ01 JJ15 LL06                       LL20 LL25 LL30 LL37 LL57                       LL61 MM16 NN02 NN03 NN17                       PP24 QQ03 QQ31 SS02 SS13                       UU06                 2H045 BA12                 2H052 AA08 AA09 AC15 AC27 AC30                       AC34 AD33 AF00 AF14 AF21                       AF25

Claims (5)

[Claims] 1. A plurality of objective lenses having different magnifications, an objective lens switching means for selectively inserting one of the plurality of objective lenses into an observation optical axis, a laser light source, and a laser beam emitted from the laser light source. Intensity adjusting means for adjusting the intensity of, a scanning means for two-dimensionally scanning the laser light on the sample, a pinhole arranged at a position conjugate with the focal plane of the objective lens inserted in the observation optical axis, A photodetector that photoelectrically converts observation light from the sample that has passed through the pinhole, an A / D converter that converts an electrical signal output from the photodetector into a digital signal, and conditions for acquiring an image of the sample are input. Operating means, the objective lens switching means, the intensity adjusting means, the scanning means, and the operating means based on the image acquisition conditions input to the operating means. A scanning laser microscope including: a microscope controller that controls an A / D converter and constructs an observation image of the sample from the digital signal; and a monitor that displays an image constructed by the microscope controller. The scanning laser microscope, wherein the image acquisition conditions input to the operating means are an irradiation spot diameter and an irradiation intensity of the laser light with which the sample is irradiated. 2. The image acquisition condition input to the operating unit is at least one of a scanning range of the laser light, a time for which the laser light is irradiated to one point of the sample, and a sampling interval of the observation light. The scanning laser microscope according to claim 1, wherein 3. A zoom optical system for adjusting a beam diameter of the laser light incident on the objective lens is further provided, and the microscope control device sets a condition of an irradiation spot diameter of the laser light input to the operation means. The scanning laser microscope according to claim 1, wherein the zoom optical system is controlled so as to satisfy the condition. 4. The laser power monitor means for detecting the intensity of the laser light is further provided, and the microscope control device controls the laser light input into the operating means based on the detection result of the laser power monitor means. 3. The scanning laser microscope according to claim 1, wherein the intensity adjusting means is controlled so that the irradiation intensity condition is satisfied. 5. The microscope control device controls the objective lens switching means so as to satisfy a condition of an irradiation spot diameter of the laser light input to the operation means, and the laser light input to the operation means. Controlling the intensity adjusting means so as to satisfy the irradiation intensity condition, controlling the scanning width of the scanning means so as to satisfy the scanning range of the irradiation spot of the laser beam input to the operating means, The scanning speed of the scanning unit is controlled so that the irradiation time of the laser light input to the sample at one point on the sample is controlled so that the resolution of the image input to the operating unit is satisfied. The scanning laser microscope according to claim 2, wherein a sampling interval of the A / D converter is controlled.