JP2010266278A - Microscope - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a microscope capable of keeping a spectrum shape of fluorescence to be detected even in the case that a plurality of kinds of fluorescence are detected and capable of easily adjusting the sensitivity of each channel. <P>SOLUTION: The microscope 1 is equipped with an object lens 114 condensing light generated in a specimen 115, a grating 117 for spectrally diffracting the condensed light into spectrum components, a PMT 118 having a large number of channels for detecting the spectrally diffracted spectrum components and a control part 301 for adjusting the sensitivity of each of the channels of the PMT 118. The control part 301 divides the channels into a plurality of groups on the basis of the data related to the wavelength range of the light to be detected and collectively adjusts the sensitivity of each of the channels at each group so that all of the channels in the groups become the same in sensitivity. <P>COPYRIGHT: (C)2011,JPO&INPIT

Description

  The present invention relates to a microscope.

  Conventionally, a microscope is known in which fluorescence emitted from a sample is divided into spectral components by a spectroscopic element, and the divided spectral components are detected by a multi-channel photodetector having a plurality of channels (for example, Patent Document 1). And 2).

JP 2006-098419 A JP 2006-138875 A

  By the way, in the microscopes disclosed in Patent Documents 1 and 2, the sensitivity of all the channels is adjusted at once by adjusting the voltage applied to the multichannel photodetector. Therefore, when observing a plurality of types of fluorescence as in the case of multiple staining observation, if the intensity of any fluorescence is increased, the spectral components may be saturated in other fluorescence. As a result, the spectrum shape of other fluorescence changes, and there is a disadvantage that observation cannot be performed with high accuracy.

  On the other hand, in order to adjust the gain while maintaining the spectrum shape in each fluorescence, the sensitivity of each channel has to be adjusted one by one, which has the disadvantage that the operation becomes very complicated.

  The present invention has been made in view of the above circumstances, and provides a microscope capable of maintaining the spectral shape of the fluorescence to be detected and easily adjusting the sensitivity of each channel when detecting multiple types of fluorescence. The purpose is to do.

In order to achieve the above object, the present invention employs the following means.
The present invention provides an objective lens that collects light generated in a specimen, a spectroscopic element that splits light collected by the objective lens into spectral components, and a plurality of spectral components that detect spectral components dispersed by the spectroscopic elements. And a sensitivity adjustment unit that adjusts the sensitivity of each channel of the photodetector, and the sensitivity adjustment unit assigns the channels to a plurality of channels based on information on the wavelength range of light to be detected. The microscope is divided into groups, and the sensitivity of the channels is collectively adjusted for each group so that all the channels in the group have the same sensitivity.

  According to the present invention, the light generated in the specimen and collected by the objective lens is split into spectral components by the spectroscopic element, and the spectral components that are split are detected by the photodetectors having a plurality of channels, respectively. . In this case, the channels of the photodetector are divided into a plurality of groups based on the information on the wavelength range of the light to be detected by the sensitivity adjustment unit, so that all the channels in the group have the same sensitivity. Sensitivity is collectively adjusted for each group.

  Thereby, the intensity | strength can be adjusted, maintaining a spectrum shape for every grouped channel group, ie, for every wavelength range of the light to detect. For example, even when observing a plurality of types of fluorescence at the same time, the intensity of each fluorescence can be individually adjusted without saturating the spectral components of each fluorescence, and can be observed with high accuracy. Further, it is possible to eliminate the necessity of individually adjusting the sensitivity of each channel, and the adjustment work can be facilitated.

In the above invention, the sensitivity adjustment unit may adjust the voltage applied to the photodetector for each group of the channels.
By adjusting the voltage applied to the photodetector for each group by the sensitivity adjustment unit, the sensitivity of the channel can be adjusted for each group, while maintaining the spectral shape for each wavelength range of light to be detected. , Its strength can be adjusted.

In the above invention, an integration unit that integrates luminance information of light detected by the photodetector may be provided, and the sensitivity adjustment unit may adjust the integration time of the integration unit for each group of the channels.
By adjusting the integration time of the integration unit for each group by the sensitivity adjustment unit, the luminance information detected by each channel can be adjusted for each group, while maintaining the spectrum shape for each wavelength range of the light to be detected , Its strength can be adjusted.

In the above invention, an amplification unit that amplifies luminance information of light detected by the photodetector may be provided, and the sensitivity adjustment unit may adjust the amplification factor of the amplification unit for each group of the channels.
By adjusting the amplification factor of the amplification unit for each group by the sensitivity adjustment unit, it is possible to adjust the luminance information detected by each channel for each group, while maintaining the spectrum shape for each wavelength range of light to be detected , Its strength can be adjusted.

The said invention WHEREIN: It is good also as providing the input part which can set the sensitivity adjustment value of the said sensitivity adjustment part for every group with respect to the group of the said several channel.
In this way, the sensitivity adjustment value of the sensitivity adjustment unit is set for each group for a group of a plurality of channels by an input unit such as GUI (Graphical User Interface), and the sensitivity of each channel is set for each group. Can be adjusted.

  In the above invention, an overall sensitivity adjustment unit that adjusts sensitivity of the entire channel, an integration unit that integrates luminance information of light detected by the photodetector, and luminance information integrated by the integration unit is equal to or greater than a predetermined value. A storage unit that stores the maximum sensitivity of the entire channel such that any one of the spectral components is saturated at the maximum integration time, and the sensitivity adjustment unit stores the sensitivity of the entire channel in the storage unit. When the sensitivity falls below the maximum sensitivity, the integration time of the integration unit may be maximized for each group of the channels.

  By adjusting the integration time of the integration unit for each group by the sensitivity adjustment unit, the luminance information detected by each channel can be adjusted for each group, while maintaining the spectrum shape for each wavelength range of the light to be detected , Its strength can be adjusted. In this case, when the sensitivity of the entire channel falls below the maximum sensitivity at which one of the spectral components is saturated, the integration time of the integrated luminance information is maximized by maximizing the integration time for each channel group. The S / N ratio can be improved.

In the above invention, the sensitivity adjustment unit may adjust the sensitivity of the channel for each group when any of the spectral components detected by the channel exceeds a predetermined threshold.
In this way, when one of the spectral components exceeds a predetermined threshold, for example, when one of the spectral components is saturated, the sensitivity of the channel is adjusted, so that any spectral component is saturated. And the intensity of the detected light can be adjusted.

In the above invention, a storage unit that stores the sensitivity and luminance information of the channel for each group in association with each other may be provided.
By storing the sensitivity of each group of channels and the luminance information in the storage unit in association with each other, the sensitivity of the channels can be reproduced for each wavelength range of light to be detected. Further, from the luminance information for each wavelength range and its sensitivity, it is possible to generate a sample image when all channels have the same sensitivity.

  In the above invention, in the image generation unit that generates an image based on the light detected by the photodetector, the display unit that displays the image generated by the image generation unit, and the screen displayed on the display unit A setting unit configured to set a predetermined range, and the sensitivity adjustment unit may adjust the sensitivity of the channel within a predetermined range set by the setting unit.

  By doing in this way, a region (predetermined range) to be noted by the setting unit is set on the screen displayed on the display unit, and in this region of interest (predetermined range), for each wavelength range of light to be detected, The intensity can be adjusted while maintaining the spectral shape. On the other hand, it is possible to eliminate the necessity of adjusting the intensity of the non-attention area in the screen displayed on the display unit.

  According to the present invention, when detecting a plurality of types of fluorescence, the spectral shape of the fluorescence to be detected is maintained, and the sensitivity of each channel can be easily adjusted.

1 is a schematic configuration diagram of a microscope according to an embodiment of the present invention. It is a functional block diagram of the signal adjustment part of FIG. It is a flowchart which shows the process performed by the microscope of FIG. It is a flowchart which shows the detailed process of step 5 of FIG. It is a figure explaining the state in which the digital signal value less than a setting value is input into CPU of FIG. It is a figure explaining the state in which the digital signal value more than a setting value is input into CPU of FIG. It is a flowchart which shows the process performed with the microscope which concerns on a 1st modification. It is a flowchart which shows the process performed with the microscope which concerns on a 2nd modification. It is a flowchart which shows the process performed with the microscope which concerns on a 3rd modification. It is a flowchart which shows the process performed by the microscope which concerns on a 4th modification. It is a figure explaining operation | movement of the microscope of FIG.

A microscope according to an embodiment of the present invention will be described with reference to the drawings.
FIG. 1 is a schematic configuration diagram of a microscope 1 according to the present embodiment. As shown in FIG. 1, the microscope 1 of this embodiment adjusts the intensity | strength etc. of the signal from the microscope main body 101 which irradiates the laser beam to the specimen 115, and detects the fluorescence which generate | occur | produced in the specimen 115. A signal adjustment unit 201, a control unit (sensitivity adjustment unit, image generation unit) 301 that controls each unit and generates an image based on a signal from the microscope main body 101, and an image of the specimen 115 generated by the control unit 301 A display unit 401 to be displayed and an input unit (input unit, setting unit) 501 for inputting setting conditions and the like by a user are provided as main components.

  The microscope body 101 includes a light source 111, a beam splitter 112, a scanning system 113, an objective lens 114, a stage 116, a grating (spectral element) 117, and a PMT (light detector) 118.

  The light source 111 emits laser light. This laser light excites a fluorescent indicator in the specimen 115 to generate fluorescence.

  The beam splitter 112 transmits the laser light from the light source 111, and reflects the fluorescence generated in the sample 115 and collected by the objective lens 114. By having such a configuration, the beam splitter 112 branches the optical path of the laser light and the optical path of the fluorescence from the specimen 115.

  The scanning system 113 has, for example, a pair of galvanometer mirrors (not shown) coated with aluminum, and is driven by a raster scan method by changing the angle of the pair of galvanometer mirrors. Thereby, the laser beam from the light source 111 is scanned two-dimensionally on the specimen 115.

  The objective lens 114 irradiates the laser beam scanned by the scanning system 113 onto the sample 115, while condensing the fluorescence generated from the sample 115.

  The grating 117 divides the fluorescence generated in the sample 115 and branched by the beam splitter 112 into spectral components for each wavelength, and causes the spectral components to be incident on the PMT 118.

  The PMT 118 is, for example, a 32-channel photomultiplier tube, and a plurality of channels for detecting spectral components are arranged in the direction in which the spectrum is dispersed by the grating 117. By having such a configuration, the PMT 118 detects the spectral components dispersed by the grating 117 by each channel, converts the luminance information of the detected spectral components into an electrical signal, and outputs the electrical signal to the signal adjustment unit 201. It is like that.

  The signal adjustment unit 201 includes an integrator (integrating unit) 201 that integrates the luminance information of the fluorescence detected by the PMT 118, an offset amplifier (amplifying unit) 212 that amplifies the luminance information of the fluorescence detected by the PMT 118 by giving an offset, And an A / D converter 213 for converting fluorescence luminance information detected by the PMT 118 into a digital signal.

  As shown in FIG. 2, the signal adjustment unit 201 is provided with an integrator 211, an offset amplifier 212, and an A / D converter 213 for each channel of the PMT 118, and is converted by the A / D converter 213. A digital signal is output to the CPU 311 of the control unit 301.

  The control unit 301 includes a memory (storage unit) 312 that stores various types of information, a multiplexer 313 that adds up digital signals from the A / D converter 213, and a CPU 311 that controls them, and a laser light source 111. , A signal for controlling each of the scanning system 113, the integrator 211, the offset amplifier 212, and the A / D converter 213 is generated.

  Further, the control unit 301 divides the channels of the PMT 118 into a plurality of groups (channel groups) based on the information on the fluorescence wavelength range detected by the PMT 118 so that all the channels in the channel group have the same sensitivity. The channel sensitivity is collectively adjusted for each channel group. Here, as a specific adjustment method, the control unit 301 will be described as adjusting the integration time of the integrator 211 for each channel group.

  In addition, when the CPU 311 recognizes that the digital signal value output from the A / D converter 213 exceeds a preset setting value, the control unit 301 determines the integration time in the integrator 211 and the time at that time. A voltage applied to the PMT 118 (hereinafter referred to as “HV”) is stored in the memory 312 by the CPU 311.

The control unit 301 generates a scan image based on the fluorescence intensity calculated by the CPU 311.
The display unit 401 is a liquid crystal display, for example, and displays an image of the specimen 115 generated by the control unit 301.

  The input unit 501 includes, for example, a keyboard and a mouse, and acquires values and operations input by the user and outputs them to the CPU 311. The input unit 501 is configured to input, for example, sensitivity adjustment, pigment selection, and threshold setting by the user, and output the input information to the control unit 301.

  With this configuration, when the user adjusts the sensitivity of the PMT 118 using the input unit 501, the sensitivity of the PMT 118 is adjusted via the control unit 301. In addition, when the user inputs the dye selection by the input unit 501, the setting is sent to the CPU 311 in the control unit 301, and the CPU 311 determines and determines the total channel (channel group) corresponding to the selected dye. The summation processing of the spectral components detected by the channel is performed.

  Further, when the user inputs the threshold setting by the input unit 501, the setting is sent to the CPU 311 in the control unit 301. The CPU 311 sets the threshold value and the intensity of the spectral component detected by each channel of the PMT 118. Comparisons are made.

In the microscope 1 having the above configuration, an operation when acquiring a fluorescent image will be described below.
In the microscope body 101, the light emitted from the laser light source 111 passes through the beam splitter 112 and enters the laser scanning system 113. The laser scanning system 113 scans the laser light through the objective lens 114 in the X direction and the Y direction on the plane of the specimen 115 placed on the stage 116.

  On the focal plane of the specimen 115, the fluorescent material in the specimen 115 is excited by the laser light, and fluorescence is generated. The fluorescence from the specimen 115 is collected by the objective lens 114, passes through the laser scanning system 113, and is branched from the optical path of the laser light by the beam splitter 112.

  The branched fluorescence is split by the grating 117, and the spectral components thus split enter each channel of the PMT 118 for each wavelength. At this time, fluorescence from at least two fluorescent dyes enters the PMT 118.

  In each channel of the PMT 118, the luminance information is converted into an electric signal according to the detected intensity of the spectral component, and is output to the signal adjustment unit 201. In the signal adjustment unit 201, the luminance information of the spectral component is integrated by the integrator 211 for each channel of the PMT 118, and the luminance information of the spectral component is amplified by the offset amplifier 212. The luminance information subjected to such processing is converted into a digital signal by the A / D converter 213 and output to the CPU 311.

The digital signal received by the CPU 311 is added to the channel group corresponding to the fluorescent dye by the multiplexer 313 inside the CPU 311. Since the summed signal is generated for each fluorescent dye, there are a plurality of signals, and each becomes a luminance signal for one pixel displayed on the display unit 401.
By repeating the above and scanning with the laser scanning system 113, a fluorescent image is generated on the display unit 401.

Detailed control executed by the signal adjustment unit 201 and the control unit 301 in the above-described fluorescence image acquisition operation will be described below.
All signals output from the respective channels of the PMT 118 are continuously accumulated by the accumulator 211 during the time for irradiating the sample 115 with the laser light to obtain a signal of one pixel. The accumulated time is determined by a clock signal generated by the CPU 311 (hereinafter referred to as “PCLK signal”). For example, integration is performed while the PCLK signal is Low, and the value of the integrator 211 is set to 0 when the PCLK signal is High.

  An offset amplifier 212 is connected after the integrator 211, and an offset is given to the signal after integration. The signal given the offset is converted into a digital signal by the A / D converter 213.

  During integration, the signal from the integrator 211 continues to be output to the CPU 311 through the offset amplifier 212 and the A / D converter 213. Here, in the A / D converter 213, the analog signal from the offset amplifier 212 is converted into a digital signal with a conversion clock having a shorter cycle than the PCLK signal generated by the CPU 311, and the CPU 311 for each conversion clock. Output a digital signal.

The processing of the CPU 311 at this time will be described using the flowchart shown in FIG.
First, in step S1, it is determined whether or not the PCLK signal is low.
If the PCLK signal is low, a digital signal value is acquired by the CPU 311 in step S2.

Next, in step S3, it is determined whether or not the digital signal value is less than or equal to the set value for each channel. At this time, as shown in FIG. 5, when the digital signal value is equal to or smaller than the set value, the digital signal value when the PCLK signal becomes High, that is, when the integration time becomes t _max is output to the CPU 311. (Step S4).

  Here, this set value is a value that the CPU 311 determines that the digital signal is out of the conversion range of the A / D converter 213, and the user can change the conversion range via the input unit 501 before starting scanning. It is a value set by inputting what percentage.

The digital signals are summed by the multiplexer 313 in the CPU 311 in step S5. The detailed procedure at that time will be described with reference to the flowchart shown in FIG.
First, a correspondence table (hereinafter referred to as “LUT”) is stored in the memory 312 so that a channel group corresponding to the fluorescent dye is determined.

  When the user selects a fluorescent dye displayed on the display unit 401 via the input unit 501, the fluorescent dye used by the CPU 311 is recognized (step S5-1), and a channel group to be added is determined by referring to the LUT. (Step S5-2).

  Next, in step S5-3, the signals from the channel group are added to form a luminance signal for one pixel displayed on the display unit 401. At this time, since a luminance signal is generated for each fluorescent dye, these luminance signals are transmitted to the display unit 401 at the same time, but may be displayed in a superimposed manner or separately for each fluorescent dye.

In step S3 of FIG. 3, when a digital signal value equal to or higher than the set value is output to the CPU 311 while the PCLK signal is low, the conversion range of the A / D converter 213 is shown in FIG. determining the CPU311 became outside, the value of the integrated time _{t s} and HV at that time is stored in the memory 312 (step S6).

  If a digital signal value equal to or larger than the set value continues for 3 pixels in one line displayed on the display unit 401, that is, while the Y-axis value scanned on the specimen 115 is constant (step S7). Then, the average of the integration time for three pixels acquired by the CPU 211 is calculated (step S8).

  Then, by dividing and applying the PCLK signal to each accumulator 211 corresponding to the channel group to be summed from the next pixel, the average accumulated time calculated in step S8 is 1 after the next pixel. The pixel integration time is set (step S9). At this time, among the signals from the channel group, the integration time applied to the integrator 211 of the channel having the maximum luminance signal is applied to each integrator 211 of the channel group.

In step S10, the value accumulated in the integrator 211 is set to 0 by waiting until the PCLK signal becomes High.
Thereby, the intensity | strength of a luminance signal can be adjusted, maintaining the shape of the spectrum formed with the signal intensity | strength from each channel of a channel group.

After the user finishes inputting the control conditions (selection of HV value, offset value, fluorescent dye) via the input unit 501, the above processing is performed for at least one frame.
The display unit 401 may display a button for turning on / off the control function for performing steps S6 to S10 in FIG. 3, and may be switched by the user via the input unit 501.

  As described above, according to the microscope 1 according to the present embodiment, the fluorescence generated in the specimen 115 and collected by the objective lens 114 is spectrally divided into spectral components by the grating 117, and a plurality of spectral components are separated. Are detected by the PMT 118 having the following channels. In this case, the channels of the PMT 118 are divided into a plurality of groups (channel groups) by the control unit 301 based on information on the wavelength range of light to be detected, so that all the channels in the channel group have the same sensitivity. In addition, the sensitivity is collectively adjusted for each channel group.

  Thereby, the intensity | strength can be adjusted, maintaining a spectrum shape for every grouped channel group, ie, for every wavelength range of the light to detect. For example, even when observing a plurality of types of fluorescence at the same time, the intensity of each fluorescence can be individually adjusted without saturating the spectral components of each fluorescence, and can be observed with high accuracy. Further, it is possible to eliminate the necessity of individually adjusting the sensitivity of each channel, and the adjustment work can be facilitated.

  Also, by adjusting the integration time of the integrator 211 for each group by the control unit 301, the luminance information detected by each channel of the PMT 118 can be adjusted for each group, and the spectrum is detected for each wavelength range of light to be detected. The strength can be adjusted while maintaining the shape.

  Also, when any of the spectral components exceeds the set value, for example, when any of the spectral components is saturated, it is detected without saturating any of the spectral components by adjusting the sensitivity of the PMT 118 channel. The intensity of fluorescence can be adjusted.

  Note that the control unit 301 may adjust the voltage applied to the PMT 118 for each channel group. In this way, the channel sensitivity of the PMT 118 can be adjusted for each channel group, and the intensity can be adjusted while maintaining the spectral shape for each wavelength range of light to be detected.

  Further, the control unit 301 may adjust the amplification factor of the offset amplifier 212 for each channel group. In this way, the luminance information detected by each channel of the PMT 118 can be adjusted for each channel group, and the intensity can be adjusted while maintaining the spectral shape for each wavelength range of light to be detected. it can.

  Further, the sensitivity of each group of channels and the luminance information may be stored in the memory 312 in association with each other. In this way, the channel sensitivity can be reproduced for each wavelength range of light to be detected. Further, an image of the specimen 115 when all the channels have the same sensitivity can be generated from the luminance information for each wavelength range and its sensitivity.

[First Modification]
Below, the 1st modification of this embodiment is demonstrated using FIG.
The microscope 1 according to the present modification includes means for allowing the user to recognize that the integration time of the integrator 211 has been changed.
The processing procedure of the CPU 311 in this modification will be described below using the flowchart shown in FIG.

First, in step S8, the CPU 311 calculates an average of integration times for three pixels.
The average accumulated time is applied to the next pixel (step S9), and at the same time, an image for one frame generated based on the luminance information obtained by adapting the accumulated time displayed on the display unit 401. Is displayed in a different color (step S11).
By doing in this way, the user can recognize that the image is generated by the luminance signal whose integration time is adjusted.

[Second Modification]
Below, the 2nd modification of this embodiment is demonstrated using FIG.
The microscope 1 of the present modification enables the user to arbitrarily change the integration time of the integrator 211, and applies the input integration time to each integrator connected to a channel in a range corresponding to the fluorescent dye. So that it is controlled.

  Specifically, for example, a slide corresponding to the integration time is displayed on the display unit 401 for each fluorescent dye, and the user changes the slide via the input unit 501 so that the integration time corresponds to the position of the slide. Is to be determined.

The processing of the CPU 311 in this modification will be described using the flowchart shown in FIG.
When the user selects the fluorescent dye displayed on the display unit 401 via the input unit 501, the CPU 311 recognizes the fluorescent dye used (step S5-1), and the channel group to be added is determined by referring to the LUT. (Step S5-2).
Then, the integration time is acquired from the position of the slide (step S12), and this integration time is applied to each integrator 211 corresponding to the determined channel group (step S13).
In this way, it is possible to generate an image from the signal intensity arbitrarily adjusted by the user.

[Third Modification]
Below, the 3rd modification of this embodiment is demonstrated using FIG.
The microscope 1 according to the present modification stores the HV when the digital signal received by the CPU 311 when the integration time is the maximum integration time based on the PCLK signal, that is, the integration time is t _max , exceeds the set value for the first time. When the HV is changed during the laser scanning, the accumulated time is returned to t _max when the HV is lowered from the stored HV.

The microscope 1 of the present modification can adjust the sensitivity of the entire channel by the control unit 301 (overall sensitivity adjustment unit).
The memory 312 stores the maximum sensitivity of the entire channel such that any spectral component is saturated during the maximum integration time when the luminance information integrated by the integrator 211 is equal to or greater than a predetermined value.

  When the sensitivity of the entire channel is lower than the maximum sensitivity stored in the memory 312, the control unit 301 maximizes the integration time of the integrator 211 for each group of channels.

The HV value can be adjusted by the user changing the HV value displayed on the display unit 401 via the input unit 501.
When the maximum integration time based on the PCLK signal, that is, the integration time is t _max , the HV when the digital signal received by the CPU 311 is equal to or higher than the set value for the first time is stored in the memory 312.

The subsequent processing procedure by the CPU 311 will be described with reference to the flowchart shown in FIG.
When HV changes, the value is acquired (step S14) and compared with the stored HV value (step S15). If the value is larger than the stored HV value, the integration time is adjusted in steps S6 to S10 in FIG. If the value is smaller than the stored HV value, the original integrated time is restored by setting the integrated time to t _max (step S16).

By doing so, there is no change in the integration time when the HV is finely moved, and this is effective in the case of a small amount of sensitivity adjustment.
Moreover, the luminance information detected by each channel can be adjusted for each group by adjusting the integration time of the integrator 211 for each group by the control unit 301, and the spectrum shape can be changed for each wavelength range of light to be detected. The strength can be adjusted while maintaining. In this case, the integrated luminance information is obtained by maximizing the integration time of the integrator 211 for each group of channels when the sensitivity of the entire channel falls below the maximum sensitivity at which any spectral component is saturated. The S / N ratio can be improved.

[Fourth Modification]
Below, the 4th modification of this embodiment is demonstrated using FIG. 10 and FIG.
The microscope 1 of the present modification is configured to adjust the integration time only by the process selected from the user in steps S6 to S10 in the range selected by the user.
A processing procedure performed by the CPU 311 in this modification will be described with reference to the flowchart of FIG.

  As shown in FIG. 11, the CPU 311 recognizes a selection range arbitrarily selected by the user in the sample image displayed on the display unit 401 (step S <b> 17), and determines whether the scan point corresponds to a pixel in the selection range. (Step S18).

  If it is within the selection range, an enable signal is output to the CPU 311 so that the integration time is determined by the same processing as in steps S6 to S10 in FIG. 3 (step S19). On the other hand, if it is outside the selection range, the enable signal is not output, the same integration time as the previous pixel is applied, and the digital signal value is acquired by the CPU 311 (step S2). As a result, the integration time is determined only within the selected range, but the determined integration time is applied to the entire laser scanning range.

By doing in this way, a luminance signal can be acquired within the dynamic range of the signal adjusting means for the range that the user wants to focus on.
Further, a region of interest (selection range) is set by the input unit 501 on the screen displayed on the display unit 401, and the intensity of the spectrum is maintained while maintaining the spectrum shape for each wavelength range of light to be detected in the selection range. Can be adjusted. On the other hand, it is possible to eliminate the necessity of adjusting the intensity of the non-attention area in the screen displayed on the display unit 401.

As mentioned above, although embodiment of this invention was explained in full detail with reference to drawings, the specific structure is not restricted to this embodiment, The design change etc. of the range which does not deviate from the summary of this invention are included.
For example, in the present embodiment, the integration time of the integrator 211, the amplification factor of the offset amplifier 212, or the HV is adjusted for each channel group. However, the sensitivity of the channel of the PMT 118 is adjusted by arbitrarily combining these. It is good to do.

  Also, in step S7 of the flowchart shown in FIG. 3, the integration time of the integrator 211 has been described as being changed when a digital signal value equal to or greater than the set value continues for 3 pixels. It is good also as changing when it continues.

DESCRIPTION OF SYMBOLS 1 Microscope 101 Microscope main body 111 Light source 112 Beam splitter 113 Scanning system 114 Objective lens 115 Sample 116 Stage 117 Grating (spectral element)
118 PMT (photodetector)
201 signal adjustment unit 211 accumulator (integration unit)
212 Offset amplifier (amplifier)
213 A / D converter 301 Control unit (sensitivity adjustment unit, image generation unit)
311 CPU
312 Memory (storage unit)
313 Multiplexer 401 Display unit 501 Input unit (input unit, setting unit)

Claims (9)

An objective lens for collecting the light generated in the specimen;
A spectroscopic element that splits light collected by the objective lens into spectral components;
A photodetector having a plurality of channels that respectively detect spectral components dispersed by the spectroscopic element;
A sensitivity adjuster for adjusting the sensitivity of each channel of the photodetector;
The sensitivity adjustment unit divides the channels into a plurality of groups based on information on the wavelength range of light to be detected, and sets the channel sensitivity for each group so that all the channels in the group have the same sensitivity. A microscope that adjusts all at once.   The microscope according to claim 1, wherein the sensitivity adjustment unit adjusts a voltage applied to the photodetector for each group of the channels. An integrating unit that integrates luminance information of light detected by the photodetector;
The microscope according to claim 1, wherein the sensitivity adjustment unit adjusts an integration time of the integration unit for each group of the channels. An amplifying unit that amplifies luminance information of light detected by the photodetector;
The microscope according to claim 1, wherein the sensitivity adjustment unit adjusts an amplification factor of the amplification unit for each group of the channels.   The microscope according to any one of claims 1 to 4, further comprising an input unit capable of setting a sensitivity adjustment value of the sensitivity adjustment unit for each group of the plurality of channels. An overall sensitivity adjuster for adjusting the sensitivity of the entire channel;
An integration unit for integrating luminance information of light detected by the photodetector;
A storage unit that stores the maximum sensitivity of the entire channel such that any spectral component is saturated in a maximum integration time in which the luminance information integrated by the integration unit is equal to or greater than a predetermined value;
The sensitivity adjustment unit maximizes the integration time of the integration unit for each group of channels when the sensitivity of the entire channel is lower than the maximum sensitivity stored in the storage unit. Microscope.   The microscope according to any one of claims 1 to 6, wherein when one of the spectral components detected by the channel exceeds a predetermined threshold, the sensitivity adjustment unit adjusts the sensitivity of the channel for each group. .   The microscope according to any one of claims 1 to 7, further comprising a storage unit that stores the sensitivity and luminance information of the channel for each group in association with each other. An image generator that generates an image based on the light detected by the photodetector;
A display unit for displaying an image generated by the image generation unit;
A setting unit for setting a predetermined range on the screen displayed on the display unit,
The microscope according to any one of claims 1 to 8, wherein the sensitivity adjustment unit adjusts the sensitivity of the channel within a predetermined range set by the setting unit.

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