JP2014202945A - Scanning laser microscope system - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a scanning laser microscope system which allows a sample to be observed on an image by accurately detecting light from the sample without being affected by light from a monitor.SOLUTION: A scanning laser microscope system 100 includes: a scanner 5 which two-dimensionally scans a sample S with laser light emitted by a light source 3; a detection unit 20 which detects light from the sample S scanned with the laser light by the scanner 5 and generates a light intensity signal representing brightness of the detected light; an image generation unit 35 which generates an image of the sample S using the light intensity signal output from the detection unit 20; a monitor 19 which displays the sample image generated by the image generation unit 35; and a wavelength selection unit 37 and a brightness modification unit 39, which make the monitor 19 emit color using a wavelength other than a wavelength detected by the detection unit 20.

Description

  The present invention relates to a scanning laser microscope system.

  2. Description of the Related Art Conventionally, there is known an apparatus that generates a sample image by detecting weak fluorescence from a sample with a microscope in a darkroom, and displays the image on a PC (Personal Computer) monitor (Patent Document 1 and Patent Document 2).

  The darkroom display control device described in Patent Document 1 discriminates between an image display area for displaying image data on a monitor and a background area for displaying other backgrounds, etc., and selectively masks the background area of the monitor. By making it dark, the influence of the light emitted from the monitor on the microscope in the darkroom is reduced. In addition, the microphotographing device described in Patent Document 2 reduces the influence of light emitted from the monitor on observation of the camera in a dark room by adjusting the brightness of the monitor in synchronization with the opening and closing of the shutter of the camera. doing.

JP 2009-251454 A JP-A-4-328712

  However, as in the dark room display control device described in Patent Document 1, when the background area of the monitor is only masked and selectively darkened, an ultrasensitive sensor such as GaAsP (Gallium Arsenide Phosphide) is used. Has the disadvantage that the microscope is affected by the light of the monitor. Further, when the monitor is turned on / off as in the microphotographing device described in Patent Document 2, it is impossible to confirm the sample being observed on the monitor while the monitor is turned off. There is an inconvenience.

  The present invention has been made in view of the circumstances described above, and is a scanning laser microscope system capable of accurately detecting light from a specimen and observing the specimen on an image without being affected by light from the monitor. The purpose is to provide.

In order to solve the above problems, the present invention employs the following means.
The present invention relates to a scanning unit that two-dimensionally scans a sample with laser light emitted from a light source, and detects light from the sample scanned with laser light by the scanning unit, and corresponds to the luminance of the detected light. A detection unit that outputs a light intensity signal to be output, an image generation unit that generates a sample image based on the light intensity signal output from the detection unit, and an image that displays the sample image generated by the image generation unit Provided is a scanning laser microscope system including a display unit and a control unit that causes the image display unit to develop color at a wavelength other than the wavelength of light detected by the detection unit.

  According to the present invention, when laser light is emitted from a light source, the scanning unit scans the laser beam two-dimensionally on the sample, and the detection unit detects light from the sample and outputs a light intensity signal. . Then, an image of the sample is generated by the image generation unit based on the light intensity signal, and the image of the sample is displayed on the image display unit. Thereby, the scanning range of the laser beam in the sample can be observed on the image of the image display unit.

  In this case, when detecting weak fluorescence generated in the specimen by irradiating laser light in a dark room or the like, the detection unit detects fluorescence or the like due to the light emitted from the image display unit. Efficiency may be reduced. In contrast, according to the present invention, the control unit causes the image display unit to develop a color other than the wavelength of the light detected by the detection unit, so that the weak light generated by the detection unit is affected by the light emitted from the image display unit. It is possible to prevent the detection efficiency from being reduced. Therefore, it is possible to accurately detect the light from the sample and observe the sample on the image without being affected by the light of the image display unit.

  In the above configuration, the control unit changes the color of the image display unit to a wavelength other than the wavelength of the light to be detected by the detection unit in conjunction with the timing at which the scanning unit scans the sample with the laser beam. It is good to do.

  With this configuration, during the blanking period in which the generation of the laser beam from the light source is stopped and the scanning line of the laser beam is returned to the original position by the scanning unit, the image display unit is colored in a color that is easy for the user to see. Can be made. Thereby, for example, when performing time-lapse observation, it is possible to make it easier for the user to see the sample image, information, and the like displayed on the image display unit during the interval between observations.

In the above configuration, the control unit may cause the image display unit to develop a color with the same wavelength as the laser beam emitted from the light source.
With this configuration, the control unit controls the color of the image display unit according to the wavelength of the laser light generated from the light source, and the light from the sample is not affected by the light of the image display unit. It can be detected with high accuracy by the detector.

  In the above configuration, the detection unit includes a plurality of detection units capable of detecting light of different wavelengths, and the control unit includes all wavelengths other than the wavelengths of light to be detected by the detection units, or the detection units. The image display unit may be colored with any wavelength selected from wavelengths other than the wavelength of the light to be detected.

  By configuring in this way, it is possible to control a plurality of light from the specimen without being affected by the light of the image display unit only by controlling the color development of the image display unit according to a wavelength other than the wavelength of the light detected by the detection unit Can be detected with high accuracy.

  In the above-described configuration, the light source includes a plurality of light sources that generate laser beams having different wavelengths, and the control unit includes all wavelengths of laser beams emitted from the light sources or lasers emitted from the light sources. The image display unit may be colored with any wavelength selected from light wavelengths.

  With this configuration, the control unit controls the color development of the wavelength image display unit according to the wavelength of the laser light generated from the light source, and images fluorescence of different wavelengths from the specimen generated for each light source. The detection unit can accurately detect the light without being affected by the light of the display unit.

  According to the present invention, there is an effect that light from a specimen can be accurately detected and the specimen can be observed on an image without being affected by light from the monitor.

1 is a schematic configuration diagram of a scanning laser microscope system according to a first embodiment of the present invention. It is a block diagram explaining the detection part of FIG. 2 is a flowchart for explaining specimen observation by the scanning laser microscope system of FIG. 1. It is a figure which shows an example of the display of the monitor of FIG. It is a block diagram explaining a part of detection part of FIG. 6 is a flowchart for explaining another example of specimen observation by the scanning laser microscope system of FIG. It is a block diagram explaining the detection part of the scanning laser microscope system which concerns on the 1st modification of 1st Embodiment of this invention. FIG. 8 is a block diagram specifically illustrating a detection unit in FIG. 7. It is a block diagram explaining the detection part of the scanning laser microscope system which concerns on the 2nd modification of 1st Embodiment of this invention. FIG. 10 is a block diagram specifically illustrating the detection unit of FIG. 9. It is a block diagram explaining the scanning laser microscope system which concerns on 2nd Embodiment of this invention. 12 is a flowchart for explaining specimen observation by the scanning laser microscope system of FIG. It is a block diagram explaining the detection part of the scanning laser microscope system which concerns on the 1st modification of 2nd Embodiment of this invention. It is a block diagram explaining the detection part of the scanning laser microscope system which concerns on the 2nd modification of 2nd Embodiment of this invention. It is a figure which shows an example of the display of the monitor of the scanning laser microscope system which concerns on 3rd Embodiment of this invention.

[First Embodiment]
Hereinafter, a scanning laser microscope system according to a first embodiment of the present invention will be described with reference to the drawings.
As shown in FIG. 1, a scanning laser microscope system 100 according to the present embodiment includes a stage 1 on which a specimen S is placed, a light source 3 that emits laser light, and laser light emitted from the light source 3 on the specimen S. The scanner (scanning unit) 5 for two-dimensionally scanning (scanning), the pupil projection lens 7 for condensing the laser beam scanned by the scanner 5, and the laser beam condensed by the pupil projection lens 7 are reflected. An excitation dichroic mirror (DM) 9 and an objective lens 11 that irradiates the sample S with the laser light reflected by the dichroic mirror 9 and collects return light returning from the sample S are provided.

  The light source 3 includes a laser diode (LD, not shown) that generates laser light, and an acousto-optic element (not shown) that performs wavelength selection of the laser light emitted from the laser diode. As the acoustooptic device, an AOTF (Acousto-Optic Tunable Filter), an AOM (Acousto-Optic Modulator), or the like can be used.

For the scanner 5, for example, a galvanometer mirror can be used.
The excitation dichroic mirror 9 reflects the laser light transmitted through the pupil projection lens 7 while allowing the fluorescence generated in the sample S and collected by the objective lens 11 to pass therethrough.

  Further, the scanning laser microscope system 100 includes a condensing lens 13 that condenses the fluorescence from the specimen S that has passed through the excitation dichroic mirror 9 and a reflecting mirror 15 that reflects the laser light collected by the condensing lens 13. And detecting the fluorescence reflected by the reflection mirror 15 and outputting a light intensity signal corresponding to the luminance of the detected light, and converting the light intensity signal output from the detection unit 20 from an analog signal to a digital signal. An A / D converter 17 for A / D conversion, a PC (Personal Computer) 30 for processing the A / D converted light intensity signal, an image of the sample S, and an instruction from the user are input A monitor (image display unit) 19 is provided.

  As shown in FIG. 2, the detection unit 20 reflects the fluorescence from the first dichroic mirror (first DM) 21 </ b> A and the second dichroic mirror (second DM) 21 </ b> B that branches the fluorescence from the incident specimen S into two optical paths. Reflecting mirror 23, and first barrier filter (first BF) 25A and second barrier filter (first) that block wavelength components other than a predetermined wavelength among the fluorescence reflected by these dichroic mirrors 21A and 21B and reflecting mirror 23. 2BF) 25B and a third barrier filter (third BF) 25C, and a first detector 27A, a second detector 27B, and a third detector 27C that detect fluorescence that has passed through these barrier filters 25A, 25B, and 25C. .

  The first dichroic mirror 21A reflects the fluorescence incident on the detection unit 20 toward the first barrier filter 25A according to the wavelength, and transmits it to the second dichroic mirror 21B. The second dichroic mirror 21B reflects the fluorescence transmitted through the first dichroic mirror 21A toward the second barrier filter 25B according to the wavelength, and transmits the reflected light to enter the reflection mirror 23. The reflection mirror 23 reflects the fluorescence transmitted through the second dichroic mirror 21B toward the third barrier filter 25C.

  As the detectors 27A, 27B, and 27C, for example, a PMT (Photo Multiplier Tube) can be used. Each detector 27A, 27B, and 27C photoelectrically converts the detected light and outputs a light intensity signal having a magnitude corresponding to the amount of light.

  The PC 30 includes an illumination control unit 31 that controls the acoustooptic device of the light source 3, an observation condition setting unit 33 that sets observation conditions, and a light intensity signal sent from the A / D conversion unit 17. An image generation unit 35 that generates an image, a wavelength selection unit (control unit) 37 that selects a wavelength for coloring the monitor 19, and a luminance change that changes the luminance of the monitor 19 based on the wavelength selected by the wavelength selection unit 37 Unit (control unit) 39.

The illumination control unit 31 selects a laser beam generated from the light source 3 using an acoustooptic device.
The observation condition setting unit 33 sets the wavelength of the laser light generated from the light source 3 by the illumination control unit 31 and the barrier filter 25A used in the detection unit 20 as the observation condition in accordance with a user instruction input to the monitor 19. Settings such as 25B and 25C are made.

  The wavelength selector 37 selects a wavelength other than the wavelength of the light detected by the detector 20. Specifically, the wavelength selection unit 37 selects a color pattern of the monitor 19 from, for example, “Dye” and “Laser” in accordance with a user instruction input to the monitor 19. “Dye” is a setting that causes the monitor 19 to develop a color at a wavelength that does not affect the staining state of the specimen S, and “Laser” is a setting that causes the monitor 19 to develop a color at the same wavelength as the laser light that irradiates the specimen S. .

  In addition, when the color development pattern of the monitor 19 is “Dye”, the wavelength selection unit 37 determines the barrier filter 25A (25B) based on the value of the barrier filter 25A (25B, 25C) set by the observation condition setting unit 33. 25C), that is, the wavelength of light blocked by the barrier filter 25A (25B, 25C) is calculated.

Further, when the color development pattern of the monitor 19 is “Laser”, the wavelength selection unit 37 calculates the wavelength (excitation wavelength) of the laser light generated from the light source 3 set by the observation condition setting unit 33. Yes.
Further, the wavelength selector 37 sends the calculated wavelength to the luminance changing unit 39.

  The brightness changing unit 39 displays the monitor 19 with the wavelength transmitted from the wavelength selecting unit 37. Thereby, the brightness changing unit 20 can color the monitor 19 with a wavelength other than the wavelength of the light detected by the detecting unit 20.

The operation of the scanning laser microscope system 100 according to the present embodiment configured as described above will be described.
When the specimen S is observed by the scanning laser microscope system 100 according to the present embodiment, first, as shown in the flowchart of FIG. 3, the user displays “monitor color display on the display screen of the monitor 19 as shown in FIG. “Mode switching” is turned ON, and the monitor luminance conversion mode is selected (step SA1). At this time, a function of “return monitor color development after scanning to the condition before scanning” on the monitor 19 may be activated. As a dye for staining the specimen S, for example, DAPI or the like can be used.

  Subsequently, the user selects the color pattern of the monitor 19 on the monitor 19 from “Dye” or “Laser” (step SA2). Next, the user selects the wavelength of the laser light generated from the barrier filter 25A (25B, 25C) to be used or the light source 3 on the monitor 19. Thereby, an observation condition is set by the observation condition setting unit 33 (step SA3).

  In step SA2, for example, when the user selects the “Dye” coloring pattern, the wavelength selection unit 37 monitors the value based on the value of the barrier filter 25A (25B, 25C) set by the observation condition setting unit 33. The wavelength for developing 19 is calculated (step SA4). The wavelength calculated by the wavelength selection unit 37 is sent to the luminance change unit 39.

  For example, as shown in FIG. 5, when the set first barrier filter 25 </ b> A has a characteristic of transmitting only fluorescence in the wavelength region of 430 to 455 nm, the wavelength that causes the monitor 19 to develop color is 430 nm or less by the wavelength selection unit 37. Or it is calculated as 455 nm or more.

  Next, when the user gives an instruction to start scanning on the monitor 19 (step SA5), the brightness changer 39 monitors the wavelength calculated by the wavelength selector 37 (wavelength of 430 nm or less or wavelength of 455 nm or more in the example of FIG. 5). 19 is developed (step SA6). In this case, the user may select whether the monitor 19 develops a color with a wavelength of 430 nm or less or a color with a wavelength of 455 nm or more. In this state, the sample S is observed by generating laser light from the light source 3.

  The laser light emitted from the light source 3 is condensed by the pupil projection lens 7 via the scanner 5 and reflected by the excitation dichroic mirror 9, and then condensed by the objective lens 11 and applied to the sample S. Thereby, the laser beam is scanned two-dimensionally on the specimen S according to the swing angle of the scanner 5.

  When fluorescence is generated in the specimen S by being irradiated with the laser light, the fluorescence is condensed by the objective lens 11 and transmitted through the dichroic mirror 9, and then condensed by the condenser lens 13 and via the reflection mirror 15. The light enters the detection unit 20. The fluorescence incident on the detection unit 20 is reflected by the dichroic mirror 21A (21B) or the reflection mirror 23, passes through the barrier filter 25A (25B, 25C), and is detected by the detector 27A (27B, 27C).

  When the fluorescence is detected by the detector 27A (27B, 27C), a light intensity signal corresponding to the luminance of the fluorescence is sent from the detector 27A (27B, 27C) to the A / D converter 17. The intensity signal is A / D converted by the A / D converter 17 and sent to the image generator 35. The image generator 35 generates an image of the sample S based on the light intensity signal and displays it on the monitor 19. Is done.

Thereby, the user can confirm the scan status of the specimen S on the monitor 19 (step SA7).
When the scanning is completed, the luminance changing unit 39 returns the color of the monitor 19 to the default luminance and displays an image, information, and the like (step SA8).

  In this case, when the detection unit 20 tries to detect weak fluorescence generated in the sample S irradiated with laser light in a dark room or the like, the detection unit 20 detects fluorescence or the like due to the influence of light emitted from the monitor 19. Efficiency may be reduced. On the other hand, according to the scanning laser microscope system 100 according to the present embodiment, the wavelength selection unit 37 and the luminance change unit 39 cause the monitor 19 to develop color at a wavelength other than the wavelength of the light detected by the detection unit 20. It is possible to prevent the detection efficiency of weak fluorescence by the detection unit 20 from being reduced due to the influence of light emitted from the monitor 19. Accordingly, it is possible to accurately detect the light from the sample S without being affected by the light of the monitor 19 and observe the sample S on the image.

  In the present embodiment, the case where the “Dye” color pattern is selected in step SA2 of the flowchart of FIG. 3 has been described. For example, the “Laser” color pattern may be selected in step SA2. In this case, as shown in the flowchart of FIG. 6, the wavelength selection unit 37 calculates the wavelength for coloring the monitor 19 based on the wavelength of the laser light generated from the light source 3 set by the observation condition setting unit 33 ( Step SB4), and sent to the luminance changing section 39.

  For example, as shown in FIG. 5, when the wavelength of the laser light generated from the light source 3 is 405 nm, the wavelength for coloring the monitor 19 is calculated by the wavelength selection unit 37 as 405 nm (step SB4). Therefore, when the user gives an instruction to start scanning on the monitor 19 (step SA5), the luminance changing unit 39 causes the monitor 19 to develop a color at a wavelength of 405 nm calculated by the wavelength selecting unit 37 (step SA6).

  Also in this case, the wavelength selection unit 37 and the luminance change unit 39 simply control the color development of the monitor 19 in accordance with the wavelength of the laser light generated from the light source 3, and from the sample S without being affected by the light of the monitor 19. Can be accurately detected by the detection unit 20.

This embodiment can be modified as follows.
In the present embodiment, the detection unit 20 including the detectors 27A, 27B, 27C and the like has been described as an example of the detection unit. However, as a first modification, for example, as illustrated in FIG. A dichroic mirror (DM) 21 that branches the fluorescence into two optical paths, a first barrier filter (first BF) 25A that blocks wavelength components other than a predetermined wavelength among the fluorescence reflected by the dichroic mirror 21, and a first barrier A first detector 27A that detects fluorescence that has passed through the filter 25A, a second barrier filter (second BF) 25B that blocks wavelength components other than a predetermined wavelength among the fluorescence that has passed through the dichroic mirror 21, and a second barrier filter 25B It is good also as employ | adopting the detection part 41 provided with the 2nd detector 27B which detects the fluorescence which passed.

  Also in this modification, when the monitor 19 is colored with the “Dye” coloring pattern, for example, as shown in FIG. 8, the wavelength range (see FIG. 8) that passes through the barrier filter 25A (25B) set by the observation condition setting unit 33 ( For example, in the case of the first barrier filter 25D, a wavelength other than 420-460 nm (wavelength of 420 nm or less or wavelength of 460 nm or more) is calculated by the wavelength selection unit 37, and the luminance change unit 39 causes the monitor 19 to develop color at that wavelength. You can do that.

  When the monitor 19 is colored with the “Laser” color pattern, the wavelength of the laser beam generated from the light source 3 set by the observation condition setting unit 33 is calculated by the wavelength selection unit 37, and the wavelength is changed by the luminance changing unit 39. Then, the monitor 19 may be colored.

  Next, as a second modification, as a detection unit, for example, as shown in FIG. 9, a dichroic mirror 21 </ b> C that splits fluorescence from an incident sample S into two optical paths, a first barrier filter 25 </ b> A, and a second barrier filter The detection unit 42 including the barrier filter 25B and the first detector 27D and the second detector 27E such as a high-sensitivity detector that detects the fluorescence that has passed through the first barrier filter 25A and the second barrier filter 25B is employed. It is good.

  Moreover, as a 3rd modification, as shown in FIG. 10, as a detection part, for example, as shown in FIG. 10, the reflection mirror 23 which reflects the incident fluorescence, and the diffraction grating 24 which disperses the fluorescence reflected by the reflection mirror 23 for every wavelength. And a detector 43 including a slit 26 that passes only a part of the wavelength separated by the diffraction grating 24, for example, a wavelength of 430 to 455 nm, and a detector 27 that detects the fluorescence that has passed through the slit 26. Also good.

  Also in this case, when the monitor 19 is colored with the “Dye” coloring pattern, a wavelength other than the wavelength range (for example, 430 to 455 nm) passing through the slit 26 (wavelength of 430 nm or less or wavelength of 455 nm or more) is selected. It is only necessary to calculate by the unit 37 and cause the luminance changing unit 39 to develop the color of the monitor 19 at that wavelength.

  When the monitor 19 is colored with the “Laser” coloring pattern, the wavelength selection unit 37 calculates the wavelength (for example, 405 nm wavelength) of the laser light generated from the light source 3 set by the observation condition setting unit 33, and the luminance The monitor 19 may be colored with the wavelength by the changing unit 39.

[Second Embodiment]
Next, a scanning laser microscope system according to a second embodiment of the present invention will be described with reference to the drawings.
As shown in FIG. 11, the scanning laser microscope system 200 according to the present embodiment includes a light source 103 having a plurality of laser diodes (LD) 102A, 102B, and 102C that generate laser beams having different wavelengths as a light source. The wavelength selector 37 and the luminance changer 39 are all wavelengths other than the wavelengths detected by the detectors 27A, 27B, and 27C, or all the wavelengths of laser beams generated from the laser diodes 102A, 102B, and 102C. This is different from the first embodiment in that the monitor 19 is colored with a wavelength of.
In the following, portions having the same configuration as those of the scanning laser microscope system 100 according to the first embodiment are denoted by the same reference numerals and description thereof is omitted.

  The light source 103 includes, for example, a first laser diode (first LD) 102A that generates laser light having a wavelength of 405 nm, a second laser diode (second LD) 102B that generates laser light having a wavelength of 473 nm, and laser light having a wavelength of 635 nm. And a third laser diode (third LD) 102 </ b> C that is generated. These three laser diodes 102A, 102B, and 102C can generate laser beams simultaneously with each other.

  Instead of the dichroic mirrors 21A and 21B, the detection unit 20 includes dichroic mirrors 21D and 21E that reflect or transmit fluorescence and branch into two optical paths. The first dichroic mirror 21D reflects the incident fluorescence toward the first barrier filter 25A according to the wavelength, while transmitting the incident fluorescence to the second dichroic mirror 21E. The second dichroic mirror 21 </ b> E reflects the fluorescence transmitted through the first dichroic mirror 21 </ b> D toward the second barrier filter 25 </ b> B according to the wavelength, and transmits the reflected light to enter the reflection mirror 23.

  When the monitor 19 is colored with the “Dye” color pattern, the wavelength selection unit 37 sets the barrier filters 25A, 25B, 25C based on the values of the barrier filters 25A, 25B, 25C set by the observation condition setting unit 33. Wavelengths that do not affect 25C, that is, all wavelengths that are blocked by the barrier filters 25A, 25B, and 25C are calculated.

Further, when the monitor 19 is colored with the “Laser” color pattern, the wavelength selection unit 37 wavelength of the laser light emitted from each of the laser diodes 102A, 102B, 102C of the light source 103 set by the observation condition setting unit 33 All wavelengths are calculated.
As a dye for staining the specimen S, for example, DAPI, Alexa Fluor 488, Cy5 or the like can be used.

The operation of the thus configured scanning laser microscope system 200 will be described.
In the scanning laser microscope system 200 according to the present embodiment, when “monitor color display mode switching” is turned ON, for example, as shown in the flowchart of FIG. 12, “Dye” or “ In accordance with “Laser”, the wavelength selection unit 37 calculates the wavelength for coloring the monitor 19 based on the values of the barrier filters 25A, 25B, and 25C set by the observation condition setting unit 33 or the wavelength of the laser light generated from the light source 103. (Step SC4).

  For example, when the monitor 19 is colored with the “Dye” coloring pattern, the wavelength band (430 to 455 nm) passing through the first barrier filter 25A set by the observation condition setting unit 33 passes through the second barrier filter 25B. All wavelengths other than the wavelength range (490-590 nm) and the wavelength range (655-755 nm) passing through the third barrier filter 25C, that is, wavelengths of 430 nm or less, 455-490 nm, 590-655 nm, or 750 nm or more are wavelength selection units 37.

  Then, the luminance changing unit 39 causes the monitor 19 to develop a color at a wavelength calculated by the wavelength selecting unit 37, that is, a wavelength of 430 nm or less, 455-490 nm, 590-655 nm, or 750 nm or more (step SA6).

  On the other hand, when the monitor 19 is colored with the “Laser” coloring pattern, the wavelength of the laser light (405 nm) generated from the first laser diode 102A set by the observation condition setting unit 33 is generated from the second laser diode 102B. The wavelength selector 37 calculates all wavelengths of the laser light wavelength (473 nm) and the wavelength of the laser light generated from the third laser diode 102C (635 nm), that is, wavelengths of 405 nm, 473 nm, and 635 nm.

  Then, the luminance changing unit 39 causes the monitor 19 to develop color at the wavelengths calculated by the wavelength selecting unit 37, that is, wavelengths of 405 nm, 473 nm, and 635 nm (step SA6).

  As described above, according to the scanning laser microscope system 200 according to the present embodiment, the monitor 19 can be monitored only by controlling the color of the monitor 19 in accordance with a wavelength other than the wavelength of the light detected by the detectors 27A, 27B, and 27C. The light from the sample S can be accurately detected by the plurality of detectors 27A, 27B, and 27C without being affected by the 19 light beams. Further, only by controlling the color development of the monitor 19 in accordance with the wavelength of the laser light generated from the laser diodes 102A, 102B, and 102C, fluorescence of different wavelengths from the specimen S generated for each of the laser diodes 102A, 102B, and 102C is obtained. The detectors 27A, 27B, and 27C can accurately detect the light without being affected by the light of the monitor 19.

This embodiment can be modified as follows.
As a first modification, for example, as shown in FIG. 13, a detection unit 41 including a dichroic mirror 21, barrier filters 25A and 25B, and detectors 27A and 27B is used as a detection unit, and a first light source is used as a light source. The light source 104 including one laser diode 102D and the second laser diode 102E may be employed.

  Also in this modification, when the monitor 19 is colored with the “Dye” coloring pattern, the wavelength other than the wavelength range (420 to 460 nm) passing through the first barrier filter 25A set by the observation condition setting unit 33, and the first 2 Wavelengths other than the wavelength band (495-540 nm) passing through the barrier filter 25B, that is, wavelengths of 420 nm or less, 460-495 nm, and 540 nm or more are calculated by the wavelength selection unit 37, and the luminance change unit 39 monitors the wavelength 19 The color may be developed.

  When the monitor 19 is colored with the “Laser” color pattern, the wavelength of the laser light generated from the first laser diode 102D of the light source 104 set by the observation condition setting unit 33 and the laser generated from the second laser diode 102E are set. All the wavelengths of the light may be calculated by the wavelength selection unit 37, and the luminance change unit 39 may cause the monitor 19 to develop color at that wavelength.

  Moreover, as a 2nd modification, as shown in FIG. 14, it is good also as employ | adopting the light source 103 as a light source, for example. In addition, as a detection unit, the first dichroic mirror 21D, the first diffraction grating 24A that splits the fluorescence reflected by the first dichroic mirror 21D for each wavelength, and a part of the wavelengths (that are split by the first diffraction grating 24A) 430-455 nm), the first detector 26A, the first detector 27A, the second dichroic mirror 21E, the second diffraction grating 24B that splits the fluorescence reflected by the second dichroic mirror 21E for each wavelength, The second slit 26B, the second detector 27B, the reflecting mirror 23, the third barrier filter 25C, and the third detector 27C that allow a part of the wavelength (490-590 nm) dispersed by the two diffraction gratings 24B to pass therethrough are provided. It is good also as employ | adopting the detection part 120 which has.

  Also in this modification, when the monitor 19 is colored with the “Dye” coloring pattern, the wavelength range (430-455 nm) passing through the first slit 26A and the wavelength range (490-590 nm) passing through the second slit 26B. The wavelength selection unit 37 calculates all wavelengths other than the wavelength range (655-755) that passes through the third barrier filter 25C, that is, wavelengths of 430 nm or less, 455-490 nm, 590-655 nm, or 750 nm or more, and changes the luminance. The monitor 19 may be colored by the wavelength by the unit 39.

  When the monitor 19 is colored with the “Laser” color pattern, the wavelength of the laser light (405 nm) generated from the first laser diode 102A of the light source 103 set by the observation condition setting unit 33, the second laser diode 102B. The wavelength selection unit 37 calculates all wavelengths of the wavelength of the laser beam generated from the laser beam (473 nm) and the wavelength of the laser beam generated from the third laser diode 102C (635 nm), that is, the wavelengths of 405 nm, 473 nm, and 635 nm. The monitor 19 may be colored with the wavelength by the changing unit 39.

[Third Embodiment]
Next, a scanning laser microscope system according to a third embodiment of the present invention will be described with reference to the drawings.
In the scanning laser microscope system 200 according to the present embodiment, the wavelength selecting unit 37 and the luminance changing unit 39 select any wavelength other than the wavelength of light detected by the detectors 27A, 27B, and 27C, or each laser. The second embodiment is different from the second embodiment in that the monitor 19 is colored with any wavelength selected from laser beams generated from the diodes 102A, 102B, and 102C.
In the following, portions having the same configuration as those of the scanning laser microscope system 100 according to the first embodiment or the scanning laser microscope system 200 according to the second embodiment are denoted by the same reference numerals and description thereof is omitted.

  The wavelength selection unit 37 calculates the wavelength to be displayed on the monitor 19 in accordance with the wavelength selection method specified by the user on the display screen of the monitor 19 as shown in FIG. As the wavelength selection method, for example, in the case of FIG. 11, “refer to the value of the barrier filter” for selecting the values of the barrier filters 25A, 25B, and 25C, and the wavelength of the laser light irradiated to the specimen S are selected. The user designates from "Refer to excitation wavelength value".

  The wavelength used when there are a plurality of values of the selected barrier filters 25A, 25B, 25C or the wavelength of the laser light irradiated on the sample S is, for example, “use the short wavelength side”. ”,“ Use long wavelength side ”that uses the longest wavelength range, and“ use multiple wavelength side ”that use multiple wavelength ranges.

  In this case, as shown in FIG. 11, the wavelength of the laser light (405 nm) generated from the first laser diode 102A, the wavelength of the laser light (473 nm) generated from the second laser diode 102B, and the third laser diode 102C are generated. The wavelength selected by the user from the wavelength of the laser beam (635 nm) or a wavelength (430 nm or less, 455-490 nm, 590-655 nm, or 750 nm or more) other than the wavelength range that passes through the barrier filters 25A, 25B, and 25C. Calculation may be performed by the wavelength selection unit 37, and the luminance change unit 39 may cause the monitor 19 to develop color at that wavelength.

  This embodiment may be applied to the configuration shown in FIG. In this case, the wavelength of the laser light generated from the first laser diode 102D of the light source 104, the wavelength of the laser light generated from the second laser diode 102E, and a wavelength other than the wavelength range passing through the barrier filters 25A and 25B (420 nm or less, 460 Any wavelength selected by the user from the wavelengths of −495 nm and 540 nm or more may be calculated by the wavelength selection unit 37, and the luminance change unit 39 may cause the monitor 19 to develop a color.

  Further, the present embodiment may be applied to the configuration shown in FIG. In this case, the wavelength (405 nm) of the laser light generated from the first laser diode 102A of the light source 103, the wavelength (473 nm) of the laser light generated from the second laser diode 102B, and the wavelength of the laser light generated from the third laser diode 102C. (635 nm), any wavelength selected by the user from wavelengths (430 nm or less, 455-490 nm, 590-655 nm, or 755 nm or more) other than the wavelength range that passes through the slits 26A and 26B and the third barrier filter 25C. Calculation may be performed by the selection unit 37, and the luminance change unit 39 may cause the monitor 19 to develop color at that wavelength.

  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, the present invention is not limited to those applied to the above-described embodiments and modifications, but may be applied to embodiments in which these embodiments and modifications are appropriately combined, and is not particularly limited. .

  Further, for example, in each of the embodiments and the modifications thereof, the wavelength selection unit 37 and the luminance change unit 39 may change the color of the monitor 19 in synchronization with the scanning of the laser beam by the scanner 5. In this case, for example, during the scanning period in which the laser beam is scanned on the sample S by the scanner 5, the monitor 19 is colored by the luminance changing unit 39 at the wavelength selected by the wavelength selecting unit 37, and the light sources 3, 103, 104 are used. During the blanking period in which the generation of the laser beam is stopped and the scanning line of the laser beam is returned to the original position, the monitor 19 may be colored with the default luminance by the luminance changing unit 39. Further, for example, when performing time-lapse observation, the monitor 19 is colored by the luminance changing unit 39 at the wavelength selected by the wavelength selecting unit 37 during observation, and the monitor 19 is colored by the default luminance by the luminance changing unit 39 during the interval. It is also possible to make it.

3, 103, 104 Light source 5 Scanner (scanning unit)
19 Monitor (image display part)
20, 41, 42, 43, 120 detection unit 35 image generation unit 37 wavelength selection unit (control unit)
39 Brightness change unit (control unit)
100,200 Scanning laser microscope system

Claims (5)

A scanning unit that two-dimensionally scans a sample with laser light emitted from a light source;
A detection unit that detects light from a sample scanned with laser light by the scanning unit and outputs a light intensity signal corresponding to the luminance of the detected light;
An image generation unit that generates an image of a specimen based on the light intensity signal output from the detection unit;
An image display unit for displaying an image of the specimen generated by the image generation unit;
A scanning laser microscope system comprising: a control unit that causes the image display unit to develop a color at a wavelength other than the wavelength of light detected by the detection unit.   The control unit changes the color development of the image display unit to a wavelength other than the wavelength of light detected by the detection unit in conjunction with the timing of scanning the sample with the laser beam by the scanning unit. The scanning laser microscope system described.   The scanning laser microscope system according to claim 1, wherein the control unit causes the image display unit to develop a color with the same wavelength as the laser light emitted from the light source. A plurality of the detection units capable of detecting light of different wavelengths,
The control unit causes the image display unit to develop a color at any wavelength selected from all wavelengths other than the wavelengths of light detected by the detection units or from wavelengths other than the wavelengths of light detected by the detection units. The scanning laser microscope system according to claim 1 or 2. A plurality of light sources that generate laser beams having different wavelengths;
The control unit causes the image display unit to develop a color at any wavelength selected from all wavelengths of laser beams emitted from the light sources or from wavelengths of laser beams emitted from the light sources. Item 5. A scanning laser microscope system according to Item 4.

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