Microscope and microscope control method

Abstract

This microscope is provided with: a light source of which an ON state in which coherent light is emitted and an OFF state in which no light is emitted can be directly controlled; an illumination optical system which irradiates a sample with light to form an illumination region; an optical path changing member which changes the optical path of the light; and a control unit which controls the light source and the optical path changing member. The control unit controls the optical path changing member, switches between a first state in which light forms the illumination region on the sample when the light source is in the ON state and does not form the illumination region when the light source is in the OFF state and a second state in which light does not form an illumination region on a sample whether the light source is in the ON state or in the OFF state, and carries out control for setting the light source to be in the ON state in the second state.

Description

[Technical Field]

[0001] The present invention relates to a microscope and a method for controlling the microscope. [Background technology]

[0002] Patent Document 1 describes a scanning confocal microscope that scans a sample with an irradiating laser beam. [Prior art document] [Patent documents] [Patent Document 1] International Publication No. 2009 / 011441 [General Disclosure]

[0003] A first aspect of the present invention provides a microscope comprising: a light source that can be directly controlled between an ON state in which coherent light is emitted and an OFF state in which the light is not emitted; an illumination optical system that irradiates the light to form an illumination area on a sample; an optical path changing member that changes the optical path of the light; and a controller that controls the light source and the optical path changing member, wherein the controller controls the optical path changing member to switch between a first state in which the light forms the illumination area on the sample when the light source is in the ON state and does not form the illumination area when the light source is in the OFF state, and a second state in which the light does not form the illumination area on the sample whether the light source is in the ON state or the OFF state, and controls the light source to the ON state in the second state.

[0004] In the second state, the control unit may control the light source so that the ON state and the OFF state are repeated at predetermined timings.

[0005] The control unit may control the light source to the OFF state during at least one of a period during which the first state is switched to the second state and a period during which the second state is switched to the first state.

[0006] The illumination optical system may further include a condensing lens, and in the first state, the light passes through the condensing lens, and in the second state, the light does not pass through the condensing lens and enters a predetermined waiting position.

[0007] The image forming apparatus may further include a holding member that holds the condenser lens, and the standby position may be a part of the holding member.

[0008] A light blocking member may be provided in the part.

[0009] The light source may further include a reflecting member provided at the standby position, which reflects the light, and an absorbing member which absorbs at least a portion of the light reflected from the reflecting member.

[0010] The optical path changing member may be a galvanometer mirror or a spatial light modulator.

[0011] In the second state, the control unit may turn the light source to the ON state, wait for a predetermined time, and then perform control to switch the light to the first state.

[0012] The predetermined time may be a time required for the output value from the light source to stabilize.

[0013] A second aspect of the present invention provides a method for controlling a microscope, the method comprising: a light source that can be directly controlled between an ON state in which coherent light is emitted and an OFF state in which the light is not emitted; an illumination optical system that irradiates the light to form an illumination area on a sample; an optical path changing member that changes the optical path of the light; and a control unit that controls the light source and the optical path changing member, the method comprising a control step of controlling the optical path changing member to switch between a first state in which the light forms the illumination area on the sample when the light source is in the ON state and does not form the illumination area when the light source is in the OFF state, and a second state in which the light does not form the illumination area on the sample whether the light source is in the ON state or the OFF state, and setting the light source to the ON state in the second state.

[0014] The above summary of the invention does not list all of the necessary features of the present invention, and subcombinations of these features may also constitute inventions. [Brief explanation of the drawings]

[0015] [Figure 1] 1 shows a schematic configuration of a microscope 200 according to a first embodiment. [Figure 2] 1 shows a schematic configuration of a scan head 100 according to a first embodiment. [Figure 3] 1 shows an example of scanning by the scan head 100 according to the first embodiment. [Figure 4] 3 shows an ON / OFF timing chart of the laser light source 101 in the first embodiment. [Figure 5] 4 is a flowchart showing the operation of the scan head 100 in the first embodiment. [Figure 6] 1 shows a schematic configuration of a scan head 110 according to a second embodiment. [Figure 7] 10 shows a schematic configuration of a scan head 120 according to a third embodiment. [Figure 8] 10(a) and 10(b) show another example of ON / OFF control of the laser light source 101. [Figure 9] An example of a computer 2200 is shown. DETAILED DESCRIPTION OF THE INVENTION

[0016] The present invention will be described below through embodiments of the invention. The following embodiments do not limit the scope of the invention. Not all of the combinations of features described in the embodiments are necessarily essential to the solution of the invention.

[0017] FIG. 1 shows a schematic configuration of a microscope 200 according to the first embodiment. As shown in FIG. 1, the microscope 200 according to the first embodiment includes a scan head 100, a laser light source 101, a photodetector 102, a microscope body 103, and a control unit 104. The microscope 200 is a microscope that scans with a laser, and may be, for example, a confocal microscope. An xyz coordinate system is shown in the figure.

[0018] The laser light source 101 emits laser light, which is excitation light, toward the sample 106. The laser light source 101 is configured to be switchable between an ON state (hereinafter simply referred to as ON) in which the excitation light is emitted to the outside and an OFF state (hereinafter simply referred to as OFF) in which the excitation light is not emitted to the outside. Examples of the laser light source 101 include a semiconductor laser, a gas laser, and a solid-state laser. That is, the control unit 104 directly controls the laser light source 101 itself to switch it ON and OFF. This differs from a system in which, in a laser light source unit including a laser light source and a mechanical shutter or an AOTF, the laser light source continuously emits excitation light and the mechanical shutter or AOTF switches it ON and OFF. The sample 106 to be observed is placed on a stage of the microscope main body 103. The microscope main body 103 has a first objective lens 105a and a second objective lens 105b (hereinafter collectively referred to as an objective optical system 105).

[0019] The photodetector 102 receives the fluorescence emitted from the sample 106 using a photomultiplier tube or the like, converts it into an electrical signal, and outputs it to the control unit 104. The control unit 104 accumulates the input data in a computer, arranges the data to form a single image, and constructs an observed fluorescence image, which is then displayed on a display. The control unit 104 is connected to each component of the microscope 200 and can control each component. The control unit 104 can, for example, control the ON / OFF of the laser light source 101, the operation of the scan head 100, or the stage of the microscope main body 103.

[0020] Fig. 2 shows a schematic configuration of a scan head 100 according to the first embodiment. As shown in Fig. 2, the scan head 100 according to the first embodiment includes an input port 11, a collimating lens 12, a dichroic mirror 13, a galvanometer mirror 14, a scanning optical system 15, a lens barrel 16, a microscope port 17, a condenser lens 18, a pinhole 19, and an output port 20. The scan head 100 also includes other optical components such as a prism and a mirror that bend the optical path of the excitation light.

[0021] The input port 11 is a port through which excitation light, which is laser light from the laser light source 101, is incident on the scan head 100, and an optical fiber for the excitation light is connected to the input port 11. The collimating lens 12 converts the excitation light input from the input port 11 into parallel light. The dichroic mirror 13 reflects the excitation light that has been converted into parallel light by the collimating lens 12.

[0022] The galvanometer mirror 14 is an example of an optical path changing member that changes the optical path of the excitation light. The galvanometer mirror 14 is composed of a pair of mirrors, one that rotates the excitation light around the x-axis and the other that rotates it around the y-axis. The pair of galvanometer mirrors 14 reflects the angle of the excitation light reflected by the dichroic mirror 13 in any two orthogonal axial directions. Therefore, the galvanometer mirror 14 can scan the excitation light on the xy plane.

[0023] The scanning optical system 15 is a condensing lens that condenses the excitation light reflected by the galvanometer mirror 14. The scanning optical system 15 has an incident surface 15a onto which the excitation light is incident. The lens barrel 16 is a holding member that holds the scanning optical system 15, which is a condensing lens. The lens barrel 16 has a side surface 16a that is parallel to the xy plane. The side surface 16a is a surface that is disposed outside the incident surface 15a. A standby position 107 is provided on the side surface 16a of the lens barrel 16 for waiting the excitation light from the laser light source 101. In this embodiment, the excitation light is input via a microscope port 17 to an objective optical system 105 disposed downstream in the optical path. The scanning optical system 15 and the objective optical system 105 function as an illumination optical system that condenses the laser light to form an illumination area on a sample 106.

[0024] The excitation light incident on the incident surface 15a of the scanning optical system 15 is collected by the scanning optical system 15, output from the microscope port 17, and irradiated onto the sample 106 via the objective optical system 105, etc. On the other hand, the excitation light incident on a location other than the incident surface 15a of the scanning optical system 15, for example, on the side surface 16a of the lens barrel 16, is not output to the microscope port 17. Therefore, the excitation light incident on the standby position 107 provided on the side surface 16a of the lens barrel 16 is not output from the microscope port 17 and is not irradiated onto the sample 106.

[0025] As described above, the excitation light emitted from the laser light source 101 passes through the input port 11, collimating lens 12, dichroic mirror 13, galvanometer mirror 14, incident surface 15a of the scanning optical system 15, microscope port 17, and objective optical system 105 in this order, before being irradiated onto the sample 106. The sample 106 is exposed to the irradiated excitation light, and the excited fluorescent dye generates fluorescence of a specific wavelength. The fluorescence, which is the observation light, passes through the objective optical system 105 of the microscope main body 103 and enters the scan head 100 from the microscope port 17.

[0026] The incident fluorescence passes through the microscope port 17, scanning optical system 15, galvanometer mirror 14, and dichroic mirror 13, and is then converted into convergent light by the condenser lens 18. The pinhole 19 is disposed at a position conjugate to the focal position of the objective lens 105a, and blocks light from areas other than the conjugate position. The output port 20 is a port for outputting the fluorescence that has passed through the pinhole 19 to the photodetector 102. The fluorescence generated in the sample 106 is output from the output port 20 and input to the photodetector 102.

[0027] As described above, the fluorescence emitted from the sample 106 passes through the objective optical system 105, microscope port 17, scanning optical system 15, galvanometer mirror 14, dichroic mirror 13, condenser lens 18, pinhole 19, and output port 20, in that order, and is finally input to the photodetector 102. In the photodetector 102, the fluorescence emitted from the sample 106 is received by a photomultiplier tube or the like, converted into an electrical signal, and output to the control unit 104.

[0028] The control unit 104 controls the galvanometer mirror 14, which is an optical path changing member, to switch between a first state in which the laser light forms an illumination area on the sample 106 when the laser light source 101 is ON and does not form an illumination area when the laser light source 101 is OFF, and a second state in which the laser light does not form an illumination area on the sample 106 whether the laser light source 101 is ON or OFF, and controls the laser light source 101 to be ON in the second state. Specifically, the control unit 104 controls the galvanometer mirror 14, which is an optical path changing member, to switch between a first optical path (optical path in the first state) in which the excitation light is output from the microscope port 17 and a second optical path (optical path in the second state) in which the excitation light is not output from the microscope port 17. In FIG. 2, the control unit 104 controls the first optical path so that the excitation light is incident on the incident surface 15a of the scanning optical system 15. The control unit 104 controls the second optical path so that the excitation light is incident on a standby position 107 on the side surface 16a of the lens barrel 16. It should be noted that the first optical path is not a single optical path, but rather a collection of multiple optical paths when scanning the sample 106 in the X and Y directions.

[0029] FIG. 3 shows an example of excitation light scanning by the scan head 100 in the first embodiment. FIG. 3 shows the observable range (maximum observable range) 108 of the objective optical system 105 of the microscope 200. The observable range 108 in FIG. 3 is the range on the sample 106. The observable range 108 on the sample 106 varies depending on the magnification of the objective lens 105a. In this embodiment, excitation light scanning is performed along multiple lines within the observable range 108 of the objective optical system 105. In FIG. 3, multiple lines, such as the first line, the second line, the nth line, and so on, are indicated by thick solid lines. Scanning is performed sequentially from left to right along the thick solid lines indicating the multiple lines in FIG. 3. The left end points of each of the multiple lines are also referred to as scan start positions. FIG. 3 also shows a standby position 107. The standby position 107 in FIG. 3 is a schematic representation of the standby position 107 provided on the side surface 16a of the lens barrel 16 in FIG. 2. In addition, optically, the side surface 16 a of the lens barrel 16 corresponds to the outside of the observable range 108 of the objective optical system 105 .

[0030] In the first embodiment, when the system of the microscope 200 is started, first the galvanometer mirror moves (rotates by a predetermined angle) so that the excitation light emitted from the laser light source 101 is directed toward the standby position 107 in Fig. 3. After the galvanometer mirror moves, the laser light source 101 is turned on to irradiate the standby position 107 with the excitation light, and the system waits until the output value of the laser light source 101 stabilizes. The excitation light irradiated toward the standby position 107 is not irradiated onto the sample 106.

[0031] Depending on the type of laser light source 101, after the laser light source 101 is turned on and the output value (intensity of the excitation light) reaches a preset value, it may take several milliseconds to several seconds for the output value to actually stabilize. For laser light sources 101 that require several milliseconds for the output value to stabilize, there is no problem in waiting for several milliseconds until the output value stabilizes. On the other hand, when using a laser light source 101 that requires several seconds for the output value to stabilize, if the laser light source 101 is completely turned off and then turned on every time the observation position of the sample 106 is changed or the sample 106 is replaced, and then several seconds are waited until the output value stabilizes, this causes a problem because the sequentiality of the observation is lost. Furthermore, if the wait is not performed, unexpected changes in the brightness of the image obtained by the observation may occur, making quantitative analysis impossible.

[0032] In contrast, in this embodiment, before irradiating the sample 106 with excitation light, the laser light source 101 is turned on after the standby position 107 is irradiated with the excitation light, and then the system waits until the output value from the laser light source 101 stabilizes. Therefore, irradiation of the sample 106 with excitation light can be started with the output value of the laser light source 101 stabilized, solving the above problem, and the brightness of images resulting from each capture can be acquired, displayed, and saved with a stable state. Note that immediately after system startup, after the standby position 107 is irradiated with the excitation light, the laser light source 101 is turned on and a wait of several seconds is performed to wait until the output value of the laser light source 101 stabilizes. The wait of several seconds is required only once, immediately after startup of the microscope 200, and thereafter, unless the laser light source 101 is turned off, a wait of several seconds is not required.

[0033] At the timing when scanning with the excitation light begins, the excitation light is scanned from the standby position 107 to the scanning start position, and the irradiation position of the excitation light is moved. Note that point 301 at the top left of the first line in FIG. 3 is the scanning start position, and the coordinates of point 301 are (Sx, Sy). Point 306 at the bottom right is the scanning end position, and the coordinates of point 306 are (Sx+m-1, Sy+n-1). m is the amount of movement in the X direction (m is an integer greater than or equal to 1), and n is the amount of movement (number of lines) in the Y direction (n is an integer greater than or equal to 1). The coordinates of the standby position 107 are (a, b). The coordinates of this standby position 107 are the position when projected directly onto the sample 106 without passing through the objective optical system 105. Here, a and b are values ​​that result in a position outside the observable range 108 of the objective optical system 105. Here, during the short time it takes for the excitation light to scan from the standby position 107 to the scan start position 301, the laser light source 101 is turned off.

[0034] At the timing when the excitation light has completed moving to the scanning start position 301, the laser light source 101 is turned ON again, the excitation light is scanned for one line in the +x direction, and the irradiation position of the excitation light is moved to point 302 (Sx+m-1, Sy) on the right side of the first line, thereby obtaining observation data of the sample 106. This observation data is stored in a memory or the like within the control unit 104.

[0035] When the irradiation position of the excitation light reaches point 302 (Sx+m-1, Sy) on the right side of the first line, the laser light source 101 is turned off, and the irradiation position of the excitation light is turned back and moved to point 303 (Sx, Sy+1) on the left side of the second line. The laser light source 101 is kept turned off until it moves to point 303 (Sx, Sy+1). In this way, as with the above, unnecessary light is not irradiated onto the sample 106 during the turning operation that does not accumulate data, thereby reducing phototoxicity and damage to the sample 106. Thereafter, scanning is similarly performed up to the third line, fourth line, and so on up to the nth line.

[0036] When the excitation light reaches the scan end position 306 (Sx+m-1, Sy+n-1) of the nth line, which is the final line, the excitation light is moved from the scan end position 306 to the standby position 107. The laser light source 101 is turned OFF during the short time it takes to move from the scan end position 306 to the standby position 107, and after the excitation light has completely moved to the standby position 107, the laser light source 101 is turned ON again and left to wait for a while, allowing the output value of the laser light source 101 to stabilize.

[0037] As described above, in order to suppress phototoxicity and damage to the sample 106, the laser light source 101 is turned off for short periods of time while the excitation light moves from the standby position 107 to the scan start position 301, during turnaround times such as from point 302 to point 303, and during movement from the scan end position 306 to the standby position 107. Although the output value of the laser light source 101 may fluctuate slightly when the laser light source 101 is turned off for a short period of time, the effect on the output value of the laser light source 101 is limited because the time during which the laser light source 101 is turned off is short. When using a sample 106 that is less susceptible to phototoxicity and damage, or a sample 106 that is not susceptible to phototoxicity and damage, the laser light source 101 does not need to be turned off during the above periods. Furthermore, the laser light source 101 may be turned off only during one or two of the three movement times.

[0038] FIG. 4 shows a timing chart of the ON / OFF control of the laser light source 101 in the first embodiment. The three solid lines in FIG. 4 indicate, from top to bottom, the x- and y-coordinates of the excitation light irradiation position and the ON / OFF control of the laser light source 101. The x- and y-coordinates in FIG. 4 correspond to those in FIG. 3. At time t0 when the system is started up, the excitation light irradiation position is at its initial position (location is undefined). Thereafter, at time t1, the irradiation position moves to standby position 107 (a, b). This is outside the observable range 108 of the objective optical system 105, and even if the laser light source 101 is output at this position, it will not irradiate the sample 106. For a predetermined time, from time t1 to time t2, the laser light source 101 is turned on and waits at standby position 107 until the output value of the laser light source 101 stabilizes.

[0039] At time t2, the laser light source 101 is turned OFF, and then the irradiation position of the excitation light is moved to point 301 (Sx, Sy), which is the scanning start position. Thereafter, at time t3, the laser light source 101 is turned ON to start scanning. After turning ON the laser light source 101, the irradiation position of the excitation light is scanned to point 302 (Sx+m-1, Sy) on the right side of the first line. During this time, the y coordinate (Sy) of the irradiation position of the excitation light is maintained. Thereafter, at time t4, the laser light source 101 is turned OFF, and the irradiation position of the excitation light is turned back and moved to start point 303 of the second line, and the laser light source 101 is turned ON again to scan the second line. Thereafter, scanning continues until the nth line is completed.

[0040] Then, at time t6, when the irradiation position of the excitation light reaches point 306 (Sx+m-1, Sy+n-1), which is the scanning end position, the laser light source 101 is turned off and moved to standby position 107(a, b). After the movement is completed, at time t7, the laser is turned on and placed on standby to wait for the next scanning command.

[0041] 5 is a flowchart showing the operation of the scan head 100 in the first embodiment. In step S01, after starting the system, the control unit moves the irradiation position of the excitation light to a standby position 107 by scanning the galvanometer mirror 14. After moving to the standby position 107, in the next step S02, the laser light source 101 is turned on, and in the next step S03, the microscope 200 waits for several seconds only once immediately after starting up until the output value of the laser light source 101 stabilizes.

[0042] In the next step S04, when the user gives an instruction to start scanning, in the next step S05, the laser light source 101 is turned OFF once, and in the next step S06, the irradiation position of the excitation light is moved to the scan start position (first, the scan start position 301 (Sx, Sy) for the first line). In the next step S07 after the movement, the laser light source 101 is turned ON, and in the next step S08, one line is scanned. Thereafter, in the next step S09, the laser light source 101 is turned OFF, the irradiation position of the excitation light is turned back, and the next line is scanned, and sequential scanning continues up to the nth line, which is the final line.

[0043] In the next step S10, if the irradiation position of the excitation light has reached point 306 (Sx+m-1, Sy+n-1), which is the scanning end position (YES in step S10), in the next step S11, the laser light source 101 is turned OFF and the irradiation position of the excitation light is moved to standby position 107. After the movement, in the next step S12, the laser light source 101 is turned ON and waits until the output value stabilizes. In the next step S13, if there is no more scanning range and scanning is completed, the experiment ends (YES in step S13). If there is still a scanning range remaining (NO in step S13), the process returns to step S04 and the above processing is performed again.

[0044] According to the microscope 200 of the first embodiment, a standby position 107 where the excitation light waits is provided, the laser light source 101 is turned on at the standby position 107 and waits until the output value stabilizes, and in the state where the output value has stabilized, irradiation of the sample 106 is started. This solves the problem of unexpected changes in the brightness of the acquired image, making it impossible to perform quantitative analysis, and makes it possible to stabilize the brightness of the acquired image.

[0045] According to the microscope 200 of the first embodiment, the laser light source 101 is turned off while moving from the standby position 107 to the scan start position 301, while turning back from point 302 to point 303, and while moving from the scan end position 306 to the standby position 107. This makes it possible to reduce phototoxicity and damage to the sample 106 caused by irradiation with excitation light.

[0046] FIG. 6 shows a schematic configuration of a scan head 110 according to the second embodiment. In FIG. 6, the same components as those in the scan head 100 according to the first embodiment shown in FIG. 2 are denoted by the same reference numerals, and their description will be omitted. As shown in FIG. 6, the scan head 110 according to the second embodiment has a light-shielding means 21 at a standby position 107 where the excitation light is kept on standby. By providing the light-shielding means 21 at the standby position 107, the generation of secondary light due to reflection of the excitation light at the standby position, which may occur if the light-shielding means 21 is not provided, is suppressed. This makes it possible to prevent stray light due to the secondary light from being irradiated onto the sample 106 via the microscope port 17.

[0047] Fig. 7 shows a schematic configuration of a scan head 120 according to the third embodiment. In Fig. 7, the same components as those in the scan head 100 according to the first embodiment shown in Fig. 2 are denoted by the same reference numerals, and their description will be omitted. As shown in Fig. 7, in the scan head 120 according to the third embodiment, in order to deal with stray light, instead of a standby position 107 where the excitation light is kept on standby, a reflecting member 22 is provided midway along the path of the excitation light toward the standby position 107, and a beam dump 23 that absorbs the excitation light is provided at the destination of the excitation light reflected by the reflecting member 22.

[0048] The reflecting member 22 has a function of redirecting the excitation light heading toward the standby position 107 away from the excitation light heading toward the incident surface 15a, which is the main beam. The beam dump 23 has a function of further absorbing, by mechanical or optical means, at least a portion of the excitation light redirected away from the main beam by the reflecting member 22. By providing the reflecting member 22 on the path to the standby position 107 and combining it with the beam dump 23, the generation of secondary light is suppressed, and it is possible to prevent stray light from being irradiated onto the sample 106 via the microscope port 17.

[0049] Fig. 8 shows another example of ON / OFF control of the laser light source 101. Fig. 8(a) illustrates a portion of the timing chart shown in Fig. 4. In Fig. 4, while the excitation light is on standby at the standby position 107, the laser light source 101 is controlled to be OFF at time t0, ON from time t1 to time t2, turned OFF once at time t2, and turned ON again at time t3. As shown in Fig. 8(a), overall, the laser light source 101 is controlled to be turned ON and OFF at a time ratio of approximately 3:1 to 4:1.

[0050] 4, the laser light source 101 is controlled to be always ON while the excitation light is on standby at the standby position 107. However, the laser light source 101 may be controlled to be ON / OFF while the excitation light is on standby at the standby position 107, as shown in FIG. 8(b). FIG. 8(b) shows another control example, and illustrates an enlarged view of the details of the period from time t1 to time t2 while the excitation light is on standby at the standby position 107.

[0051] As shown in FIG. 8(b), in another control example, during the second state in which the excitation light is waiting at the standby position 107, the laser light source 101 is not always ON, but is instead alternately turned ON and OFF at the same timing as in FIG. 8(a). As shown in FIG. 8(b), the ON / OFF time ratio of the laser light source 101 is approximately 3:1 to 4:1, similar to FIG. 8(a). In other words, the control in FIG. 8(b) is similar to the control in FIG. 8(a). By controlling the ON / OFF of the laser light source 101 as shown in FIG. 8(b) while the laser light source 101 is waiting at the standby position 107, the output timing is similar to the ON / OFF control during actual scanning, and the output state is similar, thereby further stabilizing the output value of the laser light source 101.

[0052] In the above embodiment, the galvanometer mirror 14 is used as the optical path changing member. However, instead of the galvanometer mirror 14, a DMD (Digital Mirror Device), a MEMS (Micro Electro Mechanical Systems) shutter, a spatial light modulator (SLM), or the like may be used as the optical path changing member.

[0053] In the above embodiment, the laser light source 101 is turned off while the excitation light is moving from the standby position 107 to the scan start position 301, while the excitation light is turning back from point 302 to point 303, and while the excitation light is moving from the scan end position 306 to the standby position 107, thereby reducing phototoxicity and damage to the sample 106. However, instead of this, the laser light source 101 may be temporarily shut off using a mechanical shutter using a motor or the like. In this case, scanning may be performed taking into account the opening and closing speed of the mechanical shutter.

[0054] In the above embodiment, the first and second states are switched by switching between a first optical path in which the excitation light passes through the scanning optical system 15, which is disposed downstream of the optical path of the galvanometer mirror 14, and a second optical path in which the excitation light does not pass through the scanning optical system 15. However, a mirror may be disposed between the scanning optical system 15 and the objective optical system 105, and the mirror may be controlled to switch between an optical path in which the excitation light passes through the objective optical system 105 and an optical path in which the excitation light does not pass through the objective optical system 105. Specifically, the standby position 107 may be located outside the objective optical system 105 (on the side surface of the lens barrel of the second objective lens 105b, similar to the side surface 16a of the lens barrel 16), i.e., at a position where the excitation light does not enter the objective optical system 105. Instead of the laser light source 101, a superluminescent diode (SLD) that can be directly controlled by the control unit 104 to be switched on and off may be used. In this case, a filter that passes only a predetermined wavelength of light emitted from the SLD may be disposed to generate excitation light of a desired wavelength. The laser light source 101 and the SLD are light sources that emit coherent light.

[0055] Various embodiments of the present invention may also be described with reference to flowcharts and block diagrams, where the blocks may represent (1) stages of a process in which operations are performed or (2) sections of an apparatus responsible for performing the operations. Particular stages and sections may be implemented by dedicated circuitry, programmable circuitry provided with computer-readable instructions stored on a computer-readable medium, and / or a processor provided with computer-readable instructions stored on a computer-readable medium. Dedicated circuitry may include digital and / or analog hardware circuitry, and may include integrated circuits (ICs) and / or discrete circuits. Programmable circuitry may include reconfigurable hardware circuitry, including logical AND, OR, XOR, NAND, NOR, and other logic operations, flip-flops, registers, memory elements such as field programmable gate arrays (FPGAs), programmable logic arrays (PLAs), and the like.

[0056] A computer-readable medium may include any tangible device capable of storing instructions that are executed by an appropriate device, such that the computer-readable medium having instructions stored thereon comprises an article of manufacture containing instructions that can be executed to create means for performing the operations specified in the flowcharts or block diagrams. Examples of computer-readable media may include electronic, magnetic, optical, electromagnetic, and semiconductor storage media. More specific examples of computer-readable media may include floppy disks, diskettes, hard disks, random access memory (RAM), read-only memory (ROM), erasable programmable read-only memory (EPROM or flash memory), electrically erasable programmable read-only memory (EEPROM), static random access memory (SRAM), compact disc read-only memory (CD-ROM), digital versatile disc (DVD), Blu-ray (RTM) disc, memory stick, integrated circuit card, and the like.

[0057] The computer readable instructions may include either assembler instructions, Instruction Set Architecture (ISA) instructions, machine instructions, machine-dependent instructions, microcode, firmware instructions, state-setting data, or source or object code written in any combination of one or more programming languages, including object-oriented programming languages ​​such as Smalltalk®, JAVA®, C++, etc., and conventional procedural programming languages ​​such as the “C” programming language or similar programming languages.

[0058] The computer-readable instructions may be provided to a processor or programmable circuitry of a general-purpose computer, special-purpose computer, or other programmable data processing apparatus, either locally or over a wide-area network (WAN) such as a local area network (LAN), the Internet, etc., which executes the computer-readable instructions to create means for performing the operations specified in the flowcharts or block diagrams. Examples of processors include computer processors, processing units, microprocessors, digital signal processors, controllers, microcontrollers, etc.

[0059] 9 illustrates an example of a computer 2200 in which aspects of the present invention may be embodied, in whole or in part. Programs installed on the computer 2200 may cause the computer 2200 to function as or perform operations associated with an apparatus or one or more sections of the apparatus according to embodiments of the present invention, and / or to perform a process or steps of a process according to embodiments of the present invention. Such programs may be executed by the CPU 2212 to cause the computer 2200 to perform specific operations associated with some or all of the blocks of the flowcharts and block diagrams described herein.

[0060] A computer 2200 according to this embodiment includes a CPU 2212, a RAM 2214, a graphics controller 2216, and a display device 2218, which are interconnected by a host controller 2210. The computer 2200 also includes input / output units such as a communication interface 2222, a hard disk drive 2224, a DVD-ROM drive 2226, and an IC card drive, which are connected to the host controller 2210 via an input / output controller 2220. The computer also includes legacy input / output units such as a ROM 2230 and a keyboard 2242, which are connected to the input / output controller 2220 via an input / output chip 2240.

[0061] The CPU 2212 operates according to programs stored in the ROM 2230 and RAM 2214, thereby controlling each unit. The graphics controller 2216 acquires image data generated by the CPU 2212 into a frame buffer or the like provided in the RAM 2214 or into the graphics controller 2216 itself, and causes the image data to be displayed on the display device 2218.

[0062] The communication interface 2222 communicates with other electronic devices via a network. The hard disk drive 2224 stores programs and data used by the CPU 2212 in the computer 2200. The DVD-ROM drive 2226 reads programs or data from the DVD-ROM 2201 and provides the programs or data to the hard disk drive 2224 via the RAM 2214. The IC card drive reads programs and data from an IC card and / or writes programs and data to an IC card.

[0063] The ROM 2230 stores therein a boot program or the like that is executed by the computer 2200 upon activation, and / or programs that depend on the hardware of the computer 2200. The input / output chip 2240 may also connect various input / output units to the input / output controller 2220 via a parallel port, a serial port, a keyboard port, a mouse port, etc.

[0064] The programs are provided by a computer-readable medium such as a DVD-ROM 2201 or an IC card. The programs are read from the computer-readable medium, installed in the hard disk drive 2224, RAM 2214, or ROM 2230, which are also examples of computer-readable media, and executed by the CPU 2212. Information processing described in these programs is read by the computer 2200, and brings about cooperation between the programs and the various types of hardware resources described above. An apparatus or method may be configured by realizing information manipulation or processing in accordance with the use of the computer 2200.

[0065] For example, when communication is performed between the computer 2200 and an external device, the CPU 2212 may execute a communication program loaded into the RAM 2214 and instruct the communication interface 2222 to perform communication processing based on the processing described in the communication program. Under the control of the CPU 2212, the communication interface 2222 reads transmission data stored in a transmission buffer processing area provided in the RAM 2214, the hard disk drive 2224, the DVD-ROM 2201, or a recording medium such as an IC card, and transmits the read transmission data to the network, or writes reception data received from the network to a reception buffer processing area or the like provided on the recording medium.

[0066] The CPU 2212 may also cause all or a necessary portion of a file or database stored on an external recording medium such as the hard disk drive 2224, the DVD-ROM drive 2226 (DVD-ROM 2201), an IC card, etc. to be read into the RAM 2214, and perform various types of processing on the data on the RAM 2214. The CPU 2212 then writes back the processed data to the external recording medium.

[0067] Various types of information, such as various types of programs, data, tables, and databases, may be stored on the recording medium and may undergo information processing. The CPU 2212 may perform various types of processing on data read from the RAM 2214, including various types of operations, information processing, conditional judgment, conditional branching, unconditional branching, information search / replacement, etc., as described throughout this disclosure and specified by the instruction sequences of the programs, and write the results back to the RAM 2214. The CPU 2212 may also search for information in a file, database, etc. on the recording medium. For example, if multiple entries each having an attribute value of a first attribute associated with an attribute value of a second attribute are stored on the recording medium, the CPU 2212 may search for an entry that matches a condition specified by the attribute value of the first attribute from among the multiple entries, read the attribute value of the second attribute stored in the entry, and thereby obtain the attribute value of the second attribute associated with the first attribute that satisfies a predetermined condition.

[0068] The above-described programs or software modules may be stored in a computer-readable medium on or near the computer 2200. A recording medium such as a hard disk or RAM provided in a server system connected to a dedicated communication network or the Internet can also be used as a computer-readable medium, thereby providing the programs to the computer 2200 via the network.

[0069] Although the present invention has been described above using embodiments, the technical scope of the present invention is not limited to the scope described in the above embodiments. It will be apparent to those skilled in the art that various modifications and improvements can be made to the above embodiments. It is clear from the claims that such modifications and improvements can also be included within the technical scope of the present invention.

[0070] It should be noted that the order of execution of each process, such as operations, procedures, steps, and stages, in the devices, systems, programs, and methods shown in the claims, specifications, and drawings is not specifically stated as "before," "prior to," etc., and that the processes can be performed in any order unless the output of a previous process is used in a subsequent process. Even if the operational flow in the claims, specifications, and drawings is described using "first," "next," etc. for convenience, this does not mean that the processes must be performed in this order. [Explanation of symbols]

[0071] 11 input port, 12 collimating lens, 13 dichroic mirror, 14 galvanometer mirror, 15 scanning optical system, 15a incident surface, 16a side surface, 16 lens barrel, 17 microscope port, 18 condenser lens, 19 pinhole, 20 output port, 22 reflecting member, 23 beam dump, 100 scan head, 101 laser light source, 102 light detection device, 103 microscope body, 104 control unit, 105 objective optical system, 106 sample, 107 standby position, 108 observable range, 200 microscope, 2200 computer, 2201 DVD-ROM, 2210 host controller, 2212 CPU, 2214 RAM, 2216 graphics controller, 2218 display device, 2220 input / output controller, 2222 communication interface, 2224 hard disk drive, 2226 DVD-ROM drive, 2230 ROM, 2240 input / output chip, 2242 keyboard

Claims

1. a light source that can be directly controlled between an ON state in which coherent light is emitted and an OFF state in which the light is not emitted; an illumination optical system that irradiates the sample with the light to form an illumination area on the sample; an optical path changing member that changes the optical path of the light; a control unit that controls the light source and the optical path changing member; and the control unit controls the optical path changing member to switch between a first state in which the light forms the illumination area on the sample when the light source is in the ON state and does not form the illumination area when the light source is in the OFF state, and a second state in which the light does not form the illumination area on the sample regardless of whether the light source is in the ON state or the OFF state, In the first state, after controlling the light source to the ON state and forming the illumination area on the sample with the light and completing image acquisition, while switching from the first state to the second state, the light source is controlled to the OFF state, and until a command to start image acquisition is received, in the second state, the light source is controlled to always be in the ON state or is controlled so that the ON state and the OFF state are repeated at predetermined timings; After receiving a command to start image acquisition, the light source is controlled to the OFF state while switching from the second state to the first state, and in the first state, the light source is controlled to the ON state to form the illumination area on the sample with the light and start image acquisition. microscope.

2. the illumination optical system further includes a condenser lens; 2. The microscope according to claim 1, wherein the first state is such that the light passes through the condenser lens, and the second state is such that the light does not pass through the condenser lens and enters a predetermined standby position.

3. a holding member for holding the condenser lens; The microscope according to claim 2 , wherein the standby position is a part of the holding member.

4. The microscope according to claim 3 , wherein a light-shielding member is provided in the part.

5. a reflecting member provided at the standby position and reflecting the light; an absorbing member that absorbs at least a portion of the light reflected from the reflecting member; The microscope of claim 2 further comprising:

6. 2. The microscope according to claim 1, wherein the optical path changing member is a galvanometer mirror or a spatial light modulator.

7. 7. The microscope according to claim 1, wherein the control unit, in the second state, turns the light source to the ON state, waits for a predetermined time, and then switches the light to the first state.

8. 8. The microscope according to claim 7, wherein the predetermined time is a time required for the output value from the light source to stabilize.

9. a light source that can be directly controlled between an ON state in which coherent light is emitted and an OFF state in which the light is not emitted; an illumination optical system that irradiates the sample with the light to form an illumination area on the sample; an optical path changing member that changes the optical path of the light; a control unit that controls the light source and the optical path changing member, a first state in which the light path changing member is controlled so that the light forms the illumination area on the sample when the light source is in the ON state, and does not form the illumination area when the light source is in the OFF state; a first state in which the light source is in the ON state or the OFF state, in which the light does not form the illumination area on the sample, and a second state in which the light source is in the ON state or the OFF state, In the first state, after controlling the light source to the ON state and forming the illumination area on the sample with the light and completing image acquisition, while switching from the first state to the second state, the light source is controlled to the OFF state, and until a command to start image acquisition is received, in the second state, the light source is controlled to always be in the ON state or is controlled so that the ON state and the OFF state are repeated at predetermined timings; After receiving a command to start image acquisition, the light source is controlled to be in the OFF state while switching from the second state to the first state, and in the first state, the light source is controlled to be in the ON state to form the illumination area on the sample with the light and start image acquisition. How to control the microscope.

Images