JP5466976B2 - Microscope system, observation image display method, program - Google Patents

Description

  The present invention relates to a microscope system having a microscope for performing observation or the like on a biological sample that generates weak light and a control device for controlling the microscope, an image acquisition means, and a method for displaying an observation image having a control means for controlling the image acquisition means. And related to the program.

  There has been disclosed a light emission microscope capable of taking a light emission image or a fluorescence image of a biological sample that generates self-luminescence such as bioluminescence which is weak light. This luminescence microscope has a dark box for obtaining a light-shielding state for imaging weak light, holds a biological sample imparted with a light emitting property in the dark box, and in the dark field of the biological sample. A luminescent image can be acquired. Since the light emitted from the biological sample is extremely weak, an exposure time of about 30 minutes is required to capture the luminescent image in order to acquire a clear luminescent image (see, for example, Patent Document 1).

JP 2007-108154 A

  By the way, in recent years, researches that require long-term observations such as the differentiation process of ES cells and iPS cells have been actively conducted, but various reporters called differentiation markers and undifferentiation markers have been studied in more detail. It is necessary to pick up and select colonies of cells expressing the gene. As a method for detecting the expression of a differentiation marker or an undifferentiation marker, various methods can be used depending on the type of the marker.

  For example, when a fluorescent protein such as GFP is used as a reporter gene, it is necessary to irradiate an ES cell or iPS cell with excitation light to obtain a fluorescence image. However, since the excitation light requires light with a high energy level to sufficiently excite the colonized cell group, the pick-up operation should be performed while irradiating the ES cell or iPS cell with the excitation light for a long time. Is not preferred. Therefore, the target colony is picked up based on the fluorescence image acquired in the past.

  On the other hand, when a self-luminous protein is used as a reporter gene, the luminescence derived from gene expression generated from individual cells is extremely weak, so detection capable of imaging faint light like the above-mentioned luminescence microscope In addition, it is necessary to use a total of two types of microscopes such as a stereo microscope or an upright / inverted microscope that can be observed in a bright field. This is because it is difficult to perform the pickup work because the dark box interferes with the work under the light emission microscope. Therefore, when a self-luminous protein is used as a reporter gene, a luminescence image is acquired with a luminescence microscope, and the position of a colony expressing a differentiation marker or an undifferentiation marker in a petri dish is confirmed. Based on the image, a target colony is picked up under a stereo microscope or an upright / inverted microscope. However, in such a conventional method, it cannot be said that a colony expressing a differentiation marker or an undifferentiation marker accurately corresponds to a colony actually picked up, and there is room for improvement. It was.

  The objective of this invention is providing the microscope system etc. which can visually recognize the weak light image of a biological sample in a live image.

  In order to achieve the object, a microscope system according to one aspect of the present invention is connected to a first microscope capable of acquiring a weak light image of a biological sample, the first microscope, and the weak light image. Are connected to the second microscope, the second microscope capable of acquiring a bright-field live image of the biological sample at a magnification different from that of the weak light image, and the weak And a second control device having a display unit capable of displaying an optical image superimposed on a part of the bright-field live image so as to have a magnification equivalent to that of the bright-field live image.

  In order to achieve the object, an observation image display method according to one aspect of the present invention acquires a weak light image of a biological sample in advance by a first image acquisition unit, and the biological image of the biological sample by a second image acquisition unit. A bright-field live image is acquired at a different magnification from the weak light image, and the weak light image is made to have the same magnification as the live image by the control means, and is superimposed on a part of the live image and displayed. Display on the means.

  In order to achieve the object, a program according to one aspect of the present invention includes a first image acquisition unit that acquires a weak light image of a biological sample, and a second image that acquires a live-field live image of the biological sample. A function for acquiring a weak light image of the biological sample by the first image acquisition means in an observation image display system comprising an acquisition means, a control means for controlling them, and a display means provided in the control means; A function of acquiring a live image of a bright field of a biological sample at a magnification different from that of the weak light image by the second image acquisition means, and a magnification equivalent to that of the live image by the control means. And a function of displaying the image on the display unit so as to overlap a part of the live image.

  In order to achieve the above object, a program according to another aspect of the present invention includes a first image acquisition unit that acquires a weak light image of a biological sample, and a second image that acquires a bright-field live image of the biological sample. A weak light image of the biological sample is obtained by the first image obtaining means in an observation image display system comprising an obtaining means, a control means for controlling these, a display means provided in the control means, and a display switching means. A function for acquiring at a high magnification, a function for acquiring a live image of a bright field of a biological sample at a low magnification by the second image acquisition means, and a magnification equivalent to that of the live image by the control means. And the weak light image displayed on the live image by the display switching unit, and the function of displaying the image on the display unit so as to be superimposed on a part of the live image. Display, a function of switching the non-display, to realize.

  In order to achieve the above object, a program according to another aspect of the present invention includes a first image acquisition unit that acquires a weak light image of a biological sample, and a second image that acquires a bright-field live image of the biological sample. A weak light image of the biological sample is obtained by the first image acquisition means on an observation image display system comprising an acquisition means, a control means for controlling these, a display means and a non-display means provided in the control means. A function for acquiring at a high magnification, a function for acquiring a live image of a bright field of a biological sample at a low magnification by the second image acquisition means, and a magnification equivalent to that of the live image by the control means. A function of displaying the image on the display unit so as to be overlapped with a part of the live image and a pixel value of a part of the live image within a certain time , To realize a function to zero the pixel values of a partial region of the weak light image corresponding to a partial area of the live image by the non-display means.

  According to said structure, the microscope system etc. which can visually recognize the weak light image of a biological sample in a live image can be provided.

The perspective view which showed the microscope system concerning 1st Embodiment. The block diagram which showed typically the 1st microscope of the microscope system shown in FIG. The block diagram which showed typically the 2nd microscope of the microscope system shown in FIG. The flowchart which showed the flow of the pick-up operation | work using the microscope system of 1st Embodiment. The figure which showed the state which accumulated the live image and light emission image which were obtained with the microscope system shown in FIG. 1 on the screen of the 2nd display part. The block diagram which showed typically the 2nd microscope of the microscope system concerning 2nd Embodiment. On the screen of the second display unit, an enlarged image obtained by superimposing a live image and a light emission image on a place adjacent to a display part of a live image obtained by the microscope system according to the second embodiment is displayed. The figure which showed the state displayed on. The state which displayed the enlarged image which expanded the designation | designated location in a live image on the screen of the 2nd display part in the location adjacent to the display location of the live image obtained with the microscope system concerning 2nd Embodiment is shown. Figure. The figure which showed the state before moving the live image obtained with the microscope system concerning 3rd Embodiment. The figure which showed the state after moving the live image obtained with the microscope system concerning 3rd Embodiment. The top view which showed the biological sample used for the microscope system concerning 4th Embodiment, and the thing to which the marker used as a reference position was attached | subjected to the bottom part of the petri dish.

  Hereinafter, a first embodiment of the microscope system will be described with reference to FIGS. 1 to 5. This microscope system 11 is weak over time while culturing a biological sample such as an ES cell or iPS cell into which a fluorescent protein or fluorescent dye such as a luciferase luminescent vector or a fluorescent dye is introduced as a differentiation marker in a petri dish or the like. An optical image is acquired, and after several days, it is used for operations such as picking up a colony of cultured cells in a biological sample based on this luminescent image and live image.

  As shown in FIG. 1, the microscope system 11 mainly controls a first microscope 13 (first image acquisition unit) that acquires a weak light image 21 of a biological sample 12, a first microscope 13, and a first microscope 13. A first control device 14 (control means) that stores the weak light image 21 acquired by one microscope 13 and a second microscope 15 (second image acquisition means) that mainly acquires a live image 22 of the biological sample 12. And a second control device 16 (control means) for controlling the second microscope 15 and displaying the weak light image 21 and the live image 22 acquired by the second microscope 15 in a superimposed manner.

  As shown in FIGS. 1 and 2, the first microscope 13 is a so-called luminescence microscope, and acquires a luminescence image 21 based on luminescence that is weak light emitted mainly from a biological sample such as a cell. For example, an imaging device (CCD, CMOS, etc.) that exposes light for a sufficient time (1 minute or longer) and a high NA imaging optical system (objective lens, imaging lens, etc.) are provided. Note that the faint light image referred to in the present invention is a concept including a luminescent image obtained by self-luminescence such as protein and a fluorescent image obtained by irradiating excitation light.

  The first microscope 13 can also acquire a bright field image of the biological sample 12 using the light source 23. Here, the bright field image acquired together with the luminescent image 21 is acquired mainly before and after the imaging period in which the luminescent image 21 is acquired, and provides an observation image of the whole biological sample corresponding to the luminescent image 21. In order to evaluate the activity of the biological sample in chronological order, it is preferable to acquire a plurality of luminescent images 21 at desired times, and it is more preferable to acquire a bright field image each time.

  The first microscope 13 includes a main body 24, a dark box 25 provided in a part of the main body 24, a lid 26 covering the upper portion of the dark box 25, an incubator 27 and a first stage 28 provided in the dark box 25 ,have. In the first microscope 13, an observation hole (not shown) is provided in the incubator 27 and the first stage 28.

  The first microscope 13 includes an objective optical system 32 provided below the dark box 25 and further below the first stage 28, and a first filter 33 that blocks unnecessary light passing through the objective optical system 32. And a first CCD camera 34 provided below the objective optical system 32 and the first filter 33 and capable of detecting light passing through the objective optical system 32. If only monochromatic emission observation is performed, the first filter 33 can be omitted. The objective optical system may be provided with a zoom mechanism (not shown).

  The first microscope 13 includes a light source 23 used when acquiring a bright field image or a fluorescence image, and a second filter 35 for adjusting light from the light source 23. The light source 23 includes a lamp, an LED, and the like that can emit white light used when acquiring a bright field image. The first microscope 13 may further include a fluorescence unit that can irradiate excitation light used during fluorescence observation. Examples of a microscope that acquires a luminescent image, a bright-field image, and a fluorescent image include, but are not limited to, a luminescent imaging system LUMINOWEW LV200 (LUMINOWVIEW: registered trademark) manufactured by Olympus Corporation.

  As shown in FIG. 1, the first control device 14 is a computer such as a general personal computer (PC). The first control device 14 includes a first display unit 36 constituted by a liquid crystal display or the like, and a keyboard 37 and a mouse 38 as input means. The first control device 14 can control the photographing conditions of the first CCD camera 34, image the acquired image, display it on the first display unit 36, and the light amount of the light source 23. The first control device 14 stores the light emission image 21, the fluorescence image, and the bright field image acquired by the first microscope 13, and performs image processing and image analysis of the light emission image 21, the fluorescence image, and the bright field image. Is also possible. The first control device 14 can also store the result data of the image analysis together with the observation conditions. The first control device 14 can also control switching of the first filter 33 and the second filter 35. Further, the first control device 14 can perform zoom control of the objective optical system 32.

  When light emission observation is performed with the first microscope 13 and the first control device 14, the light emitted from the biological sample 12 proceeds to the objective optical system 32, and unnecessary light is blocked by the first filter 33. 1 is detected by one CCD camera 34. The light detected by the first CCD camera 34 is sent to the first control device 14 and imaged.

  When acquiring a bright field image, the second filter 35 illuminates the sample 12 so that the white illumination light is irradiated, and the transmitted light travels through the objective optical system 32, and the first filter 33 The light travels as it is without being blocked, and is detected by the CCD camera 34. The light detected by the CCD camera 34 is sent to the first control device 14 and imaged.

  When acquiring a fluorescent image, the light emitted from the light source 23 is transmitted through only the light having a wavelength that excites the fluorescent protein or the like by the second filter 35 and is applied to the specimen 12. The light emitted from the specimen 12 travels through the objective optical system 32, unnecessary light is blocked by the first filter 33, and is detected by the CCD camera 34. The light detected by the CCD camera 34 is sent to the first control device 14 and imaged.

  The second microscope 15 is a so-called upright microscope used for obtaining a live image 22 of the biological sample 12 in a bright field and picking up a colony 41 of cultured cells in the biological sample 12. The second microscope 15 is installed in, for example, a clean bench where bacteria or the like do not enter the biological sample. As shown in FIG. 3, the second microscope 15 includes a second main body 40, an eyepiece unit 42 provided on the second main body 40, and a second CCD camera provided on the upper part of the eyepiece unit 42. 43, a plurality of objective lenses 45 that are rotatably fixed to the second main body 40 via a revolver 44, and a cube unit 46 provided between the eyepiece unit 42 and the objective lens 45. doing.

  The eyepiece unit 42 includes an imaging lens (not shown) and an eyepiece lens. The eyepiece unit 42 incorporates a filter and a mirror for branching the optical path, and branches the observation optical path with the eyepiece and the observation optical path with the second CCD camera 43.

  Objective lenses 45 having different magnifications are detachably attached to the revolver 44. The cube unit 46 holds a plurality of mirrors, half mirrors, filters, and the like (not shown), and branches an illumination optical path illuminated by the illumination unit 47 and an observation optical path from the eyepiece unit 42 and the second CCD camera 43. ing.

  The second microscope 15 further includes a second stage 31 provided below the second body 40, a specimen (biological sample 12) placed on the second stage 31, and the second body 40. And a lighting unit 47 for projecting the specimen (biological sample 12). The control unit 51 includes a drive control circuit that performs drive control of each part of the second microscope 15. The second stage 31 can be manually moved, for example, not only in the X-axis direction and the Y-axis direction (horizontal direction) but also in the Z-axis direction (vertical direction). The revolver 44 of this embodiment is configured by an electric one. The illumination unit 47 includes a plurality of light sources, a collector lens, a relay lens, an aperture stop unit, a field stop unit, and the like (not shown).

  Light from the illumination unit 47 is projected onto the biological sample 12 through the objective lens 45 via the cube unit 46.

  When the biological sample 12 is observed by the eyepiece unit 42, light such as fluorescence or reflected light emitted from the biological sample 12 is transmitted to the eyepiece unit 42 through the objective lens 45 and the cube unit 46, and is provided in the eyepiece unit 42. Focused on the imaging lens. This light is imaged on a predetermined imaging surface and can be visually observed through an eyepiece.

  In the observation by the second CCD camera 43, the light branched in the direction of the second CCD camera 43 by a mirror in the eyepiece unit 42 is imaged on the CCD surface of the second CCD camera 43, and the biological sample 12 is observed. The observation image can be displayed on the second display unit 48 of the second control device 16 connected to the second CCD camera 43.

  As shown in FIG. 1, the second control device 16 is configured by a computer such as a general personal computer (PC). The second control device 16 has a second display section 48 (display means) constituted by a liquid crystal display or the like, a keyboard 49 and a mouse 50 as input means, and the like. The second control device 16 can control the shooting conditions of the second CCD camera 43, convert the acquired image into an image and display it on the second display unit 48, and control the light quantity of the illumination unit 47. In addition to storing the live image 22 acquired by the second microscope 15, the second control device 16 can also perform image processing and image analysis of the luminescent image 21 and the live image 22. The second control device 16 can also store the result data of the image analysis together with the observation conditions. As shown in FIG. 3, the second control device 16 can control a plurality of cubes and control the rotation of the revolver 44.

  Luminescence observation using the first microscope 13, the first control device 14, the second microscope 15, and the second control device 16 configured as described above and picking up the colonies 41 of cultured cells in the biological sample 12. The operation will be described with reference to FIG.

  First, the procedure for observing light emission for several days is as follows.

  After the biological sample 12 is placed on the first microscope 13, observation conditions are set according to the type of the biological sample 12 and the culture conditions. Since the amount of light emitted from the biological sample 12 is extremely small, a clear image is acquired (captured) with a sufficient exposure time. The first CCD camera 34 is operated from the first control device 14 to acquire an image, and the image is stored in a storage device built in the first control device 14. The emission image 21 is acquired, for example, several times to several tens of times a day (S11). The luminescent image 21 uses an objective lens (for example, a 20 × objective lens) for obtaining a total magnification of a sufficiently high magnification (for example, 40 to 150 times) that can individually identify cells that generate weak self-luminescence. And obtain at high magnification. Here, the optical conditions such as the magnification of the objective lens are determined according to other imaging optical systems (imaging lens, relay lens, etc.) of the microscope 13. At this time, when acquiring the luminescent image 21 and the fluorescence image, a bright field image of the biological sample 12 may be taken together and the bright field image may be stored together.

  Subsequently, after observing for several days, when the colony 41 is formed in the cultured cells of the biological sample 12, the procedure for picking up the colony 41 is as follows.

  The petri dish in which the cells (biological sample 12) have been cultured is moved from the first microscope 13 to the second microscope 15. At this time, various images acquired by the first microscope 13 in the past are copied to the second control device 16 connected to the second microscope 15 (S12).

  Then, bright field observation is performed visually from the eyepiece unit 42 (S13). Subsequently, using an objective lens (for example, 10-times objective lens 45) for obtaining a total magnification that is low enough (for example, 0.5 to 30 times) sufficient to make the entire individual cell an observation field, The second stage 31 is moved to focus on the biological sample 12 to obtain a low magnification image.

  In this state, switching to observation with the second CCD camera 43 is started, real-time display (preview display, live display) of the bright field image is started, and acquisition of the low-magnification live image 22 is started.

  Subsequently, when setting is made to display a superimposed display (S14) and the user instructs display of the past light emitting image 21 including the same colony 41 as the colony 41 included in the live image 22, the second control is performed. The device 16 reads the light emission image 21 (S15). In the second control device 16, the luminescent image 21 is displayed so as to overlap a part of the live image 22 (S 16), and the luminescent image 21 is displayed in the second so that the display position can be manually changed via the mouse 50 or the like. Is displayed on the display unit 48. At this time, for example, the bright-field live image 22 is acquired using, for example, a 10 × objective lens 45, and the luminescent image 21 previously captured before that is captured using a 20 × objective lens. For this reason, the luminescent image 21 is reduced and displayed in accordance with the magnification of the displayed live image 22. Note that when the magnification of the bright-field live image 22 is changed, the reduction rate of the light-emitting image 21 is also changed accordingly, and the live image 22 is displayed superimposed on the live image 22.

  In the first embodiment, the user manually moves the luminescent image 21 using the mouse 50 or the like so that the colony image in the luminescent image 21 overlaps the corresponding colony image in the live image 22 and fine-tunes it. Then, the luminescent image 21 is arranged (S17).

  In this way, as shown in FIG. 5, the light emission image 21 is superimposed on the bright field live image 22 and displayed. At this time, when there are a plurality of light-emitting images 21 corresponding to the bright-field live image 22 corresponding to different areas on the live image (for example, a plurality of selected partial images corresponding to each of a plurality of cells or colonies). As shown in FIG. 5, a plurality of light-emitting images 21 may be coincident with the corresponding regions on the live image and overlapped and displayed at the same time. In addition, the time-series emission image 21 may be displayed as a moving image while the live image is continuously displayed. By displaying a time-series luminescence image at the same time, the activity such as gene expression may be confirmed for each individual cell, intracellular site, and colony.

  In this way, the differentiated colony 41 becomes very easy to understand, and the user can clearly recognize which colony 41 should be picked up. Thus, the desired colony 41 can be efficiently picked up with a pipette tip (S18).

  When the necessary pick-up work is completed (S19), the overlay display is canceled (S20), and the real-time display is terminated (S21).

  The various controls and display methods described above can be realized by software (computer program) installed in the first control device 14 and the second control device 16. The processing program is recorded in a recording medium such as a CD-ROM or a storage device such as a hard disk built in the first control device 14 and the second control device 16. The first control device 14 and the second control device 16 read the processing program onto the memory as necessary, and are executed by the CPU, and each part of the first control device 14 and the second control device 16, and these The first microscope 13 and the second microscope 15 connected to are controlled.

  The recording device may be a computer such as a program server connected to the first control device 14 and the second control device 16 via a communication line. In this case, the transmission signal obtained by modulating the carrier wave with the data signal representing the control program is transmitted from the program server to the first control device 14 and the second control device 16 through the communication line, and the first The controller 14 and the second controller 16 demodulate the transmission signal received to reproduce the processing program, and the CPU executes the processing program.

  According to the first embodiment, the user can manually and efficiently superimpose the luminescent image 21 on the live image 22, and can efficiently recognize the differentiated colony 41 and facilitate the pick-up operation. it can.

  Next, a second embodiment of the microscope system will be described with reference to FIG. In the microscope system 11 of the second embodiment, the second control device has an image recognition unit 52, a display switching unit 53, and a non-display unit 54, and the display method in the second display unit 48 is the same. Although different from that of the first embodiment in different points, other parts are common to the first embodiment. For this reason, mainly different parts will be described, and common parts will be denoted by common reference numerals and description thereof will be omitted. The microscope system 11 of the second embodiment has the same appearance as that shown in FIG.

  The second control device 16 is configured by a computer such as a general personal computer (PC). As shown in FIGS. 1 and 6, the second control device 16 includes a second display unit 48 configured by a liquid crystal display and the like, and an image recognition unit that is software installed in a storage device such as a built-in hard disk. 52, a display switching means 53, a non-display means 54, and a keyboard 49 and a mouse 50 as input means.

  The image recognition unit 52 is software that performs image recognition between a bright field image acquired together with the light emission image 21 with the first microscope 13 in the past and a bright field live image 22 acquired with the second microscope 15. Yes, it is automatically determined as to which position in the live image 22 the luminescent image 21 overlaps.

  For example, the image recognition unit 52 performs image recognition in the following procedure.

  When the emission image 21 is acquired using the first microscope 13, a bright field image is also acquired at the same time. In the temporal observation at this time, normally, the luminescent image 21 and the bright field image are acquired under the same observation conditions every time. When the pickup work is performed under the second microscope 15, the user designates the colony 41 with respect to the bright-field live image 22 displayed during the pickup work. Various methods can be used to specify the colony 41. For example, the colony 41 in the live image 22 can be specified by surrounding it with a free curve area using a pointer operated by the mouse 50 or the like. Alternatively, the target colony 41 may be designated by clicking the corresponding colony 41 with the mouse 50 and recognizing an area exceeding a certain luminance value centered on the clicked point as a light emitting area.

  When the target colony 41 is instructed, the image recognition unit 52 searches for a bright field image having a colony 41 that matches or is closest to the target colony 41 from a plurality of bright field images stored together with the light emitting image 21.

  Specifically, a rectangular region surrounding the target colony 41 is cut out from the bright-field live image 22 and enlarged to the same magnification as that of the luminescent image 21. At this time, the square area may be set to have the same magnification as that of the light emitting image 21 in advance. The image recognition unit 52 compares pixel values while shifting the position between the square area of the live image 22 and the bright-field image acquired simultaneously with the light-emitting image 21, and finds the position where the difference in pixel values is the smallest. . At this time, the difference between the pixel values is sequentially recorded in a storage device such as a memory built in the second control device 16. A search is performed for each of a plurality of bright field images, and a bright field image including a position having the smallest difference in pixel values is searched from among the plurality of bright field images.

  Subsequently, the emission image 21 acquired simultaneously with the bright field image is read out. The light emission image 21 is displayed at a position where the bright field image exactly overlaps with the live image 22, and the live image 22 and the light emission image 21 are superimposed and displayed.

  The above-described function of acquiring the luminescent image 21 of the biological sample 12 at a high magnification with the first microscope 13 (first image acquisition means) and the biological sample 12 with the second microscope 15 (second image acquisition means). The bright image live image 22 is obtained at a low magnification, and the second control device 16 (second control means) reduces the light-emitting image 21 to the same magnification as the live image 22 to obtain a live image. And a bright field image of the biological sample 12 together with the luminescent image 21 by the first microscope 13 (first image acquisition means). The image recognition unit 52 determines whether or not the partial area of the live image 22 matches the bright field image, and the live image 22 is displayed at the position where these match. Overlaid on the table A function of, is implemented by software installed in the first controller 14 and second controller 16 (computer program).

  The display position of the light emission image 21 can be finely adjusted. If the position is slightly shifted, the user can correct the position. There are various methods for setting the user to finely adjust the display position. For example, when the light emission image 21 is selected, the fine adjustment can be automatically performed, or a fine adjustment button provided on the screen of the second display unit 48 is provided. It may be finely adjusted by pressing. Further, when the fine adjustment is completed, the selection of the light emitting image 21 may be made unselected or the display position may be confirmed by pressing a position confirmation button provided on the screen of the second display unit 48. As a result, the display position of the luminescent image 21 cannot be changed.

  In the present embodiment, the overlapping position of the light emission image 21 is searched using the difference in pixel value. However, the present invention is not limited to this, and various general image recognition methods may be used. Further, in the present embodiment, a bright field image is acquired at the same time as the emission image 21 is acquired and is used by the image recognition unit 52. However, when the image recognition can be sufficiently performed using the emission image 21. The light emission image 21 may be directly used for image recognition.

  Further, when searching for the luminescent image 21, it may take time to search for an image including the target colony 41 from the plurality of luminescent images 21 depending on the specifications of the second control device 16. Thus, if you know it contains the target colony 41 on which the light-emitting image 21 in advance, and specify the luminous image 21, the position of the target colonies from the image looking at the image recognition, the live image 22 The luminescent image 21 may be displayed superimposed.

  Further, in the present embodiment, the colony 41 in the bright-field live image 22 is designated and the light-emitting image 21 that matches the colony 41 is read out, but the opposite method may be used. That is, the user designates in advance the light-emitting image 21 that is desired to be overlaid. The image recognition unit 52 compares the pixel value of the bright field image acquired simultaneously with the light emission image 21 with the pixel value of the live image 22 and finds the position where the difference in pixel value is the smallest. When a region where superimposition is possible exists in the live image, the luminescent image 21 is displayed so as to overlap the corresponding region.

  Subsequently, display switching of the luminescent image 21 using the display switching unit 53 will be described. Although various modes can be considered for the display switching unit 53, in this embodiment, the display switching unit 53 is configured as a part of software (program) that performs image processing on the live image 22 and the light-emitting image 21. As a switch for operating the display switching means 53, an icon (button) is provided on the second display section 48, and the display / non-display of the luminescent image 21 can be switched. That is, in the present embodiment, the light emission image 21 is displayed superimposed on the live image 22 while the button for operating the display switching unit 53 is pressed, and the button of the display switching unit 53 is not pressed. The luminescent image 21 is not displayed.

  This display switching means 53 can be used as follows, for example. With the button of the display switching means 53 being pressed, the colony 41 in the biological sample 12 should be picked up with the luminescent image 21 and the live image 22 superimposed and displayed. Then, this time, the button of the display switching means 53 is not pressed and the light emitting image 21 is not displayed. As a result, only the bright-field live image 22 is displayed on the second display unit 48. As a result, the light emission image 21 does not interfere with the operation, and the pickup operation can be performed efficiently.

  As described above, the function of switching between display and non-display of the luminescent image 21 displayed on the live image 22 by the display switching unit 53 is realized by software (computer program) installed in the second control device 16. Yes.

  Next, the non-display unit 54 will be described. In the present embodiment, the non-display means 54 is configured as a part of software (program) that performs image processing on the live image 22 and the light-emitting image 21. The non-display means 54 acquires the live image 22 at a constant time interval, and the brightness value (pixel value) of one live image 22 and the brightness value (pixel value) of the live image 22 after a lapse of a fixed time. Compare Then, the non-display unit 54 determines that the pick-up operation has been completed because the colony 41 has disappeared in the region where the luminance value (pixel value) has greatly decreased. The non-display means 54 sets all the pixel values to zero for a partial area of the luminescent image 21 corresponding to this area, and cancels the overlapping display of this partial area.

  Thus, when there is a change in the pixel value of a partial area of the live image 22 within a certain time, a part of the light emitting image 21 corresponding to the partial area of the live image 22 is displayed by the non-display unit 54. The function of setting the pixel value of the region to zero is realized by software (computer program) installed in the second control device 16.

  In the present embodiment, the non-display unit 54 is automatically operated based on a change in luminance value (pixel value), but the present invention is not limited to this. The non-display means 54 may be manually operated after the pickup operation is completed as described below.

  That is, the user can designate a colony 41 to be hidden on the luminescent image 21 by surrounding the corresponding colony 41 with a free curve area using the mouse 38 of the second control device 16 or the like. Similarly, the user can also click on the corresponding position with the mouse 38 to recognize the area exceeding a certain luminance value centered on that point as the colony 41 and indicate the colony 41 to be hidden. . Then, when an operation indicating that the work of the instruction portion is finished (for example, pressing a picked up button or the like provided on the second display unit 48), all the pixel values in the corresponding area of the light emitting image 21 are set to zero. Thus, the overlapped display of the specified partial area is canceled.

  Further, in the second embodiment, as shown in FIG. 7, the live image 22 and the light emitting image 21 are superimposed and displayed at a location adjacent to the display location of the live image 22 in the second display unit 48. An enlarged image 55 obtained by enlarging the object is displayed. The enlarged image 55 displays a portion where the superimposed display is performed, for example, at a magnification when the light-emitting image 21 is acquired. When a plurality of light-emitting images 21 are included in the bright-field live image 22, one of them can be selected and displayed as the enlarged image 55.

  In the case illustrated in FIG. 7, when the lower left emission image 21 in the live image 22 is selected, the emission image 21 is displayed at the resolution at which the emission image 21 was acquired. In addition, the live image 22 is enlarged to the same magnification as that of the luminescent image 21, and the live image 22 is displayed so as to be superimposed on the luminescent image 21, and the enlarged image 55 is displayed. Further, as shown in FIG. 8, when the target area of the enlarged image 55 is designated, the designated area is displayed as the enlarged image 55 adjacent to the display location of the live image 22 regardless of the presence or absence of the light emitting image 21. You can also

  The live image 22 and the live image 22 are overlapped at a position adjacent to the location where the live image 22 is displayed in the second display unit 48 by the second control device 16 (second control means). The function of displaying the enlarged image 55 obtained by enlarging the emitted light image 21 is realized by software (computer program) installed in the second control device 16.

  According to the microscope system 11 of the second embodiment, the image recognition unit 52 can automatically and easily recognize the position where the live image 22 and the luminescent image 21 match. It can be done smoothly.

  Further, in the second display unit 48, the enlarged image 55 obtained by enlarging the live image 22 and the light emitting image 21 superimposed on the live image 22 may be displayed at a position adjacent to the location where the live image 22 is displayed. Therefore, when the light emitting image 21 is reduced and displayed, it becomes difficult to see, so that the overlapped portion can be easily seen.

  Further, since the display switching means 53 and the non-display means 54 are provided, the luminescent image 21 can be prevented from being displayed when the luminescent image 21 is disturbed during the pickup operation. The display on the second display unit 48 can be flexibly switched.

  Subsequently, a third embodiment of the microscope system will be described with reference to FIGS. In the microscope system 11 of the third embodiment, the first stage 28 of the first microscope and the second stage 31 of the second microscope are each configured by an electric stage, and the light emission image 21 is superimposed. Although it differs from that of the first embodiment and the second embodiment in that the absolute position information of the luminescent image 21 is used for display, the other parts are common to the first embodiment and the second embodiment. doing. For this reason, mainly different parts will be described, and common parts will be denoted by common reference numerals and description thereof will be omitted. The microscope system 11 of the third embodiment has the same appearance as that shown in FIG.

  In the second embodiment, the image recognition unit 52 uses the bright field image acquired together with the luminescent image 21 to be used for alignment of the luminescent image 21, but in the third embodiment, the luminescent image. The light emission image 21 is aligned using the absolute position information of the light emission image 21 together with the light emission image 21.

  In the present embodiment, the first stage 28 of the first microscope and the second stage 31 of the second microscope are each constituted by an electric stage. The first stage 28 of the first microscope is controlled by the first control device 14 in a plane direction perpendicular to the optical axis (X-axis direction, Y-axis direction) and a direction along the optical axis (Z-axis direction). ) And can be moved respectively. Similarly, the second stage 31 of the second microscope is controlled by the second control device 16 in a plane direction (X-axis direction and Y-axis direction) perpendicular to the optical axis and a direction along the optical axis ( In the Z-axis direction).

  When acquiring the luminescent image 21, the first control device 14 stores the position of the first stage 28 at the time of acquiring the luminescent image 21 in association with the luminescent image 21. More specifically, the first control device 14 stores at least the X coordinate and the Y coordinate of the first stage 28 when the emission image 21 is captured, and more preferably the X coordinate, the Y coordinate, and the Z coordinate. Remember everything.

  A method for overlay display of the luminescent image 21 and the live image 22 according to the present embodiment will be described.

  The observation for several days is continued using the first microscope 13 as in the first embodiment. At this time, when the luminescent image 21 is stored, the absolute position information of the first stage 28 is also recorded in association with the luminescent image 21. At this time, a bright-field image and a fluorescent image may be stored in addition to the light-emitting image 21. When acquiring the light emission images 21 at a plurality of locations, the positional information of the first stage 28 is stored in association with each light emission image 21.

  Subsequently, when picking up a few days later, first, the plurality of light-emitting images 21 and the absolute position information stored in association therewith are copied from the first controller 14 to the second controller 16. To do.

  After the biological sample 12 is placed at the same position on the second stage 31 with the second microscope 15, real-time display (live display) is started with the second CCD camera 43, and then the luminescent image 21 is superimposed and displayed. Set to be performed. The light emission image 21 is automatically read according to the absolute position information of the second stage 31. The light-emitting image 21 is reduced and displayed on the live-field image 22 of the bright field image, and is superimposed and displayed. When a plurality of light emission images 21 are included in one live image 22, all the light emission images 21 are displayed according to the absolute position information of the second stage 31.

  At this time, when the stage types of the first microscope 13 and the second microscope 15 are the same, the position coordinates of the stage are the same. Therefore, the display position of the luminescent image 21 is recorded by the first microscope 13. The position information is used as it is. If the stage types are different, the coordinates do not match, so that the position information between the stages is converted, and then the luminescent image 21 is displayed superimposed on the bright field live image 22.

  Further, as in the first and second embodiments, the user can finely adjust the display position of the light-emitting image 21 after superposition. Further, the enlarged image 55 obtained by enlarging the live image 22 and the light emitting image 21 superimposed and displayed at a location adjacent to the display location of the live image 22 is the same as in the second embodiment. Further, the display switching means 53 and the non-display means 54 are also provided as in the second embodiment.

  Subsequently, the second stage 31 is moved to search for a pickup target. When the biological sample 12 is moved by operating the position of the second stage 31, the live image 22 is also updated according to the position of the second stage 31. At this time, for example, when the live image 22 is moved from the position shown in FIG. 9 to the position shown in FIG. 10, the light-emitting image 21 superimposed thereon is also moved together. At this time, the second control device 16 always acquires the position information of the second stage 31 and updates the display position of the light emission image 21 in accordance with the position information after the movement. Further, if another live light image 21 taken in the past is included in the moved live image 22, the corresponding light image 21 is automatically read out and displayed superimposed on the live image 22. On the contrary, when the light emission image 21 is not included in the live image 22 after the movement, the light emission image 21 is not displayed. When only a part of the luminescent image 21 is included in the live image 22 before and after the movement, only a part of the luminescent image 21 is superimposed on the live image 22 and displayed.

  As described above, the function of storing the position at which the light emission image 21 is acquired together with the light emission image 21 by the first control device 14 (first control means) and the light emission by the second control device 16 (second control means). Based on the position where the image 21 is acquired, a function for displaying the luminescent image 21 superimposed on a part of the live image 22 and displaying it on the second display unit 48 by the second control device 16 (second control means). When the live image 22 is moved, the function of moving the luminescent image 21 superimposed on the live image 22 together with the software (computer) installed in the first control device 14 and the second control device 16 Program).

  In the present embodiment, the first stage 28 and the second stage 31 are electric. However, the first stage 28 and the second stage may be a manual stage instead of the electric stage. The same thing can be realized by adding a means for grasping the position of 31, for example, a position sensor.

  Next, a fourth embodiment of the microscope system will be described with reference to FIG. The microscope system 11 of the fourth embodiment differs from that of the first to third embodiments in that the display is superimposed using the relative position information of the luminescent image 21, but the other parts are the first. This is common to the first to third embodiments. For this reason, mainly different parts will be described, and common parts will be denoted by common reference numerals and description thereof will be omitted. The microscope system 11 of the fourth embodiment has the same appearance as that shown in FIG.

  In the present embodiment, as shown in FIG. 11, for example, a mark 62 is attached to the bottom or edge of the petri dish 61 as a reference position that serves as a reference for the observation position of the biological sample 12. There are various other methods for determining the reference position, and fluorescent beads that emit fluorescence without affecting cells may be embedded in the biological sample.

  When acquiring the luminescent image 21 with the first microscope 13, the reference position is first recorded. In this case, a bright field image including the reference position may be acquired and stored in the first control device 14, or only the coordinates of the reference position may be stored separately in the first control device 14. Subsequently, the luminescence image 21 of the biological sample 12 is acquired as usual. The position information stored in association with the luminescent image 21 is relative (relative position information) with reference to the reference position.

  A few days later, when picking up the colony 41, first, the plurality of luminescent images 21 and the relative position information stored in association with them are copied from the first control device 14 to the second control device 16. . Further, the reference position information stored in the first control device 14 is also copied.

  Subsequently, after placing the biological sample 12 at the same position on the second stage 31 with the second microscope 15, after starting the acquisition (real-time display) of the live image 22 with the second CCD camera 43, The reference position acquired by the first microscope 13 is read out. Next, when setting to execute the superimposed display of the luminescent image 21, the second control device 16 starts the current position of the live image 22 displayed on the second display unit 48 from the current stage position. The relative position with respect to is calculated. The second control device 16 automatically reads the light emission image 21 including the calculated relative position information, and displays the light emission image 21 so as to overlap the live image 22.

  The bright field image including the reference position acquired by the first microscope 13 and the live image 22 including the reference position are manually aligned by the user as described in the first embodiment. It may be.

  Further, as in the first and second embodiments, the user can finely adjust the display position of the light-emitting image 21 after superposition. Also in the fourth embodiment, as in the second embodiment, the live image 22 and the light-emitting image 21 are superimposed and displayed at a location adjacent to the display location of the live image 22 in the second display unit 48. An enlarged image 55 obtained by enlarging the image is displayed. Further, the display switching means 53 and the non-display means 54 are also provided as in the second embodiment.

  As described above, as in the fourth embodiment, the luminescent image 21 may be displayed superimposed on the live image 22 using the relative position information.

  The microscope system 11 described in the first to fourth embodiments can be implemented with various modifications without departing from the gist of the invention. For example, in the first to third embodiments, the light emission image 21 and the live image 22 are displayed on the second display unit 48 of the second control device 16, but the EVF ( An electrical view finder) may be provided to display the light emission image 21 on the eyepiece unit 42 so as to overlap the bright field image. Further, in the microscope system 11 described in the first to third embodiments, the first microscope 13 and the second microscope 15 are assumed to be installed at locations physically separated from each other. Although the control apparatus 14 and the 2nd control apparatus 16 are comprised separately, it is not limited to this. Of course, the first control device 14 and the second control device 16 may be realized by one control means (computer). Similarly, the first display unit 36 and the second display unit 48 may be displayed on any one display unit or may be displayed on each other. Further, the microscope system including the first microscope 13 and the second microscope 15 may be realized with a microscope optical system in which at least a part of the optical systems necessary for both microscopes is shared and integrated. Good.

In the first to third embodiments, the first microscope 13 is a light emission microscope, but may be a fluorescence microscope. Moreover, although the 2nd microscope 15 is comprised with the upright microscope, various microscopes, such as an inverted microscope and a fluorescence microscope, may be sufficient. In the above-described example, the pickup operation has been described. However, the present invention can be applied to various cell operations including other operations such as separation of desired cells and intracellular parts in a container or the like. In addition, the first microscope 13 is provided with an incubator 27 for keeping cells, tissues, and the like as biological samples alive, so that the first microscope 13 can be applied to culturable micro organisms such as embryos, microorganisms, and bacteria. However, such an incubator may be provided in the second microscope 15. In addition, when observing a living individual such as a small animal (mouse, rabbit, etc.) with the first microscope 13 and the second microscope 15, the incubator 27 is unnecessary.
Hereinafter, the invention described in the scope of claims of the present application will be appended.
[1]
A first microscope capable of acquiring a faint light image of a biological sample;
A first control device connected to the first microscope and storing the weak light image;
A second microscope capable of acquiring a bright-field live image of the biological sample at a magnification different from that of the weak light image;
A display unit connected to the second microscope and capable of displaying the feeble light image on a part of the bright-field live image so as to have a magnification equivalent to that of the bright-field live image; A second control device having;
A microscope system comprising:
[2]
The first microscope can obtain a bright field image of the biological sample together with the weak light image at a high magnification,
The second control device includes an image recognition unit that can determine whether or not a partial region of the live image and the bright field image match, and the weak light image is displayed at a position where they match. The microscope system according to [1], wherein the microscope system displays the image superimposed on a live image.
[3]
The first control device stores absolute position information of the weak light image together with the weak light image,
The microscope system according to [1], wherein the second control device displays the weak light image superimposed on a part of the live image based on the absolute position information.
[4]
The second control device moves the live image displayed on the display unit together with the weak light image superimposed on the live image, according to [3]. Microscope system.
[5]
The biological sample is provided with a reference point used for overlay display of the weak light image and the live image,
The first control device stores relative position information of the weak light image with respect to the reference point together with the weak light image,
The microscope system according to [1], wherein the second control device displays the weak light image superimposed on a part of the live image based on the relative position information.
[6]
The second control device displays an enlarged image obtained by enlarging the live image and the weak light image superimposed on the live image at a position adjacent to a location where the live image is displayed in the display unit. The microscope system according to any one of [1] to [5], wherein:
[7]
The second control device includes a display switching unit, and the display switching unit can switch between display and non-display of the faint light image displayed superimposed on the live image [1. ] The microscope system of any one of [5].
[8]
The second control device has a non-display unit that hides the weak light image,
The non-display unit is configured to detect a pixel value of a partial region of the weak light image corresponding to a partial region of the live image when a pixel value of the partial region of the live image changes within a predetermined time. The microscope system according to any one of [1] to [5], wherein zero is set to zero.
[9]
A weak light image of the biological sample is acquired in advance by the first image acquisition means,
A live image of a bright field of a biological sample is acquired at a magnification different from the weak light image by the second image acquisition means,
An observation image display method, wherein the weak light image is made to have a magnification equivalent to that of the live image by a control means, and is displayed on a display means so as to be superimposed on a part of the live image.
[10]
The first image acquisition means acquires a bright field image of the biological sample together with the weak light image at a high magnification,
The image recognition means determines whether or not the partial area of the live image matches the bright field image, and displays the weak light image superimposed on the live image at a position where they match. The observation image display method according to [9], which is characterized.
[11]
The control means stores the position where the weak light image is acquired together with the weak light image, and displays the weak light image superimposed on a part of the live image based on the position where the weak light image is acquired. [9] The method for displaying an observation image according to [9].
[12]
The observation image according to [11], wherein when the live image displayed on the display unit is moved, the control unit moves the weak light image superimposed on the live image together. How to display.
[13]
The control means displays a magnified image obtained by enlarging the live image and the weak light image superimposed on the live image at a position adjacent to the place where the live image is displayed in the display means. The method for displaying an observation image according to any one of [9] to [12], which is characterized.
[14]
The control means includes display switching means, and the display switching means can switch between display and non-display of the weak light image displayed superimposed on the live image [9] to [9]. [13] The method for displaying an observation image according to any one of [13].
[15]
The control means has a non-display means for hiding the weak light image,
The non-display unit is configured to display a partial area of the weak light image corresponding to the partial area of the live image when a pixel value of the partial area of the live image is changed within a predetermined time. The display method of an observation image according to any one of [9] to [14], wherein the pixel value is set to zero.
[16]
First image acquisition means for acquiring a weak light image of a biological sample, second image acquisition means for acquiring a bright-field live image of the biological sample, control means for controlling these, and display provided in the control means An observation image display system comprising means,
A function of acquiring a weak light image of the biological sample by the first image acquisition means;
A function of acquiring a bright-field live image of a biological sample by the second image acquisition means at a magnification different from that of the weak light image;
A function of causing the weak light image to have a magnification equivalent to that of the live image by the control unit, and displaying the image on the display unit so as to overlap with a part of the live image;
A program characterized by realizing.
[17]
A function of acquiring a bright-field image of the biological sample together with the weak light image at a high magnification by the first image acquisition means;
The image recognition unit provided in the control unit determines whether or not the partial area of the live image matches the bright field image, and the weak light image is converted into the live image at a position where they match. A function to display them in a superimposed manner,
The program according to [16], wherein the program is realized.
[18]
A function for storing the position at which the weak light image is acquired together with the weak light image by the control means;
A function of superimposing and displaying the weak light image on a part of the live image based on the position where the weak light image is acquired by the control means;
The program according to [16], wherein the program is realized.
[19]
[18], wherein when the live image displayed on the display unit is moved by the control unit, a function of moving the weak light image superimposed on the live image together is realized. Program.
[20]
The control means realizes a function of displaying the enlarged image obtained by enlarging the live image and the weak light image superimposed on the live image at a position adjacent to the place where the live image is displayed in the display unit. The program according to any one of [16] to [19], wherein the program is executed.
[21]
First image acquisition means for acquiring a weak light image of a biological sample, second image acquisition means for acquiring a bright-field live image of the biological sample, control means for controlling these, and display provided in the control means An observation image display system comprising: means and display switching means;
A function of acquiring a weak light image of the biological sample at a high magnification by the first image acquisition means;
A function of acquiring a live image of a bright field of a biological sample at a low magnification by the second image acquisition means;
A function of reducing the weak light image by the control means so as to have a magnification equivalent to that of the live image, and displaying the image on the display means so as to be superimposed on a part of the live image;
A function for switching between display and non-display of the weak light image displayed on the live image by the display switching means;
A program characterized by realizing.
[22]
First image acquisition means for acquiring a weak light image of a biological sample, second image acquisition means for acquiring a bright-field live image of the biological sample, control means for controlling these, and display provided in the control means An observation image display system comprising means and non-display means,
A function of acquiring a weak light image of the biological sample at a high magnification by the first image acquisition means;
A function of acquiring a live image of a bright field of a biological sample at a low magnification by the second image acquisition means;
A function of reducing the weak light image by the control means so as to have a magnification equivalent to that of the live image, and displaying the image on the display means so as to be superimposed on a part of the live image;
When there is a change in the pixel value of a part of the live image within a predetermined time, the non-display unit causes a pixel of the part of the weak light image corresponding to the part of the live image. The ability to zero the value,
A program characterized by realizing.

DESCRIPTION OF SYMBOLS 11 ... Microscope system, 12 ... Biological sample, 13 ... 1st microscope, 14 ... 1st control apparatus, 15 ... 2nd microscope, 16 ... 2nd control apparatus, 21 ... Luminous image, 22 ... Live image, 48 ... second display unit, 52 ... image recognition unit, 53 ... display switching unit, 54 ... non-display unit, 55 ... enlarged image

Claims (22)

A first microscope capable of acquiring a faint light image of a biological sample;
A first control device connected to the first microscope and storing the weak light image;
A second microscope capable of acquiring a bright-field live image of the biological sample at a magnification different from that of the weak light image;
A display unit connected to the second microscope and capable of displaying the feeble light image on a part of the bright-field live image so as to have a magnification equivalent to that of the bright-field live image; A second control device having;
A microscope system comprising: The first microscope can obtain a bright field image of the biological sample together with the weak light image at a high magnification,
The second control device includes an image recognition unit that can determine whether or not a partial region of the live image and the bright field image match, and the weak light image is displayed at a position where they match. The microscope system according to claim 1, wherein the microscope system displays the image superimposed on a live image. The first control device stores absolute position information of the weak light image together with the weak light image,
The microscope system according to claim 1, wherein the second control device displays the weak light image superimposed on a part of the live image based on the absolute position information.   The said 2nd control apparatus moves the said weak light image superimposed on the said live image together, when moving the said live image displayed on the said display part, It is characterized by the above-mentioned. Microscope system. The biological sample is provided with a reference point used for overlay display of the weak light image and the live image,
The first control device stores relative position information of the weak light image with respect to the reference point together with the weak light image,
2. The microscope system according to claim 1, wherein the second control device displays the weak light image superimposed on a part of the live image based on the relative position information.   The second control device displays an enlarged image obtained by enlarging the live image and the weak light image superimposed on the live image at a position adjacent to a location where the live image is displayed in the display unit. The microscope system according to any one of claims 1 to 5, wherein:   The said 2nd control apparatus has a display switching means, This display switching means can switch the display of the said weak light image displayed on the said live image, and non-display. The microscope system according to any one of claims 1 to 5. The second control device has a non-display unit that hides the weak light image,
The non-display unit is configured to detect a pixel value of a partial region of the weak light image corresponding to a partial region of the live image when a pixel value of the partial region of the live image changes within a predetermined time. The microscope system according to claim 1, wherein zero is set to zero. A weak light image of the biological sample is acquired in advance by the first image acquisition means,
A live image of a bright field of a biological sample is acquired at a magnification different from the weak light image by the second image acquisition means,
The weak light image is made to have a magnification equivalent to that of the live image by the control means, and is displayed on the display means while being superimposed on a part of the live image ,
The first image acquisition means acquires a bright field image of the biological sample together with the weak light image,
The image recognition means determines whether or not the partial area of the live image matches the bright field image, and displays the weak light image superimposed on the live image at a position where they match. A method for displaying a characteristic observation image. The first image acquisition unit, the display method of the observation image according to claim 9, characterized in that together with the weak light image to obtain a bright field image of the biological sample at a high magnification.   The control means stores the position where the weak light image is acquired together with the weak light image, and displays the weak light image superimposed on a part of the live image based on the position where the weak light image is acquired. The observation image display method according to claim 9, wherein the observation image is displayed.   The observation image according to claim 11, wherein when the live image displayed on the display unit is moved, the control unit also moves the weak light image superimposed on the live image. How to display.   The control means displays a magnified image obtained by enlarging the live image and the weak light image superimposed on the live image at a position adjacent to the place where the live image is displayed in the display means. 13. The observation image display method according to claim 9, wherein the observation image is displayed.   The control unit includes a display switching unit, and the display switching unit is capable of switching between display and non-display of the weak light image displayed to be superimposed on the live image. Item 14. The method for displaying an observation image according to any one of Items13. The control means has a non-display means for hiding the weak light image,
The non-display unit is configured to display a partial area of the weak light image corresponding to the partial area of the live image when a pixel value of the partial area of the live image is changed within a predetermined time. 15. The observation image display method according to claim 9, wherein a pixel value is set to zero. First image acquisition means for acquiring a weak light image of a biological sample, second image acquisition means for acquiring a bright-field live image of the biological sample, control means for controlling these, and display provided in the control means An observation image display system comprising means,
A function of acquiring a weak light image of the biological sample by the first image acquisition means;
A function of acquiring a bright-field live image of a biological sample by the second image acquisition means at a magnification different from that of the weak light image;
A function of causing the weak light image to have a magnification equivalent to that of the live image by the control unit, and displaying the image on the display unit so as to overlap with a part of the live image;
A function of acquiring a bright field image of the biological sample together with the weak light image by the first image acquisition means;
The image recognition unit provided in the control unit determines whether or not the partial area of the live image matches the bright field image, and the weak light image is converted into the live image at a position where they match. A function to display them in a superimposed manner,
A program characterized by realizing. The program according to claim 16, wherein the first image acquisition unit realizes a function of acquiring a bright field image of the biological sample together with the weak light image at a high magnification. A function for storing the position at which the weak light image is acquired together with the weak light image by the control means;
A function of superimposing and displaying the weak light image on a part of the live image based on the position where the weak light image is acquired by the control means;
The program according to claim 16, wherein the program is realized.   19. The function for moving the weak light image superimposed on the live image together with the live image displayed on the display unit by the control unit is realized. Program.   The control means realizes a function of displaying the enlarged image obtained by enlarging the live image and the weak light image superimposed on the live image at a position adjacent to the place where the live image is displayed in the display unit. The program according to any one of claims 16 to 19, wherein the program is executed. First image acquisition means for acquiring a weak light image of a biological sample, second image acquisition means for acquiring a bright-field live image of the biological sample, control means for controlling these, and display provided in the control means An observation image display system comprising: means and display switching means;
The program according to claim 16, wherein the display switching unit realizes a function of switching between display and non-display of the faint light image displayed superimposed on the live image. First image acquisition means for acquiring a weak light image of a biological sample, second image acquisition means for acquiring a bright-field live image of the biological sample, control means for controlling these, and display provided in the control means An observation image display system comprising means and non-display means,
A function of acquiring a weak light image of the biological sample at a high magnification by the first image acquisition means;
A function of acquiring a live image of a bright field of a biological sample at a low magnification by the second image acquisition means;
A function of reducing the weak light image by the control means so as to have a magnification equivalent to that of the live image, and displaying the image on the display means so as to be superimposed on a part of the live image;
When there is a change in the pixel value of a part of the live image within a predetermined time, the non-display unit causes a pixel of the part of the weak light image corresponding to the part of the live image. The ability to zero the value,
A program characterized by realizing.

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