JP2012159854A - Inverted microscope system - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an inverted microscope system shortening a time for applying exciting light to an observation sample in a time lapse observation.SOLUTION: In the time lapse observation, while performing an autofocus, the inverted microscope system does not allow an application of the exiting light to the observation sample, when photographing the observation sample after finishing the autofocus, allows the application of the exiting light to the observation sample, and after finishing the photographing, does not allow the application of the exciting light to the observation sample again.

Description

  The present invention relates to a microscope system, an observation method, and an observation program for observing a specimen as an observation body using a microscope, and in particular, a microscope used for so-called time-lapse observation that performs observation and recording of observation images at predetermined time intervals. The present invention relates to a system, an observation method, and an observation program.

  Conventionally, as a method of observing a specimen which is an observation body using a microscope, there is a method of observing and photographing a microscopic image at regular time intervals to observe temporal changes in the form of the specimen. In the case of such an observation method, in general, in order to focus on the vicinity of the sample first and observe the change of the sample including the movement of the sample, a plurality of images are obtained while changing the coordinates at regular intervals with respect to the optical axis direction. The image of is taken. Such a method is extremely effective as a method of observing a temporal change of a specific specimen, for example, a living cell.

  However, even if the objective lens is focused on the specimen during specimen observation, for example, the dimensions of each machine part change due to changes in ambient temperature due to changes in temperature or the operation of air conditioning equipment, etc. There is a problem that the focal point shifts due to the large change in the distance. In view of this, various focusing devices have been considered to correct these defocuses. However, the specimen is generally placed on a slide glass or enclosed in a slide glass and a cover glass, and the focusing device requires a large driving distance in order to cope with various slide glass and cover glass thicknesses. Come. On the other hand, when shooting multiple images while changing the distance with respect to the optical axis direction, particularly in the case of fluorescent specimens, a very rapid drive operation is performed to shorten the time for irradiating the specimen with excitation light. It is desirable. However, it is difficult to increase the driving speed if the driving distance is large, and it is difficult to increase the driving distance if the driving speed is high.

  As a technique for solving this problem, a technique is disclosed in which coarse driving means and fine driving means are provided, and after focusing is performed by the coarse driving means, focusing is performed by the fine driving means (for example, (See Patent Document 1).

Further, a technique in which a piezo element is used for fine movement driving means is disclosed (for example, see Patent Document 2).
JP-A-5-346528 JP 2003-172878 A

  However, the conventional technology is a technology related to the focusing operation in the optical axis direction, and has not yet reached the improvement of the focusing operation for the specimen placed on the slide glass or enclosed in the slide glass and the cover glass. First, quick and appropriate focusing operation for specimens placed on individual slide glasses, or quick and appropriate focusing operation for specimens enclosed in slide glass and cover glass. There was a problem that it was difficult.

In addition, there is a problem that it is difficult to quickly and appropriately perform the focusing operation at a plurality of points in the XY directions and the change of the focusing position accompanying the change of the objective lens and the excitation light.
The present invention has been made in view of the above-mentioned drawbacks of the prior art, and maintains a high focusing speed even for specimens placed on slide glasses of various thicknesses or encapsulated in slide glasses and cover glasses. An object of the present invention is to provide a microscope system, an observation method, and an observation program that enable quick focusing operation.

  In order to solve the above problems, an inverted microscope system of the present invention is an inverted microscope system capable of observing fluorescence, and a stage for placing an observation sample comprising a transparent body and an observation body placed on the transparent body, and the stage An objective lens disposed below the stage so as to face the observation sample placed on the stage, and focusing for driving at least one of the stage and the objective lens in a direction parallel to the observation optical axis A drive unit, an excitation light source for exciting the observation sample, excitation light introducing means disposed in the observation optical path for introducing the excitation light into the observation optical path, and excitation light from the excitation light source to the observation sample The excitation light control means for controlling whether to irradiate or not to irradiate and the excitation light source are different from the excitation light source. A focus detection light source that emits detection light for autofocus, and a detection that is disposed in the observation optical path, reflects the wavelength component of the detection light with respect to the optical axis direction of the observation optical path, and transmits the wavelength of the excitation light A light introducing unit; a light receiving unit configured to receive the detection light reflected from the transparent body in the observation sample through the objective lens; and the focusing unit is controlled based on a result of the light receiving unit to control the transparent body. An autofocus unit for autofocusing at a predetermined position, an offset amount recording means for recording a distance away from the predetermined position in the optical axis direction, and photographing the observation sample via the objective lens. An imaging device to perform, a time lapse measurement interval setting means for setting a time lapse measurement interval which is an intermittent imaging interval of the imaging device, the imaging device, and the autofocuser An imaging control means for controlling the part, and with respect to the predetermined position which is a position where the detection light is irradiated by the imaging control means and the focus detection light is reflected from the transparent body by the autofocus part. While performing autofocus, the excitation light control means causes the excitation light from the observation excitation light source to be unirradiated to the observation sample, and after the autofocus is completed at the predetermined position, the imaging device When imaging the observation sample at a distance away from the predetermined position in the optical axis direction by the excitation light control means, the excitation light from the excitation light source is irradiated onto the observation sample by the excitation light control means, and the imaging device After the imaging of the observation image is completed by the excitation light control means, the observation sample is again non-irradiated with the excitation light from the excitation light source, It is performed for each time lapse measurement interval recorded in the time lapse measurement interval setting means.

  Further, the inverted microscope system of the present invention is an inverted microscope system capable of fluorescence observation, a stage for placing an observation sample comprising a transparent body and an observation body placed on the transparent body, and the stage placed on the stage. An objective lens disposed below the stage so as to face the observation sample, a focusing drive unit that drives at least one of the stage and the objective lens in a direction parallel to the observation optical axis, and An excitation light source for exciting the observation sample, excitation light introducing means arranged in the observation optical path for introducing the excitation light into the observation optical path, and whether the observation sample is irradiated with excitation light from the excitation light source The excitation light control means for controlling whether to enter the irradiation state and the excitation light source are different from the excitation light source, and the autofocus is applied to the observation sample from below the objective lens. A focus detection light source that emits detection light for detection, and a detection light introduction unit that is disposed in the observation optical path and reflects the wavelength component of the detection light with respect to the optical axis direction of the observation optical path and transmits the wavelength of the excitation light And a light receiving means for receiving the detection light reflected from the transparent body in the observation sample through the objective lens, and controlling the focusing drive unit based on a result of the light receiving means, thereby determining a predetermined value of the transparent body. An autofocus unit that autofocuses the position, an offset amount recording unit that records a distance from the predetermined position in the optical axis direction, and two or more different shooting positions based on the offset An imaging range setting means, an imaging device that images the observation sample through the objective lens, and a time lapse that sets a time lapse measurement interval that is an intermittent imaging interval of the imaging device A fixed interval setting unit; and an imaging control unit configured to control the imaging device and the autofocus unit. The imaging control unit irradiates the detection light, and the autofocus unit reflects the focus detection light from the transparent body. While the autofocus is being performed on the predetermined position, the excitation light from the observation excitation light source is not irradiated on the observation sample by the excitation light control means, and the predetermined position is not irradiated. After the autofocus is completed, the imaging device captures the observation sample at two or more imaging positions that are apart from the predetermined position in the optical axis direction by a certain amount set by the imaging range setting unit. When performing, the excitation light from the excitation light source is irradiated onto the observation sample by the excitation light control means, and the imaging device captures the image. After the imaging of the observation image set by the range setting unit is completed, the time-lapse measurement interval setting unit is configured to cause the excitation light from the excitation light source to be non-irradiated to the observation sample again by the excitation light control unit. It is performed at every time lapse measurement interval recorded in the above.

  According to the present invention, when performing long-term observation analysis of an observation object (test object), it is possible to perform a quick focusing operation, shorten the excitation light irradiation time, and enable a good focusing operation observation. It becomes.

Embodiments of the present invention will be described below with reference to the drawings.
(First embodiment)
FIG. 1 is a diagram showing a configuration of a first embodiment to which a microscope system according to the present invention is applied.

  The technical feature of the first embodiment is that the semi-focusing section motor 22 and the motor control section 24 function as the first driving means according to the present invention, and the piezo element 21 and the piezo control section 25 according to the present invention. It functions as the second drive means.

  In FIG. 1, in an inverted microscope system 1A as an example of a microscope system according to the present invention, a specimen 2 as a test object is placed on a slide glass 3, and the slide glass 3 on which the specimen 2 is further placed is an electric stage. 4 is fixed. The electric stage 4 can be electrically controlled in the XY direction orthogonal to the optical axis, and the control is performed by the stage controller 5.

  The fluorescent light source 6 is a light source for performing fluorescent illumination on the specimen 2. The excitation light emitted from the fluorescent light source 6 is collected by the collector lens 7 and irradiated to the specimen 2 fixed on the electric stage 4 through the excitation filter 8, the dichroic mirror 9, and the objective lens 10a. The specimen 2 is illuminated by The fluorescence emitted from the specimen 2 by being illuminated with the excitation light passes through the dichroic mirror 9 and the absorption filter 11, and is guided to the eyepiece 13 or the CCD camera unit 14 by the optical path switching unit 12. The image of the specimen 2 captured by the CCD camera unit 14 is acquired by the host PC 16 by the video capture board 15. The host PC 16 stores the acquired image on an image memory (not shown). This image memory is usually capable of storing a plurality of images. The electric shutter 17 is a shutter for shielding the excitation light emitted from the fluorescent light source 6, and the control unit 18 can control the shielding of the excitation light.

  The objective lens 10a is attached to the electric revolver 19, and the electric revolver 19 has a function of inserting an arbitrary objective lens 10a or objective lens 10b into the optical path by a signal from the control unit 18. The electric revolver 19 is fixed to a pedestal 23 driven by a focusing motor 22 via a piezo element 21 and is moved in the optical axis direction (Z direction) by a motor control unit 24 controlled from the control unit 18. By doing so, it is possible to relatively drive the distance between the sample 2 and the objective lens 10a.

  On the other hand, the piezo element 21 is disposed between the electric revolver 19 and the gantry 23, and the thickness of the piezo element 21 in the optical axis direction is electrically changed by a piezo drive unit 25 controlled by the control unit 18. The structure is such that the position of the objective lens 10a attached to the interlocking revolver 19 is moved in the direction of the optical axis. Similarly, the distance between the specimen 2 and the objective lens 10a can be relatively driven. The piezo element 21 is an element having characteristics of high speed and high resolution, although the driving range is shorter than the focusing motor 22.

FIG. 2 shows a driving position relationship between the electric revolver 19 and the piezo element 21.
As shown in FIG. 2, the electric revolver 19 can be driven by a semi-focusing motor 22 within a driving distance L between “electric revo Upper Limit” and “electric revo Lower Limit”, and further within that range. In the inside, a high-speed, high-resolution drive is possible in the range of L_p between “piezo upper limit” and “piezo lower limit” by the piezo element 21. In FIG. 2, “Piezo Center” indicates the center coordinates of the drive range L_p by the piezo element 21.

Returning to the description of FIG.
The control unit 18 is connected to the host PC 16 and performs various controls of the inverted microscope system 1A. Further, a jog encoder 38 for a user to input an instruction to drive the electric revolver 19, a pulse counter 39, and an operation unit 40 for inputting various other instructions are connected.

  On the other hand, an infrared semiconductor laser 26 that is an invisible wavelength region corresponds to a measurement light source for performing focusing, and is controlled by a laser control unit 27 connected to the control unit 18.

  Laser light emitted from the semiconductor laser 26 passes through a collimating lens 28 for maintaining parallel light, and half of the beam diameter is cut by the light projecting side stopper 29. Thereafter, only the P-polarized light component is reflected by a PBS (Polarization Beam Splitter) 30 and guided to the sample 2 side. The light beam once condensed by the condenser lens group 31 passes through the chromatic aberration correction lens group 32. The light that has passed through the chromatic aberration correction lens group 32 is polarized by 45 ° by the λ / 4 plate 33 and enters the dichroic mirror 34. Here, since only the infrared region is reflected by the dichroic mirror 34, the reflected light beam forms a spot-shaped image on the surface of the slide glass 3 by the objective lens 10a.

  The light beam reflected by the slide glass 3 is then further polarized by 45 ° by the λ / 4 plate 33 via the objective lens 10a and the dichroic mirror 34, and switched to the S polarization component. Further, the light returns from the chromatic aberration correction lens group 32 and the condenser lens group 31 and enters the PBS 30. Then, since the light flux is an S-polarized component, it passes through the PBS 30 as it is, and passes through the condenser lens group 35 and is imaged on the light receiving sensor 36.

  The light receiving sensor 36 is a two-divided photodiode (A region and B region) arranged around the optical axis. When the sample 2 is in the focus position, as shown in FIG. 3, the spot imaged on the light receiving sensor 36 is a narrow and high-intensity signal, and the sample 2 is above the focus position in the Z direction (rear side). 4, the signal intensity distribution is biased toward the range of the B region as shown in FIG. 4, and when the sample 2 is below the focus position in the Z direction (front pin position), As shown in FIG. 5, the signal intensity distribution is biased toward the range of the A region, and each is converted into a sensor signal.

Then, the detection signal divided and converted into the ranges of the A region and the B region is calculated by the focus determination unit 37 in the sum of the intensities in the respective ranges. That is, as shown in FIG. 6, when the horizontal axis is the Z direction of the electric revolver 19 and the vertical axis is the light intensity incident on each light receiving sensor 36, the A range signal and the B range signal that are symmetric with respect to the focus position. These two curves can be detected. Next, A + B as shown in FIG. 7 and (A−B) / (A + B) as shown in FIG. 8 are calculated from the A range signal and B range signal. In particular, the characteristics as shown in FIG. 8 are called S-shaped curves, and the values are called evaluation function values (hereinafter abbreviated as Ef values).

  The control unit 18 determines the in-focus position direction based on the sign of the Ef value from the focus determination unit 37. For example, when AF is started from the position “1” in FIG. 8, the sign of the Ef value is negative. When the AF is started from the position “2” in FIG. 8, the control of lowering the electric revolver 19 is performed because the sign of the Ef value is positive. The motor control unit 24 drives the distance between the objective lens 10a and the surface of the slide glass 3 relatively in the optical axis direction so that the value becomes 0, and the focusing operation is performed. In addition, the thing in the case of having two front and rear reflecting surfaces like the slide glass 3 also has a function of automatically focusing on the surface of the slide glass 3 from the relationship between the two coordinate positions.

Next, the operation of the first embodiment will be described.
In the first embodiment, as shown in FIG. 9, the case where the specimen 2 is a culture specimen existing in the culture solution on the slide glass 3 will be described.

In FIG. 9, the coordinates indicated by “Center” are the observation center coordinates in the Z direction when the observation of the specimen 2 is started. “Pitch” is an interval at which shooting is performed in the Z direction. When the number of shots from the center is set to 3, the number of shots n is 7, and 7 images are taken at the coordinates shown in (1) to (7). Yes. “Lower Limit” in FIG. 9 is a shooting start coordinate at this time, and “Upper Limit” in FIG. 9 is also a shooting end coordinate. It is assumed that the sample 2 has moved from the position A to the position B.

  When performing observation, the user sets shooting conditions. First, the user instructs to focus on the position of the “Base” coordinate on the surface of the slide glass 3. Then, the control unit 18 drives the semi-focusing unit motor 22 via the motor control unit 24 and moves the electric revolver 19 in the Z direction so that the Ef value of the focusing determination unit 37 becomes 0, Focus on "Base" coordinates.

  Subsequently, the in-focus position is moved from the “Base” coordinate to “Center” which is a coordinate to be observed. The user inputs from the jog encoder 38 connected to the control unit 18, and similarly drives the semi-focusing unit motor 22 via the motor control unit 24 to move the electric revolver 19 in the Z direction, thereby “Center” coordinates. Focus on. After focusing on the “Center” coordinate, the distance from the “Base” coordinate to the “Center” coordinate is stored. That is, the value “Center” − “Base” is the offset amount Off_1.

  Subsequently, the shooting pitch p from the “Center” coordinate and the number of shots n_s on one side are set. In this example, the shooting pitch p and the number of shots n_s are set to be the same around the “Center” position, but may be set separately. In the first embodiment, the shooting pitch is set to p1 and the number of shots on one side is set to 3 in both directions, and the total number of shots n is 7. Next, based on the set shooting pitch and the number of shots, “Lower Limit” coordinates, which are the start coordinates of shooting shown in FIG. 9, are calculated.

  Next, the photographing interval time tp and the number of photographing repetitions n_t in time lap observation are set. In the first embodiment, it is assumed that the shooting interval time is set to tp1 and the number of shooting repetitions is set to five. The setting is performed via the host PC 16. Based on the shooting interval time tp and the number of repetitions of shooting n_t, the shooting end time end_time is calculated from the shooting start time start_time set at the start of shooting.

FIG. 10 is a flowchart showing the flow of the measurement process in the first embodiment.
The operation of the inverted microscope system 1A set as described above will be described using the flowchart of FIG.

  At the start of imaging, the driving coordinates by the piezo element 21 controlled by the piezo driving unit 25 are moved to the “piezo center” coordinates that are the center position. Then, when the measurement is started, in step S101, it is checked whether or not the time lapse measurement is completed by reaching the photographing end time end_time. When shooting is finished (step S101: Y), the process is finished.

  If shooting has not been completed (step S101: N), the process waits until the time lapse shooting interval time tp1 set by the host PC 16 is reached in step S102. When the shooting interval time is reached (step S102: Y), AF control is performed on the “Base” coordinate of the slide glass 3 in step S103, and the “Base” coordinate shown in FIG. To drive.

  Subsequently, after moving to the “Base” coordinate, in step S104, the electric revolver 19 is driven by the “Center” coordinate preset by the host PC 16, that is, the offset amount. Furthermore, in step S105, in order to acquire the Z direction images (1) to (7) in FIG. 9, the focusing position is set to “Lower” by driving the piezo element 21 configured in the electric revolver 19. Move to the “Limit” coordinate.

  After that, in step S106, the excitation light is allowed to pass by opening the shutter 17 for excitation light, and in step S107, the specimen 2 is illuminated with the excitation light and photographing is performed. After this photographing is performed, in step S109, the piezo element 21 is driven, so that the number of photographings n set by the host PC 16 is the same as the photographing pitch set by the host PC 16. Photographing is performed while moving the in-focus position by p1.

  When the predetermined number of shots is completed (step S108: Y), the excitation light is shielded by closing the shutter 17 in step S110. This operation is repeated until the photographing end time end_time is reached and the measurement is completed. With the above operation, the time-lapse image shown in FIG. 11 is acquired.

  The inverted microscope system 1A of the first embodiment configured and controlled as described above applies autofocus to the slide glass 3, so that even when the focal position of the objective lens 10a changes due to a change in ambient temperature. Since the observation object photographing position is set with an offset while the reference position is fixed, the movement of the specimen 2 as the observation object can be reliably measured, and the movement of the photographing point in the Z direction is quickly performed by the piezo element 21. Since it is performed with a simple drive, the time for applying the excitation light can be made extremely short.

  In the first embodiment, the focus is set on the surface of the slide glass 3. However, the present invention is not limited to this, and a culture dish or the like may be used.

The inverted microscope system 1A according to the first embodiment is a mechanism for moving the electric revolver 19 up and down, but may have a function of moving the electric stage 4 up and down, or a so-called upright microscope system. However, the same effect can be obtained.
(Second Embodiment)
Next, a second embodiment to which the microscope system according to the present invention is applied will be described.

FIG. 12 is a diagram showing the configuration of the second embodiment to which the microscope system according to the present invention is applied.
The same components as those in the first embodiment are denoted by the same reference numerals and description thereof is omitted.

  The technical feature of the microscope system 1B according to the second embodiment is that an XY stage that can be electrically driven in the X direction or the Y direction according to the present invention is provided on the electric stage 4, the stage controller 5, and the optical axis. In contrast, it moves in the XY direction, which is a parallel direction, and corresponds to an XY coordinate recording unit having a function of recording coordinates in the XY coordinate recording unit 43. The XY stage is connected to the control unit 18.

In the second embodiment, as shown in FIG. 13, a case will be described in which the culture medium on the slide glass 3 exists at different coordinates like the specimens 2a and 2b.
In FIG. 13, “Center_a” coordinates are the observation center coordinates in the Z direction at the start of observation of the sample 2a, and “Center_b” coordinates are the observation center coordinates in the Z direction at the start of observation of the sample 2b. It is assumed that the XY coordinate of the sample 2a is X-Ya and the XY coordinate of the sample 2b is XYb.

Returning to FIG.
As in the first embodiment, first, the user sets shooting conditions. The motorized stage 4 drives the X-Ya coordinates that are the coordinates of the sample 2a. The user focuses on the position of the “Base” coordinate which is the surface of the slide glass 3.

  Subsequently, the in-focus position is moved from the “Base” position to the “Center_a” coordinate at the start of observation. After focusing on the “Center” coordinate, the distance from the “Base” coordinate to the “Center_a” coordinate is stored. That is, the value “Center_a” − “Base” is the offset amount Off — 1a in the X-Ya coordinates.

  Subsequently, the shooting pitch p from the “Center_a” coordinate and the number of shots on one side n_s are set. In this example, the shooting pitch p and the number of shots n_s are set to be the same around the “Center” position, but may be set separately for each XY coordinate. In the second embodiment, the shooting pitch p1 and the number of shots on one side are set to 3 in both directions. Next, “Lower Limit_a” coordinates, which are the start coordinates of shooting, are calculated from the set shooting pitch and number of shots.

  Next, the motorized stage 4 drives the X-Yb coordinates that are the coordinates of the sample 2b. As in the case of the sample 2 a, the user focuses on the position of the “Base” coordinate that is the surface of the slide glass 3. Subsequently, the in-focus position is moved from the “Base” position to the “Center_b” coordinate to be observed. After focusing on the “Center_b” coordinate, the distance from the “Base” coordinate to the “Center_b” coordinate is stored. That is, the offset amount Off_1b in the value X-Yb coordinates of "Center_b"-"Base".

  Subsequently, the shooting pitch p and the number of shots on one side n_s from the “Center_b” coordinates are set. Here, similarly to the sample 2a, it is assumed that the shooting pitch p1 and the number of shots on one side are set to three. Next, based on the set shooting pitch p and the number of shots n_s, “Lower Limit_b” coordinates, which are the shooting start coordinates, are calculated.

  Next, the shooting interval time tp and the number of shooting repetitions n_t are set. In the second embodiment, it is assumed that the shooting interval time is set to tp1 and the number of shooting repetitions is set to five. The setting is performed via the host PC 16. Based on the shooting interval time tp and the number of shooting repetitions n_t, the shooting end time end_time is calculated from the shooting start time start_time set at the start of shooting.

FIG. 14 is a flowchart showing the flow of measurement processing in the second embodiment.
At the start of imaging, the driving coordinate by the piezo element 21 is moved to the “piezo center” coordinate which is the center position. Then, when the measurement is started, in step S101, it is checked whether or not the measurement is finished by reaching the photographing end time end_time. When the measurement is finished (step S101: Y), the process is finished.

  If the measurement has not ended (step S101: N), the process waits until the measurement interval set by the host PC 16 is reached in step S102. When the measurement interval is reached (step S102: Y), in step S201, the XY drive of the electric stage 4 is performed in the order of setting a plurality of measurement points. Then, the stage XY is driven to the X-Ya coordinate which is the first measurement point, and in step S103, AF control is performed on the “Base” coordinate of the slide glass 3 of the X-Ya coordinate, and the semi-focusing is performed. The motor 22 is driven to the “Base” coordinate shown in FIG.

  Subsequently, after moving to the “Base” coordinate, in step S104, the electric revolver 19 is driven by the “Center_a” coordinate preset by the host PC 16, that is, the offset amount. Furthermore, in step S105, in order to acquire a Z direction image, the focus position is driven to “Lower_Limit_a” by driving the piezo element 21 configured in the electric revolver 19.

  After that, in step S106, the excitation light is allowed to pass by opening the shutter 17 for excitation light, and in step S107, the specimen 2 is illuminated with the excitation light and photographing is performed. After this shooting is performed, in step S109, the piezo element 21 is driven by the number of shots set by the host PC 16 and by the shooting pitch set by the host PC 16 as well. Shoot while moving.

Then, after the photographing is finished (step S108: Y), the excitation light is shielded by closing the shutter 17.
Thereafter, in step S202, it is confirmed whether or not the processing for all the XY coordinates has been completed. If not completed (step S202: N), the process returns to step S201, and the electric stage is set to the next set XYb coordinates. 4 is driven. Similarly, AF control (step S103) is performed on the “Base” coordinate of the slide glass 3 having the XYb coordinate, and the set number of shots is measured (step S107).

This operation is repeated until the measurement is completed.
The inverted microscope system 1B according to the second embodiment configured and controlled as described above performs autofocus on the slide glass 3 for each of a plurality of measurement points, so that the focus of the objective lens 10a is changed due to a change in ambient temperature. Even when the position changes, the observation object photographing position is set as an offset while the reference position is fixed, so that the movement of the specimen 2 as the observation object can be measured, and the photographing point at each photographing coordinate can be measured. Since the movement in the Z direction is performed by the piezo element 21 with rapid driving, the time for applying the excitation light can be extremely shortened.

In the second embodiment, the case of the electric XY stage has been described. However, the present invention is not limited to this in terms of driving the specimen 2 in the XY direction. There may be.
(Third embodiment)
Next, a third embodiment to which the microscope system according to the present invention is applied will be described.

FIG. 15 is a diagram showing a configuration of a third embodiment to which the microscope system according to the present invention is applied.
The same components as those in the first embodiment are denoted by the same reference numerals and description thereof is omitted.

  The technical feature of the microscope system 1C according to the third embodiment is that the object detection unit 20 corresponds to the object determination unit that determines the type of the inserted objective lenses 10a and 10b according to the present invention. is there.

  As in the first embodiment, the user sets shooting conditions. First, the objective lens 10a having a magnification to be observed is inserted into the optical path, and focusing is performed on the position of the “Base” coordinate which is the surface of the slide glass 3 shown in FIG.

  Subsequently, the in-focus position is moved from the “Base” position to the “Center_ob_a” coordinate which is the observation center coordinate for the objective lens 10a to be observed. Input is made from the jog encoder 38 connected to the control unit 18, and similarly, by driving the focusing unit motor 22 via the motor control unit 24, the electric revolver 19 is moved in the Z direction to match the “Center_ob_a” coordinate. Do chariot. After focusing on the “Center_ob_a” coordinate, the distance from the “Base” coordinate to the “Center_ob_a” coordinate is stored. That is, the value “Center_ob_a” − “Base” is the offset amount Off_ob_a of the selected objective lens 10a.

  Subsequently, the shooting pitch p_ob_a from the “Center_ob_a” coordinate and the number of shots n_s_ob_a on one side are set. In the third embodiment, the shooting pitch p1 and the number of shots on one side are set to 3 in both directions. Next, “Lower Limit_ob_a” coordinates, which are the start coordinates of shooting, are calculated from the set shooting pitch and the number of shots.

  Next, the objective lens 10b having a magnification to be observed next is inserted into the optical path by the electric revolver 19. Subsequently, the in-focus position is moved from the “Center_ob_a” coordinate of the previous objective lens 10a to the “Center_ob_b” coordinate for the selected objective lens 10b to be observed. The movement to the in-focus coordinates is performed by driving a piezo element 21 configured in the electric revolver 19. After focusing on the “Center_ob_b” coordinate, the distance from the “Center_ob_a” coordinate to the “Center_ob_b” coordinate is stored. That is, the value “Center_ob_b” − “Center_ob_a” is the offset amount Off_ob_b of the selected objective lens 10a.

  Subsequently, the shooting pitch p_ob_b from the “Center_ob_b” coordinate and the number of shots n_s_ob_b on one side are set. In the third embodiment, the shooting pitch p1b and the number of shots on one side are set to 3 in both directions. Next, the “Lower Limit_ob_b” coordinates, which are the start coordinates of shooting, are calculated from the set shooting pitch and the number of shots.

  Next, the shooting interval time tp and the number of repeated shootings n_t are set. In the third embodiment, it is assumed that the shooting interval time is set to tp1 and the number of shooting repetitions is set to five. The setting is performed via the host PC 16. From the shooting interval time tp and the number of repetitions of shooting n_t, the shooting end time end_time is calculated from the shooting start time start_time set at the start of shooting.

FIG. 17 is a flowchart showing the flow of measurement processing in the third embodiment.
When the measurement is started, the driving coordinate by the piezo element 21 is moved to the “piezo Center” coordinate which is the center position, and in step S101, it is checked whether the time lapse measurement corresponding to the photographing end time end_time is completed. When the measurement is finished (step S101: Y), the process is finished.

  If the measurement has not been completed (step S101: N), the process waits until the time lapse measurement interval tp1 set by the host PC 16 is reached in step S102. When the measurement interval is reached (step S102: Y), in step S301, the set objective lens 10a is inserted into the optical path. When the objective lens 10a is inserted into the optical path, AF control is performed on the “Base” coordinate of the slide glass 3 in step S103, and the “Base” coordinate shown in FIG. Drive.

  Subsequently, after moving to the “Base” coordinate, in step S104, the electric revolver 19 is driven by the “Center_ob_a” coordinate preset by the host PC 16, that is, the offset amount. Furthermore, in step S105, in order to acquire a Z direction image, the focus position is driven to “Lower_Limit_ob_a” by driving the piezo element 21 configured in the electric revolver 19.

  After that, in step S106, the excitation light is allowed to pass by opening the shutter 17 for excitation light, and in step S107, the specimen 2 is illuminated with the excitation light and photographing is performed. After this photographing is performed, in step S109, the piezo element 21 is driven, so that the number of photographings n set by the host PC 16 is the same as the photographing pitch set by the host PC 16. Photographing is performed while moving the in-focus position by p1.

Then, after the photographing is finished (step S108: Y), the excitation light is shielded by closing the shutter 17.
Thereafter, in step 302, it is confirmed whether or not the processing for the set objective lens 10a or 10b has been completed. If it has not been completed (step S302: N), in step S303, the next set objective lens 10b is selected. Insert into the optical path. Then, after the objective lens 10b is inserted into the optical path, the process returns to step S105, and the piezoelectric element 21 configured in the electric revolver 19 is driven to move to the “Lower_Limit_ob_b” coordinates set in advance by the host PC 16. Subsequently, in step S106, the excitation light is allowed to pass by opening the shutter 17 for excitation light. In step S107, the specimen 2 is illuminated with the excitation light and imaging is performed. After this photographing is performed, in step S109, the piezo element 21 is driven, so that the number of photographings n set by the host PC 16 is the same as the photographing pitch set by the host PC 16. Shooting is performed while moving the in-focus position by p1b.

  Then, after the photographing is finished (step S108: Y), the excitation light is shielded by closing the shutter 17. This operation is repeated until the photographing end time end_time is reached and the measurement is completed.

  The inverted microscope system 1C of the third embodiment configured and controlled as described above has a case where there is a focus shift when observations with a plurality of objective lenses 10a and 10b are included in the time lapse measurement condition. Since the observation object photographing position is set with an offset in a state where the reference position is fixed, it is possible to reliably measure the movement of the specimen 2 as the observation object even with a plurality of measurements.

In the third embodiment, the objective lenses 10a and 10b have been described. However, a zoom lens that can continuously change the magnification may be used.
(Fourth embodiment)
Next, a fourth embodiment to which the microscope system according to the present invention is applied will be described.

FIG. 18 is a diagram showing the configuration of the fourth embodiment to which the microscope system according to the present invention is applied.
The same components as those in the first embodiment are denoted by the same reference numerals and description thereof is omitted.

  The technical feature of the microscope system 1D according to the fourth embodiment is that the excitation light selection means for switching the excitation light to be irradiated according to the present invention includes a plurality of excitation filters 8, dichroic mirrors 9, and absorption filters 11. It corresponds to the electric cube turret 42 in which the fluorescent cube unit 41 to be configured can be selectively inserted into the optical path. The electric cube turret 42 is controlled by the control unit 18.

In FIG. 19, the “Center_q_a” coordinate in the sample 2 indicates the observation center coordinate by the excitation light a, and the “Center_q_b” coordinate indicates the observation center coordinate by the excitation light b.
Returning to the description of FIG.

As in the first embodiment, the user sets shooting conditions. First, a cube unit 41 having a magnification to be observed is inserted into the optical path, and the position of the “Base” coordinate on the surface of the slide glass 3 shown in FIG. 19 is focused. Subsequently, the in-focus position is moved from the “Base” position to the “Center_q_a” coordinate that is the observation center position for the cube unit 41 to be observed. Input is made from the jog encoder 38 connected to the control unit 18, and similarly, by driving the focusing unit motor 22 via the motor control unit 24, the electric revolver 19 is moved in the Z direction to match the “Center_ob_a” coordinate. Do chariot. After focusing on the “Center_q_a” coordinate, the distance from the “Base” coordinate to the “Center_q_a” coordinate is stored. That is, the value “Center_q_a” − “Base” is the offset amount Off_q_a in the selected cube unit 41.

  Subsequently, the shooting pitch p_q_a from the “Center_q_a” coordinates and the number of shots n_s_q_a on one side are set. In the fourth embodiment, the shooting pitch p1 and the number of shots on one side are set to 3 in both directions. Next, “Lower Limit_q_a” coordinates, which are the start coordinates of shooting, are calculated from the set shooting pitch and the number of shots.

  Next, the electric cube turret 42 is used to insert a cube unit 41b having a magnification to be observed next into the optical path. Subsequently, the in-focus position is moved from the “Center_q_a” coordinate of the previous cube unit 41 to the “Center_q_b” coordinate for the selected cube unit 41b to be observed. The movement to the in-focus coordinates is performed by driving a piezo element 21 configured in the electric revolver 19. After focusing on the “Center_q_b” coordinate, the distance from the “Center_q_a” coordinate to the “Center_q_b” coordinate is stored. That is, the value “Center_q_b” − “Center_q_a” is the offset amount off_q_b in the selected cube unit 41b.

  Subsequently, the shooting pitch p_q_b from the “Center_q_b” coordinates and the number of shots n_s_q_b on one side are set. In the fourth embodiment, the shooting pitch p1b and the number of shots on one side are set to 3 in both directions. Next, based on the set shooting pitch p_q_b and the number of shots on one side n_s_q_b, “Lower Limt_q_b” coordinates, which are shooting start coordinates, are calculated.

  Next, the shooting interval time tp and the number of shooting repetitions n_t are set. In the fourth embodiment, it is assumed that the shooting interval time is set to tp1 and the number of shooting repetitions is set to five. The setting is performed via the host PC 16. Based on the shooting interval time tp and the number of repetitions of shooting n_t, the shooting end time end_time is calculated from the shooting start time start_time set at the start of shooting.

FIG. 20 is a flowchart showing the flow of measurement processing in the fourth embodiment.
When the measurement is started, the driving coordinate by the piezo element 21 is moved to the “piezo Center” coordinate which is the center position, and in step S101, it is checked whether or not the time lapse measurement is completed by reaching the photographing end time end_time. When the measurement is finished (step S101: Y), the process is finished.

  If the measurement has not ended (step S101: N), the process waits until the time lapse measurement interval tp1 set by the host PC 16 is reached in step S102. When the measurement interval is reached (step S102: Y), the set cube unit 41 is inserted into the optical path in step S401. When the cube unit 41 is inserted into the optical path, AF control is performed on the “Base” coordinate of the slide glass 3 in step S103, and the “Base” coordinate shown in FIG. Drive.

  Subsequently, after moving to the “Base” coordinate, in step S104, the electric revolver 19 is driven by the “Center_q_a” coordinate preset by the host PC 16, that is, the offset amount. Furthermore, in step S105, in order to acquire a Z direction image, the focus position is driven to “Lower_Limit_q_a” by driving the piezo element 21 configured in the electric revolver 19.

  After that, in step S106, the excitation light is allowed to pass by opening the shutter 17 for excitation light, and in step S107, the specimen 2 is illuminated with the excitation light and photographing is performed. After this photographing is performed, in step S109, the piezo element 21 is driven, so that the number of photographings n set by the host PC 16 is the same as the photographing pitch set by the host PC 16. Photographing is performed while moving the in-focus position by p1.

Then, after the photographing is finished (step S108: Y), the excitation light is shielded by closing the shutter 17.
Thereafter, in step S402, it is confirmed whether or not the processing for the set cube unit 41 or 41b has been completed. If it has not been completed (step S402: N), the next set cube unit 41b is determined in step S403. Insert into the optical path. Then, the process returns to step S105, and the piezoelectric element 21 configured in the electric revolver 19 is driven to move to the “Lower_Limit_q_b” coordinate set in advance by the host PC 16. Subsequently, in step S106, the excitation light is allowed to pass by opening the shutter 17 for excitation light. In step S107, the specimen 2 is illuminated with the excitation light and imaging is performed. After this photographing is performed, in step S109, the piezo element 21 is driven, so that the number of photographings n set by the host PC 16 is the same as the photographing pitch set by the host PC 16. Shooting is performed while moving the in-focus position by p1b.

  Then, after the photographing is finished (step S108: Y), the excitation light is shielded by closing the shutter 17. This operation is repeated until the photographing end time end_time is reached and the measurement is completed.

  The inverted microscope system 1D according to the fourth embodiment configured and controlled as described above has a focus shift even when observation with a plurality of excitation lights a and b is included in the time lapse measurement condition. Since the photographing position is set as an offset, it is possible to measure the movement of the specimen 2 as an observation body, and the movement of the photographing point in each photographing coordinate in the Z direction is performed quickly by the piezo element 21. The time for applying the excitation light can be made extremely short.

  In the fourth embodiment, the focus detection means is not limited to the method described in the fourth embodiment, and can be applied to other known methods. In the fourth embodiment, the microscope apparatus has been described with the inverted microscope system 1D as a representative. However, the present invention is not limited to such a microscope apparatus, but can be applied to various systems such as a line apparatus incorporating the microscope apparatus. Is also possible.

  The embodiments of the present invention have been described above with reference to the drawings. However, the microscope system to which the present invention is applied is limited to the above-described embodiments and the like as long as the function is executed. Needless to say, a single device, a system composed of a plurality of devices, or an integrated device may be used.

  That is, the present invention is not limited to the embodiments described above, and can take various configurations or shapes without departing from the gist of the present invention.

It is a figure which shows the structure of 1st Embodiment to which the microscope system concerning this invention is applied. It is a drive positional relationship by an electric revolver and a piezo element. It is a figure for demonstrating the relationship (focus position) between a light receiving sensor and a focus position. It is a figure for demonstrating the relationship (rear pin position) between a light receiving sensor and a focus position. It is a figure for demonstrating the relationship (front pin position) of a light receiving sensor and a focus position. FIG. 6 is a diagram (part 1) for explaining the relationship between defocus and incident light intensity; FIG. 6 is a second diagram for explaining the relationship between defocus and incident light intensity; FIG. 10 is a diagram (No. 3) for explaining the relationship between defocus and incident light intensity; It is a figure for demonstrating the relationship between the sample and slide glass in 1st Embodiment. It is a flowchart which shows the flow of the measurement process in 1st Embodiment. It is a figure for demonstrating acquisition of a time-lapse image by the measurement process in 1st Embodiment. It is a figure which shows the structure of 2nd Embodiment to which the microscope system concerning this invention is applied. It is a figure for demonstrating the relationship between the sample and slide glass in 2nd Embodiment. It is a flowchart which shows the flow of the measurement process in 2nd Embodiment. It is a figure which shows the structure of 3rd Embodiment to which the microscope system concerning this invention is applied. It is a figure for demonstrating the relationship between the sample and slide glass in 3rd Embodiment. It is a flowchart which shows the flow of the measurement process in 3rd Embodiment. It is a figure which shows the structure of 4th Embodiment to which the microscope system concerning this invention is applied. It is a figure for demonstrating the relationship between the sample and slide glass in 4th Embodiment. It is a flowchart which shows the flow of the measurement process in 4th Embodiment.

DESCRIPTION OF SYMBOLS 1A Inverted microscope system 1B Inverted microscope system 1C Inverted microscope system 1D Inverted microscope system 2 Specimen 2a Specimen 2b Specimen 3 Slide glass 4 Electric stage 5 Stage control part 6 Fluorescence light source 7 Collector lens 8 Excitation filter 8b Excitation filter 9 Dyclock mirror 9b Die Clock mirror 10a Objective lens 10b Objective lens 11 Absorption filter 11b Absorption filter 12 Optical path switching unit 13 Eyepiece 14 CCD camera unit 15 Video capture board 16 Host PC
DESCRIPTION OF SYMBOLS 17 Electric shutter 18 Control part 19 Electric revolver 20 Objective detection part 21 Piezo element 22 Semi-focusing part motor 23 Base 24 Motor control part 25 Piezo control part 26 Infrared semiconductor laser 27 Laser control part 28 Collimating lens 29 Light emission side stopper 30 PBS
Reference Signs List 31 Condensing Lens Group 32 Chromatic Aberration Correction Lens Group 33 λ / 4 Plate 34 Dichroic Mirror 35 Condensing Lens Group 36 Light Receiving Sensor 37 Focus Determination Unit 38 Jog Encoder 39 Pulse Counter 40 Operation Unit 41 Fluorescent Cube Unit 41b Fluorescent Cube Unit 42 Electric Cube turret 43 XY coordinate recording unit

In order to solve the above-mentioned problem, the inverted microscope system of the present invention is the inverted microscope system in which time-lapse observation is performed in which imaging is repeatedly performed at every imaging interval time and morphological change of the test object is observed over time. A transparent member holding the transparent member, a stage on which the transparent member is placed, an objective lens for observing the specimen, an imaging means for photographing the specimen, and the stage and the objective lens in the optical axis direction A focusing driving means for relatively driving, an excitation light source for emitting excitation light for exciting the test object, and the focusing driving means for controlling autofocusing on the transparent member at every imaging interval time. A control means, wherein the control means is in a state in which the test object is not irradiated with the excitation light while the autofocus is performed, and after the autofocus is completed. Wherein while performing imaging of the test object by imaging means is characterized by a state of irradiating the excitation light to the test object.

Further, the inverted microscope system of the present invention is an inverted microscope system capable of observing fluorescence, a transparent member that holds a test object, a stage on which the transparent member is placed, and an objective lens that is disposed below the stage. A focusing drive means for driving at least one of the stage and the objective lens in an optical axis direction, an excitation light source for emitting excitation light for exciting the test object, and an excitation light source disposed in an observation optical path. Excitation light introducing means for introducing into the observation optical path, a focus detection light source that emits detection light for autofocus from the lower side of the objective lens to the test object, using a light source different from the excitation light source, and the observation optical path A detection light introducing means for reflecting the wavelength component of the detection light with respect to the optical axis direction of the observation optical path and transmitting the wavelength component of the excitation light; and A light receiving means for receiving the detection light reflected from the transparent member through the objective lens; an autofocus means for performing autofocus on the transparent member by controlling the focusing drive means from the result of the light receiving means; An offset amount recording unit that records an offset amount from the transparent member to a position that is apart from the transparent member by a certain amount in the optical axis direction, an imaging unit that captures an image of the test object via the objective lens, A time lapse measurement interval setting means for setting an imaging interval time in time lapse observation, and a control means for controlling the auto focus means so as to auto focus the transparent member at each imaging interval time, the control means, While performing autofocus on the transparent member, the excitation light is not irradiated onto the test object. After the autofocus is completed, while the imaging object is imaged by the imaging means at a position away from the transparent member by the offset amount, the imaging object is irradiated with the excitation light, and the imaging is performed. After the imaging by the means is completed, the state in which the excitation light is not irradiated onto the test object is performed at each imaging interval time.
Further, the inverted microscope system of the present invention is an inverted microscope system capable of observing fluorescence, a transparent member that holds a test object, a stage on which the transparent member is placed, and an objective lens that is disposed below the stage. A focusing drive means for driving at least one of the stage and the objective lens in an optical axis direction, an excitation light source for emitting excitation light for exciting the test object, and an excitation light source disposed in an observation optical path. Excitation light introducing means for introducing into the observation optical path, a focus detection light source that emits detection light for autofocus from the lower side of the objective lens to the test object, using a light source different from the excitation light source, and the observation optical path A detection light introducing means for reflecting the wavelength component of the detection light with respect to the optical axis direction of the observation optical path and transmitting the wavelength component of the excitation light; and A light receiving means for receiving the detection light reflected from the transparent member through the objective lens; an autofocus means for performing autofocus on the transparent member by controlling the focusing drive means from the result of the light receiving means; A photographing range setting means for setting two or more different photographing positions separated by a certain amount in the optical axis direction from the transparent member, an imaging means for photographing the observation sample via the objective lens, and the imaging means A time lapse measurement interval setting means for setting an imaging interval time in time lapse observation, and a control means for controlling the auto focus means so as to auto focus the transparent member at each imaging interval time, the control means, While the auto focus is being performed on the transparent member, the excitation light is not irradiated onto the test object, After the focus is completed, a state in which the excitation light is irradiated on the test object while the test object is imaged by the imaging unit at two or more different positions set by the imaging range setting unit In addition, after the imaging by the imaging unit is completed, the excitation light is not irradiated onto the test object at every imaging interval time.

Claims (3)

In an inverted microscope system that can observe fluorescence,
A stage for placing an observation sample comprising a transparent body and an observation body placed on the transparent body;
An objective lens disposed below the stage so as to face the observation sample placed on the stage;
A focusing drive unit for driving at least one of the stage and the objective lens in a direction parallel to the observation optical axis;
An excitation light source for exciting the observation sample;
Excitation light introducing means arranged in the observation optical path for introducing the excitation light into the observation optical path;
Excitation light control means for controlling whether excitation light from the excitation light source is applied to the observation sample or in a non-irradiation state, and a light source different from the excitation light source, from below the objective lens to the observation sample A focus detection light source for emitting detection light for autofocus,
A detection light introducing means that is disposed in the observation optical path, reflects the wavelength component of the detection light with respect to the optical axis direction of the observation optical path, and transmits the wavelength of the excitation light;
A light receiving means for receiving the detection light reflected from the transparent body in the observation sample through the objective lens;
An autofocus unit that performs autofocus on a predetermined position of the transparent body by controlling the focusing drive unit from the result of the light receiving means;
Offset amount recording means for recording a distance away from the predetermined position in the optical axis direction by a certain amount;
An imaging device that photographs the observation sample via the objective lens;
A time lapse measurement interval setting unit that sets a time lapse measurement interval that is an intermittent shooting interval of the imaging device; and a shooting control unit that controls the imaging device and the autofocus unit;
Excitation light control is performed while auto-focusing is performed on the predetermined position, which is a position where the auto-focus unit irradiates the detection light by the imaging control unit and reflects the focus detection light from the transparent body. Means for irradiating the observation sample with the excitation light from the observation excitation light source,
After the autofocus is completed with respect to the predetermined position, when the observation sample is imaged at a distance away from the predetermined position in the optical axis direction by the imaging device, the excitation light control unit performs the imaging. The observation sample is irradiated with excitation light from an excitation light source,
After the imaging of the observation image is completed by the imaging apparatus, the time-lapse measurement interval setting unit records that the excitation light from the excitation light source is not irradiated again by the excitation light control unit. An inverted microscope system, which is performed at each time lapse measurement interval. In an inverted microscope system that can observe fluorescence,
A stage for placing an observation sample comprising a transparent body and an observation body placed on the transparent body;
An objective lens disposed below the stage so as to face the observation sample placed on the stage;
A focusing drive unit for driving at least one of the stage and the objective lens in a direction parallel to the observation optical axis;
An excitation light source for exciting the observation sample;
Excitation light introducing means arranged in the observation optical path for introducing the excitation light into the observation optical path;
Excitation light control means for controlling whether excitation light from the excitation light source is applied to the observation sample or in a non-irradiation state, and a light source different from the excitation light source, from below the objective lens to the observation sample A focus detection light source for emitting detection light for autofocus,
A detection light introducing means that is disposed in the observation optical path, reflects the wavelength component of the detection light with respect to the optical axis direction of the observation optical path, and transmits the wavelength of the excitation light;
A light receiving means for receiving the detection light reflected from the transparent body in the observation sample through the objective lens;
An autofocus unit that performs autofocus on a predetermined position of the transparent body by controlling the focusing drive unit from the result of the light receiving means;
Offset amount recording means for recording a distance away from the predetermined position in the optical axis direction by a certain amount;
Shooting range setting means that is two or more different shooting positions based on the offset,
An imaging device that photographs the observation sample via the objective lens;
A time lapse measurement interval setting unit that sets a time lapse measurement interval that is an intermittent shooting interval of the imaging device; and a shooting control unit that controls the imaging device and the autofocus unit;
Excitation light control is performed while auto-focusing is performed on the predetermined position, which is a position where the auto-focus unit irradiates the detection light by the imaging control unit and reflects the focus detection light from the transparent body. Means for irradiating the observation sample with the excitation light from the observation excitation light source,
After the autofocus is completed with respect to the predetermined position, at two or more shooting positions that are separated from the predetermined position by the imaging device in the optical axis direction by a certain amount set by the shooting range setting unit. When photographing the observation sample, the excitation light control means causes the observation sample to be irradiated with excitation light from the excitation light source,
After the imaging of the observation image set by the imaging range setting unit is completed by the imaging device, the excitation light from the excitation light source is again non-irradiated to the observation sample by the excitation light control unit. An inverted microscope system, which is performed for each time lapse measurement interval recorded in the time lapse measurement interval setting means. The stage is an XY stage that is electrically movable in the X direction or the Y direction, and the inverted microscope system sets a driving amount with respect to the XY coordinates that are pre-positioned on the XY stage. Further comprising means for
After the first driving means focuses on the transparent member, the second driving means performs predetermined drive control on the coordinates positioned on the XY stage. The inverted microscope system according to claim 1 or 2.

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