JPH07318782A - Automatic focus detecting device - Google Patents

Abstract

PURPOSE:To provide an automatic focus detecting device constituted so that an AF control action is surely executed by a vertical dark visual field microscope and in the case of fluorescent observation and the discoloring of a sample is minimized at the time of the fluorescent observation. CONSTITUTION:The sample 2 is irradiated with exciting light by operating an external controller 15. A fluorescent image emitted by the sample 2 is observed by an observer through an eyepiece 4 and projected to an image sensor 5 for focusing. The prescribed processing of an image signal obtained in prescribed timing is executed by an analog signal processing circuit 7 and the evaluation calculation thereof is executed according to an operation expression designated by an evaluation function computing element 8. Then, the calculated result is transmitted to a CPU 9. Based on the calculated result, a control signal is transmitted to a stage driving device 14 and a stage 1 is moved in upper and lower directions. Besides, a relative distance between the sample 2 and an objective lens 3 is adjusted so as to execute an automatic focusing action.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an optical device for optically obtaining an image, and more particularly to an automatic focus detecting device in a microscope.

[0002]

2. Description of the Related Art The epi-illumination dark-field spectroscopic method using a microscope is conventionally known as one of the methods for visually detecting defects such as metal scratches. In addition, recently, when observing living tissues and cells, fluorescent staining with a microscope and fluorescence observation thereof have been actively performed.

In the field of microscopes, on the other hand, research and development for mounting an automatic focus detection device (hereinafter referred to as AF device) on the microscope have been actively conducted. For example, in Japanese Unexamined Patent Publication No. 64-54408, the mounting table (stage) is moved by a predetermined constant distance up to a predetermined number of times, and the contrast at each position is detected. A position that is equal to or higher than a predetermined threshold is searched, and the mounting table is moved by a predetermined distance based on the search result, and the contrast at the intermediate position becomes maximum within a range where the contrast is equal to or higher than the threshold. Three consecutive positions are obtained, the contrast value of each position is used, and the position obtained by the weighted average calculation is used as the focal position for alignment.

[0004]

However, there are the following problems when the above-mentioned automatic focusing technique is applied to the epi-illumination dark-field microscope and fluorescence observation. First, as a common point when performing fluorescence observation with the epi-illumination dark-field microscope, when the sample is in the defocused state, the obtained image becomes dark. Further, in the epi-illumination dark-field microscope or fluorescence observation, even if no illumination light is given, it becomes dark.

That is, the conventional AF apparatus cannot distinguish whether the sample is in a dark state due to defocus or simply because there is no illumination light. For example, consider an example of AF control when the shutter of the illumination light is closed in the continuous focus mode in which the subject is once brought into focus and the AF operation is performed by following the movement of the subject. Holds the state.

(2) The shutter is closed so that the dark state occurs. However, the AF device determines that defocusing is the cause of the darkness, and searches the object. (3) However, since there is no illumination light and it is in a dark state, the subject that cannot be captured will continue to be searched.

In order to avoid such a search, when the illumination light is shut off, the microscopist must perform an essentially unnecessary work of temporarily interrupting the AF operation. Efficiency deteriorates and usability is poor.

If the speculum neglects this interruption operation, the operation is performed based on a false judgment, and therefore there is a possibility that an operation that should not be originally performed may occur. Alternatively, the optical element or the like may be damaged.

By the way, a characteristic of the fluorescence observation is the fading of the specimen, which is different from the above-mentioned one. The fading of the sample means that the fluorescence light emitted from the sample is attenuated by the fluorescence-stained sample being irradiated with the excitation light (illumination light). The attenuation of this light is proportional to the intensity of the excitation light, that is, the intensity of the excitation light times the irradiation time of the excitation light. For this reason, the microscopist must avoid irradiating the specimen with excitation light more than necessary when observing the fluorescent specimen. Due to these factors, the AF device must shorten the irradiation time of the excitation light as much as possible in the time required for the AF operation.

Since the conventional AF device does not deal with such a point, it means that the illumination light is emitted even during the time other than the time required for the AF operation, and the fading of the sample proceeds more than necessary. There was a problem that caused it.

Therefore, the present invention provides an automatic focus detection device capable of surely performing AF control in epi-illumination dark-field spectroscope and fluorescence observation, and minimizing fading of a specimen during fluorescence observation. The purpose is to

[0012]

In order to achieve the above object, the present invention provides a light source for irradiating a specimen, which is an object, with illumination light according to a spectroscopic method, and an opening / closing device for blocking or passing the illumination light. There is provided an automatic focus detection device having means and a switch for turning on or off the light source, wherein at least one of the opening / closing means and the switch is linked with the automatic focus detection operation.

Further, the light source has a light source for irradiating a sample as an object with illumination light according to a microscopic method, an opening / closing means for blocking or passing the illumination light, and a switch for turning on or off the light source. Provided is an automatic focus detection device, which forcibly stops or prohibits the automatic focus detection operation when the illumination light is blocked by the opening / closing means or the light source is turned off by the switch.

Further, an automatic light source for irradiating a specimen serving as a subject with illumination light according to a microscopic method, an opening / closing means for blocking or allowing the illumination light to pass therethrough, and an image pickup device for picking up an image of the subject are provided. Provided is an automatic focus detection device, wherein the opening / closing means allows the illumination light to pass therethrough while the subject is being imaged in the focus detection operation, and the opening / closing means blocks the illumination light in other states.

[0015]

In the automatic focus detection device having the above-described structure, at least one of the opening / closing means and the switch for illuminating light performs AF control in cooperation with the automatic focus detecting device, and the illumination light is switched by the opening / closing means. When the light source is turned off by shutting off or the switch, the AF operation is forcibly interrupted. Further, in the AF operation, the opening / closing means allows the illumination light to pass through while the subject image is being captured, and in other states, the opening / closing means blocks the illumination light.

[0016]

Embodiments of the present invention will now be described in detail with reference to the drawings. FIG. 1 shows, as a first embodiment, an example in which the automatic focusing device according to the present invention is applied to an epi-fluorescence observation microscope.

In the structure of the microscope for epi-illumination fluorescence observation, the excitation light emitted from the light source such as the mercury lamp 13 passes through the epi-illumination fluorescent light projection tube 11 and the objective lens 3,
The sample 2 is irradiated. The fluorescence image emitted from the sample 2 passes through the objective lens 3, a part of which is observed by the spectroscope through the eyepiece 4, and a part of which is projected on the image sensor 5 for focus adjustment.

The image sensor drive circuit 6 is a CP
Under the control of U9, a drive pulse is always output to the image sensor 5 at an appropriate timing, and the image sensor 5
The received image is converted into an electric signal at a timing according to the input drive pulse. The image signal converted into an electric signal by the image sensor 5 is subjected to predetermined processing by the analog signal processing circuit 7 and sent to the evaluation function calculator 8.

The evaluation function calculator 8 performs the evaluation calculation according to the calculation formula designated by the CPU 9 and outputs the result to the CPU.
Send to 9. The CPU 9 sends a control signal to the stage driving device 14 on the basis of the result of the evaluation calculation to move the stage 1 in the vertical direction, thereby adjusting the relative distance between the sample 2 and the objective lens 3 for automatic focusing. Is configured to get. Further, the CPU 9 outputs a shutter open / close control signal to the shutter drive circuit 10 as needed to open / close the shutter 12.

The external controller 15 is a device having a switch and a jog dial used by the observer to control the motorized portion of the microscope, and observes operations such as starting and stopping the automatic focusing operation and opening and closing the shutter 12. If the person uses the external controller 15, the information is
Since it is sent to U9, the CPU 9 performs an operation corresponding to the operation.

Next, the operation of the present embodiment configured as described above will be described with reference to the flow chart shown in FIG. First, when operating the external controller 15,
The AF operation is started (step S1), the CPU 9 detects the state of illumination light (step S2), and determines the open / closed state of the shutter 12 (step S3). If the shutter 12 is closed in this determination (NO), the AF operation is prohibited (step S4), a warning to that effect is displayed on the display unit of the external controller 15 (step S5), and the process ends.

However, in step S3, the shutter 1
When 2 is in the open state (YES), the defocus amount is calculated from the image signal of the image sensor 5, and the focus detection operation is performed by the CPU 9 by driving the stage 1.
Is performed (step S6). Even during this focus detection operation, the CPU 9 always detects the open / closed state of the shutter 12 (step S7), and in the state determination of the shutter 12 (step S8), if the shutter 12 is closed (NO). , The AF operation is immediately interrupted (step S
9) A warning is displayed on the external controller 15 (step S10), and the process ends.

However, if it is determined in step S8 that the shutter 12 is held in the open state (YES), the focus detection operation is continued, and then it is determined whether or not the sample 2 is in focus (step S11). If it is not in focus (NO), the process returns to step S6, and if it is in focus (YES), it is determined whether the AF control input by the spectrographer to the external controller 15 is the single mode or the continuous mode (following mode). Yes (step S12). If the AF control is the single mode in this determination, the AF operation ends here (step S
13). On the other hand, if the AF control is in continuous mode (following mode), until the AF operation is canceled,
The focus detection operation is always performed (steps S14 to S17).

Further, steps S14 to S17
In the routine, the CPU 9 detects and monitors the open / closed state of the shutter 12 even during the focus detection operation in the continuous mode (steps S15 and S16), and if the shutter 12 is in operation during the continuous operation. If it is closed (NO), the process proceeds to step S9, the AF operation is cut off, a warning is issued, and the process ends. In step S17, A
If the F operation is canceled (YES), this routine is ended.

According to the configuration and operation of such an embodiment, since unnecessary operation such as continuing to search for a sample that cannot be captured is not performed, the working time performed by the spectroscope is shortened, that is, working efficiency is improved. The usability of the AF device is also improved. Furthermore, it is possible to prevent damage to the sample to be observed and the optical element.

Further, in the present embodiment, only the shutter 12 and the AF operation are interlocked, but the same effect can be obtained by interlocking the switch of the main power source of the mercury lamp 13 which is the light source. .

Further, although the shutter 12 is one of the constituent factors in this embodiment, this is used as an ND filter.
The AF operation may be interrupted by detecting a change in the optical condition of illumination light such as a filter with a sensor or the like.

Further, in this embodiment, A is driven by driving the stage.
Although the F operation is performed, the same effect can be obtained by driving the objective lens. Further, in the present embodiment, only the control for interrupting the AF operation when the shutter is closed during the continuous AF operation,
If the continuous AF is automatically restarted when the shutter is opened again, the usability of the AF device is further improved.

Next, as a second embodiment, an example in which the automatic focusing device according to the present invention is applied to an epi-fluorescence observation microscope will be described.
The construction of the epi-fluorescence observation microscope and the automatic focusing device of the second embodiment is the same as that of the first embodiment, but the operation is different.

The operation of the second embodiment will be described with reference to the flow chart shown in FIG. First, when the external controller 15 is operated, the AF operation is started (step S21), the CPU 9 detects the state of the illumination light (step S22), and determines whether or not the shutter 12 is in the open state (step S23). . Shutter 12
If is closed (NO), the AF operation is prohibited (step S24), a warning to that effect is displayed on the display unit of the external controller 15 (step S25), and the process ends.

However, if the shutter 12 is open in step S23 (YES), the image sensor 5 captures the optical image of the sample 2 (step S2).
6). After that, the CPU 9 closes the shutter 12 (step S27) and calculates the defocus amount of the sample 2 (step S28).

Further, the CPU 9 determines whether or not it is in focus based on the calculated defocus amount (step S2).
9) If not focused (NO), the stage 1 is driven (step S30), and after the driving is completed, the shutter 12 is opened to capture the optical image of the sample 2 (step S3).
1). The CPU 9 performs these controls until focusing is completed. When focusing is completed in step S29 (YE
S), AF control input by the spectrograph to the external controller 15 is single mode or continuous (following mode)
It is determined (step S32), and if it is the single mode, the AF operation is ended. On the other hand, in step S32, A
If the F control is continuous (following mode), the shutter 12 is kept open (step S3).
3), the same AF operation as in steps S26 to S28 described above is continuously performed, and the external controller 15
Then, the operation is performed until the spectroscope cancels the AF operation (steps S35 to S38).

When the spectroscope cancels the AF operation in step S38 (YES), the series of operations ends. According to the operation of the second embodiment, the excitation light is radiated to the sample only during the time of capturing the optical image of the sample in the time required for the AF operation, so that the fluorescence fading can be suppressed as much as possible.

Therefore, the AF apparatus according to the present embodiment can provide a focused and clear sample image with very little fading. Further, in the present embodiment, when detecting that the shutter is closed in the detection of the state of the illumination light after the AF operation is started, the AF operation is prohibited. The operation may be started.

Further, from the viewpoint of suppressing fading of the sample as much as possible, the shutter is automatically closed when the CPU determines that focusing is impossible or when focusing is completed in the single focus mode. Good.

Further, by combining this embodiment with the first embodiment, it becomes possible to provide the image of the sample for which the AF operation is surely performed and the fading is suppressed as much as possible.
In particular, although the present embodiment is only for fluorescence observation, it is obvious that it can be applied to various speculums from the viewpoint of not damaging the specimen.

As described above, according to the automatic focus detecting device of the present invention, the following configurations and effects can be obtained. (1) Automatic focus detection having a light source for irradiating a specimen as a subject with illumination light according to a speculum method, opening / closing means for blocking or passing the illumination light, and a switch for turning on or off the light source In the apparatus, at least one of the opening / closing means and the switch is interlocked with an automatic focus detection operation.

As a result, even in the epi-illumination dark-field spectroscope and the special spectroscope for performing fluorescence observation as described in the embodiment, the malfunction of the focus detection caused by the illumination light can be surely suppressed, and the specimen can be detected. The damage done can be suppressed as much as possible.

(2) It has a light source for irradiating a specimen as a subject with illumination light according to a speculum method, an opening / closing means for blocking or allowing the illumination light to pass through, and a switch for turning on or off the light source. In the automatic focus detection device,
Thus, the automatic focus detection device is characterized in that the automatic focus detection operation is forcibly stopped or prohibited when the illumination light is blocked by the opening / closing means or the light source is turned off by the switch. When the illumination light is not applied to the sample, the malfunction during the AF operation can be suppressed by prohibiting or interrupting the AF operation.

(3) A light source for irradiating a specimen serving as an object with illumination light according to a microscopic method, an opening / closing means for blocking or allowing the illumination light to pass therethrough, and a switch for turning on or off the light source in succession In an automatic focus detection device having means for switching between a focus mode and a single focus mode, while the continuous focus mode is being selected by the switching means, the opening / closing means shuts off the illumination light or the switch turns off the light source. The automatic focus detection device is characterized in that the focus detection operation is interrupted when it is turned on, and the focus detection operation is restarted when the illumination light is passed again by the opening / closing means or the light source is turned on by the switch.

Thus, when the illumination light is not applied to the sample, the AF operation is prohibited or interrupted,
In addition, during continuous focus mode, A
The F operation can be interrupted and restarted automatically. Therefore, the work efficiency of the speculum is improved.

(4) An automatic system having a light source for irradiating a specimen serving as a subject with illumination light according to a microscopic method, an opening / closing means for blocking or allowing the illumination light to pass therethrough, and an image pickup device for picking up an image of the subject. In the focus detecting device, the illumination light is passed by the opening / closing means only when the subject is being imaged during focus detection, and the illumination light is blocked by the opening / closing means in other states. .

As a result, the sample is irradiated with the illumination light only during the time when the sample light image is taken during the AF operation.
Discoloration of the sample during the AF operation can be suppressed as much as possible. Therefore, it is possible to provide the speculum with a clear image with little damage.

(5) A light source for irradiating a specimen as a subject with illumination light according to a spectroscopic method, an opening / closing means for blocking or allowing the illumination light to pass therethrough, and a switching means for switching between a continuous focus mode and a single focus mode. In the automatic focus detection device having the above-mentioned, when the single focus mode is selected by the switching means, the illumination light is blocked by the opening / closing means at the end of focusing.

As a result, in the AF single mode, once the object is focused, the shutter of the illumination light is closed to prevent the sample from being damaged, so that the discoloration of the sample after the AF is completed can be suppressed.

(6) In the case where the light source for irradiating the specimen as a subject with the illumination light according to the microscopic method, the opening / closing means for blocking or allowing the illumination light, and the element for focus detection are missing In an automatic focus detection device having focusability determination means for determining that focus detection is impossible, when the focusability determination means determines that focus detection is impossible, the opening / closing means shuts off the illumination light. An automatic focus detection device characterized by:

As a result, when the focus cannot be detected, the shutter of the illumination light is closed to prevent the specimen from being damaged, so that the fading of the specimen, which cannot be expected by the spectroscope, can be suppressed. As described above, the automatic focus detection apparatus according to the present embodiment is capable of reliably performing the AF operation and damaging the sample even with a speculum having a special characteristic during defocusing such as episcopic dark field or fluorescence observation. Can be suppressed as much as possible. Further, the present invention is not limited to the above-described embodiments, and it goes without saying that various modifications and applications can be made without departing from the scope of the invention.

[0048]

As described above in detail, according to the present invention, it is possible to surely perform AF control in epi-illumination dark-field spectroscope and fluorescence observation, and to minimize discoloration of a specimen during fluorescence observation. It is possible to provide an automatic focus detection device capable of performing the above.

[Brief description of drawings]

FIG. 1 is a diagram showing an example in which an automatic focusing device according to the present invention is applied to a microscope for epi-illumination fluorescence observation as a first embodiment.

FIG. 2 is a flowchart for explaining an operation of the automatic focusing device according to the first embodiment.

FIG. 3 is a flowchart for explaining an operation of the automatic focusing device according to the second embodiment.

[Explanation of symbols]

1 ... Stage, 2 ... Sample, 3 ... Objective lens, 4 ... Eyepiece lens, 5 ... Image sensor, 6 ... Image sensor drive circuit, 7 ... Analog signal processing circuit, 8 ... Evaluation function calculator,
9 ... CPU, 10 ... Shutter drive circuit, 11 ... Epi-fluorescent light projection tube, 12 ... Shutter, 13 ... Mercury lamp, 14 ... Stage drive device, 15 ... External controller.

Claims (3)

[Claims] 1. A light source for irradiating a specimen as a subject with illumination light according to a microscopic method, an opening / closing means for blocking or allowing the illumination light to pass therethrough, and a switch for turning on or off the light source. In the automatic focus detection device, at least one of the opening / closing means and the switch is interlocked with the automatic focus detection operation. 2. A light source for irradiating a specimen as a subject with illumination light according to a microscopic method, an opening / closing means for blocking or passing the illumination light, and a switch for turning on or off the light source. In the automatic focus detection device, the automatic focus detection operation is forcibly stopped or prohibited when the illumination light is blocked by the opening / closing means or the light source is turned off by the switch. 3. An automatic system comprising: a light source for irradiating a specimen as a subject with illumination light according to a microscopic method; an opening / closing means for blocking or passing the illumination light; and an image pickup device for picking up the subject. In the focus detection device, the illumination light is passed by the opening / closing means while the subject is being imaged in the focus detection operation, and the illumination light is blocked by the opening / closing means in other states.