JPH08286116A - Microscopic photographing device - Google Patents

Abstract

PURPOSE: To provide a microscopic photographing device capable of automatically photographing a good microscopic photograph in focus even in the case an objective lens has a low magnification of about 1 to 4, and also capable of automatically focusing while visually confirming the situation of the visual field and the focusing situation. CONSTITUTION: The image of a sample 0 is photographed on the film 16A of the camera 16 by a microscopic optical system constituted of the objective lens 10 and a photographic lens 12, and a half mirror 14. And also, a luminous flux is partly separated by the half mirror 14 so as to be guided to a CCD camera 20 through the relay lens 18 and the half mirror 30, then, in response to the output, a motor 24 is driven in an automatic focusing device 22 so as to execute an automatic focusing operation of the microscopic optical system. Besides, the luminous flux is partly separated again by the half mirror 30, then, guided to an eyepiece 36, so that the situation of the visual field and the focusing situation are visually confirmed.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a microscope photographing device used for photographing a sample subject to be observed with a microscope.

[0002]

2. Description of the Related Art Conventionally, various microscope image pickup devices are known in relation to image pickup of medical specimen samples and the like.

Conventionally, in a microscope photographing device, a photographer manually manually focuses and then photographs are taken by the microscope photographing device. For example, in FIG.
In the microscope image capturing apparatus as shown in (1), the photographer uses the focusing viewer 50 to focus on the sample subject 53 placed on the stage 54 using a knob (not shown). That is, when focusing visually, after deciding the photographing field of view of the sample subject 53, a part of the light flux imaged on the photographing film 56 is branched just before the photographing film 56, and the photographer's eyes are in focus. Focusing is performed using the focusing viewer 50 (magnification of about 4 times) whose diopter is adjusted as described above. After focusing is complete,
A photograph of the sample subject 53 was taken by using the photograph taking device 55 installed in the upper part of FIG.

[0004]

However, when the magnification of the microscope optical system is as low as 1 to 4 times, the visual depth of focus becomes (very) deep, which makes focusing difficult. Therefore, in order to make the depth of focus shallow, the focusing telescope 51 is used to increase the magnification of the visual optical system. However, even if it is visually determined that the subject is in focus, when the photographer actually shoots the image, the photograph is often out of focus.

In spite of the above measures, this is
This is because the visual depth of focus is deeper than the depth of focus of the photographic film. For example, in the case of the conventional microscope image pickup apparatus, the magnification of the objective lens 52 is 4.0 (NA).
= 0.13), the magnification of the photographing lens is 3.3 times, the magnification of the focusing viewer 50 is 3.9 times, and the magnification of the focusing telescope 51 is 4.0 times. DF = n ・ (ε / m) ・ (1 / N.
When the depth of focus DF is obtained by A.), the visual depth of focus is 44.6 μm, and the depth of focus of the photographic film is 26.
The depth is 6 μm, and the visual depth of focus is still deeper than the depth of focus of the photographic film. The present invention has been made in view of the above circumstances, and it is possible to take a good photograph in focus even when the magnification of the microscope optical system is as low as about 1 to 4 times. An object of the present invention is to provide a novel microscope imaging device that allows an observer to visually check the focusing state at the same time.

[0006]

In order to achieve such an object, a microscope photographing device used for photographing a sample subject to be observed by a microscope according to a first aspect of the present invention, Photographing means for taking a picture, a microscope optical system for enlarging an image of the sample subject and for forming an image on a photographing film surface set in the photographing means, and an optical path from the microscope optical system to the photographing film surface. And a part of the separated light flux for the observation optical system, which is capable of observing the image of the sample subject by the separated light flux. , An image pickup means for picking up an image of the subject by the separated luminous flux, and an image of the sample subject on the basis of an image pickup signal output from the image pickup means. And a automatic focusing means to I, and the depth of focus of the imaging means is shallower than the depth of focus of the photographing means.

A microscope photographing apparatus according to a second aspect of the present invention is a microscope photographing apparatus used for photographing a sample subject to be observed with a microscope, and is a photograph photographing the sample subject. Means, a photographing means for photographing the sample subject, a microscope optical system for enlarging an image of the sample subject and for forming an image on a photographing film surface set in the photographing means, and the microscope optical system An image pickup means for separating a part of the light flux from the optical path reaching the photographic film surface and for picking up an image of the sample subject by the separated light flux, and further separating a part of the light flux separated by the image pickup means. Then, the image of the sample subject is raised based on the observation optical system that allows the image of the subject to be observed by the separated light flux and the image pickup signal output from the image pickup means. And a automatic focusing means to focus the imaging film surface, the depth of focus of the imaging means is shallower than the depth of focus the photographing means.

Further, a microscope photographing apparatus according to a third aspect of the present invention is the apparatus according to the first or second aspect of the invention, wherein the image pickup means has a resolution ε ₁ , the image pickup means has a resolution ε ₂ , and the image pickup means has the same. Assuming that the magnification of the imaging optical system is m ₁ and the magnification of the imaging optical system of the photographing means is m ₂ , the value of (ε ₁ / m ₁ ) of the imaging means is (ε ₂ / m ₂ ) of the photographing means. The value is less than or equal to the value.

Further, in the microscope photographing apparatus according to the invention of claim 4, in the apparatus according to the invention of claim 3, the magnification m ₁ of the image forming optical system of the image pickup means is set higher than the magnification m _{2 of the} photographing means. ing.

According to a fifth aspect of the present invention, there is provided a microscope photographing apparatus according to the third aspect of the invention.
The resolution ε _{1 of the} image pickup means is set higher than the resolution ε _{2 of the} image pickup means.

According to a sixth aspect of the invention, there is provided a microscope photographing apparatus according to the third aspect of the invention.
The resolution ε _{1 of the} image pickup means is set higher than the resolution ε _{2 of} the image pickup means, and the magnification m ₁ of the imaging optical system of the image pickup means is set higher than the magnification m _{2 of the} image pickup means.

[0012]

When the magnification of the microscope optical system is m, the numerical aperture is NA, the refractive index is n, the resolution is ε, and this ε is used as the minimum circle of confusion of the image sensor or the film, the depth of focus: DF As is well known, it is represented by DF = n · (ε / m) · (1 / NA).

Here, since the refractive index n and the numerical aperture NA are determined by the objective lens used in the microscope and the usage conditions, (ε / m) is adjusted and the depth of focus of the image pickup means is taken as described above. The depth of focus is adjusted to be equal to or shallower than the depth of focus. In other words, the value of (ε ₁ / m ₁ ) of the image pickup means is adjusted to be the value of (ε ₂ / m ₂ ) or less of the image pickup means.

As a method of adjusting the above (ε / m) so that the depth of focus of the image pickup means is equal to or less than the depth of focus of the image pickup means, the magnification m _{1 of the} image pickup means is made higher than the magnification m ₂ of the image pickup means. There is a way. On the contrary, there is also a method of making the resolution ε ₁ of the image pickup means higher than the resolution ε ₂ of the image pickup means. There is also a method of adjusting both values of the above (ε / m). In short, when one of the factors (ε / m) has a value in the direction in which the depth of focus of the image pickup unit is made deeper than the depth of focus of the image pickup unit, the other factor is further extracted. The depth of focus of the device is adjusted to be shallow, and the depth of focus as a whole is made shallow.

The "magnification" is defined as the ratio of the size of the image of an object produced by the optical system to the size of the object. High-to-low magnification means that the numerical value of the magnification m is large to small. Is expressed as Also, "resolution" means a camera, a microscope,
It is defined as the ability to distinguish between two points or between two lines with the eyes. High resolution → low resolution means small numerical value of resolution ε →
Expressed as large.

[0016]

EXAMPLES Specific examples will be described below. In FIG. 1, reference numeral 0 indicates a sample subject observed with a microscope. The sample subject 0 is illuminated toward the microscope optical system by the illumination means 11 provided on the opposite side of the microscope optical system called Koehler illumination. The microscope optical system is composed of an objective lens 10 and a lens 12 for photographing, and magnifies the image of the sample subject 0. In addition, the lens 12 for taking a photograph displays the magnified image of the sample subject 0.
An image is formed on the film surface of the photographing film 16A set on the photographing means (for example, still camera) 16.

A half mirror 14 is provided in the optical path from the objective lens 10 to the film surface of the photographic film 16A, and the half mirror 14 separates a part of the light beam. Part of the separated light flux is the relay lens 1
8, the half mirror 30 and the eyepiece 36 can be used for observation by an observer, and a part thereof is further separated by the half mirror 30 and then the imaging lens 3
An image of the sample subject 0 is formed on the light receiving surface of the CCD camera 20 via the image pickup device 2. That is, the CCD camera 20 constitutes “imaging means” and captures an image of the sample subject 0. In this case, the half mirror 30 is the relay lens 18
May be placed in front of. The imaging lens 32 magnifies the image of the sample subject 0 on the CCD camera 20 to achieve autofocus at a desired position, while the observer can view the same field of view as the still camera 16 via the eyepiece lens 36. Can be observed.

The output signal of the CCD camera 20 is sent to the autofocus device 22, and the autofocus device 22 receives the output signal.
The signal input to is output to the display 26 as an image signal, and the image captured by the CCD camera 20 is displayed on the display 2.
It is displayed in 6. At the same time, the autofocus device 22 drives the motor 24 based on the input signal to perform autofocus control of the microscope optical system. The autofocus control method described later is incorporated as a program in the control device 28 such as a host computer, and is implemented by a program process.

The above program may be incorporated in the autofocus device 22 and processed by the program. The autofocus device 22, the motor 24, and the control device 28 constitute "autofocus means". The auto-focus control is performed so that a focused image of the sample subject 0 is formed on the light receiving surface of the CCD camera 20, and at this time, the focused image of the sample subject 0 is automatically taken. The image is formed on 16A. A publicly-known control method can be used for the above-described autofocus control, but a brief description is as follows.

A signal obtained by performing differential or difference calculation on an image signal obtained from a CCD camera 20 as an image pickup means, that is, a contrast signal is used, and the objective lens 10 and the sample subject 0 having the maximum contrast signal. The autofocus control method is performed by detecting the position of the distance (working distance) and determining the focus position. Generally, there are a full scan method, a peak stop method, and a hill climbing method depending on the method of moving the optical system including the objective lens 10. The full scan method is to move the optical system sufficiently from the front focus position to the rear focus position sufficiently, and store the position where the intensity of the obtained contrast signal is maximum, and the optical system It is a method of returning to the position.

The peak stop system is a system in which the taking lens is moved from the front focus position and the taking lens is stopped at the position where the intensity of the obtained contrast signal has the maximum value. The hill climbing method is to move the taking lens in one arbitrary direction first, compare the intensity of the obtained contrast signal before and after the movement, and move it in the same direction if the intensity of the obtained contrast signal increases. Conversely, when the intensity of the contrast signal obtained after the movement is decreased, the contrast signal is moved in the opposite direction, and the photographing lens is stopped at a position where the intensity is maximized during the subsequent movement.

That is, in the "microscope optical system" composed of the objective lens 10 and the photographing lens 12, the "objective lens side object plane" optically conjugate with the film surface of the photographing film 16A is the objective lens. In the optical system composed of 10, the lens 12 for photographing, the relay lens 18, and the image forming lens 32, the light receiving surface of the CCD camera 20 coincides with the object surface on the objective lens side that is optically conjugate, and autofocus control is performed. Is performed so that the object plane on the objective lens side coincides with the sample subject 0. In the above embodiment, the motor 24 is used to move the optical system, that is, the entire microscope to change the distance between the objective lens 10 and the sample subject 0. However, instead of moving the optical system, the sample subject 0 is placed. The stage 37 may be moved.

When a focused image of the sample subject 0 is formed on the film surface of the photographic film 16A by the autofocus control, the photographic device 16 may take a picture. As shown by a broken line in the figure, a half mirror 30a and an eyepiece lens 36a may be provided to visually observe the sample. Also in this case, the sample subject 0
When the focused image is formed on the film surface of the photographic film 16A, the sample subject 0 is brought into focus even in the visual field of the eyepiece lens 36a.

Next, in the above embodiment, the magnification of the objective lens 10 is set to 4 times, the numerical aperture NA is set to 0.13, the magnification of the photographic lens 12 is set to 3.3 times, and the relay lens 1 is used.
8. The following is an example of the case where the magnification of the imaging lens 32 is 1.0 and the CCD camera 20 is a 1/3 inch type (horizontal resolution: 380) as a photographing means.

At this time, the depth of focus DF film is 26.6 μm when the diameter of the image side minimum circle of confusion in the photographic optical system of the objective lens 10 and the photographic lens 12 is 30 μm (resolution 20 lines / mm). The refractive index n is
A cover glass refractive index of 1.52 was used for the sample. Objective lens 10, photo-taking lens 12, relay lens 18,
The diameter ε of the minimum confusion circle on the image side in the “imaging optical system” by the imaging lens 32 is ε (CCD) = (3L) /, where L is the horizontal length of the CCD element and K (TV lines) is the horizontal resolution. (2K), so the depth of focus of the CCD camera DF
_CCD is DF _CCD = [n ・ (3L)] / [(2mK) ・ (1 / N.
A.)], and DF _CCD = 17.1 μm.

DF _CCD is about 1/2 of DF film,
If an autofocus system as an autofocus means is used for focusing, it is possible to always obtain a focused image on the film surface. By the way, the depth of focus DFeye in the viewer system shown in FIG. 1 at this time is DF = n · [(εeye / NA.) · (250 / M) +1/2 ・
(λ / (NA) ² )], and εeye = 1.4 μm and λ = 550 nm, DFeye = 44.6 μm, so that DFeye is D
It is about twice as much as F film, and even if you focus visually
In many cases, the photos are out of focus.

Under these conditions, five photographers perform visual focusing with a visual observing system to focus 20 persons per person.
The standard deviation of the focus shift when it is carried out once each is 9.5.
μm. On the other hand, the standard deviation of the focus shift was 2.3 μm when the autofocus control performed 100 times of focusing.

That is, in the embodiment, it is possible to realize focusing with much higher accuracy than by visual observation by autofocus control. In the example,
The imaging magnification m in the image pickup means is set so that the magnification of the imaging lens 18 is higher than 1, or the CCD as the image pickup means
The resolution ε of the camera 20 is increased, the value of (ε / m) is increased, and the depth of focus of the image pickup means is reduced. Further, when the image of the sample subject which is in focus on the image pickup means is formed, the image of the sample subject which is in focus may be adjusted so as to be surely formed.

Also, in the embodiment, the half mirror 30 is used.
a and the eyepiece lens 32a may be omitted, the display 26 is for confirmation, and the magnification of the imaging lens 18 is 1.
Since it is 0 times, it can be omitted. In the embodiment, the resolution ε _{1 of the} CCD camera is set smaller than the resolution ε _{2 of the} photographic film and m ₁ and m ₂ are the same, but they may not be the same. When the magnification of the imaging lens 18 is increased, there is an advantage that a CCD camera having a low resolution ε ₁ (that is, an inexpensive CCD camera) can be used. Further, the autofocus accuracy can be improved by increasing the magnification m _{1 of the} image pickup means and the resolution ε ₁ .

FIG. 2 shows another embodiment of the present invention, in which a relay lens 18, a half mirror 30, and an imaging lens 32 are used to
The half mirror 30 projects the light flux that is focused on the CD camera 20.
And further separate the separated light by the eyepiece lens 36.
The image can be observed by an observer through the, and other configurations are the same as those in FIG.

[0031]

As is apparent from the above description, in the present invention, since the depth of focus in the image pickup means is shallower than the depth of focus in the photography means, when the magnification of the microscope optical system is low, for example, Even with about 1 to 4 times, it is possible to take a good photograph in focus with higher accuracy than by visual observation. Also, the observer can visually observe the focusing state at the same time.

[Brief description of drawings]

FIG. 1 is a diagram for explaining an embodiment of the present invention.

FIG. 2 is an explanatory diagram of another embodiment of the present invention.

FIG. 3 is a diagram for explaining a conventional example of the present invention.

[Explanation of symbols]

 0 Sample subject 10 Objective lens 12 Photographic lens 14 Half mirror 16 Photographing device 16A Photographic film 18 Relay lens

Claims (6)

[Claims] 1. A microscope photographing device used for photographing a sample subject to be observed with a microscope, comprising: a photographing means for photographing the sample subject; and enlarging an image of the sample subject. A microscope optical system for forming an image on a photographing film surface set in a photographing means, and a part of a light beam is separated from an optical path from the microscope optical system to the photographing film surface, and the sample light is separated from the sample subject by the separated light beam. An observing optical system capable of observing an image, an image pickup means for further separating a part of the separated light beam for the observation optical system, and picking up an image of the subject by the separated light beam, And an autofocus means for focusing the image of the sample subject on the photographic film surface based on an image pickup signal output from the image pickup means. A microscope photographing apparatus characterized in that a point depth is shallower than a focal depth in the photographing means. 2. A photographing device for a microscope used for photographing a sample subject to be observed with a microscope, comprising: a photographing means for photographing the sample subject, and enlarging an image of the sample subject. A microscope optical system for forming an image on a photographing film surface set in a photographing means, and a part of a light beam is separated from an optical path from the microscope optical system to the photographing film surface, and the sample light is separated from the sample subject by the separated light beam. An image pickup means for picking up an image, an observation optical system for further separating a part of the separated light flux for the image pickup means, and making it possible to observe the image of the subject by the separated light flux, and the image pickup means. An autofocus means for focusing the image of the sample subject on the photographic film surface based on an image pickup signal output from A microscope photographing apparatus characterized in that the degree is shallower than the depth of focus of the photographing means. 3. The microscope photographing apparatus according to claim ₁ , wherein the resolution of the imaging means is ε ₁ , the resolution of the photography means is ε ₂ ,
When the magnification of the imaging optical system from the subject to the imaging means is m ₁ and the magnification of the imaging optical system from the subject to the imaging means is m ₂ , the value of (ε ₁ / m ₁ ) of the imaging means is the imaging means. Of (ε ₂ / m ₂ )
Microscopic imaging device with a value less than or equal to the value. 4. The microscope photographing apparatus according to claim 3, wherein the magnification m ₁ of the imaging optical system from the subject to the imaging means is higher than the magnification m ₂ of the subject to the photographing means. 5. The microscope imaging apparatus according to claim 3, wherein the resolution ε ₁ of the imaging means is higher than the resolution ε ₂ of the imaging means. 6. The microscope photographing apparatus according to claim 3, wherein the resolution ε ₁ of the image pickup means is higher than the resolution ε _{2 of} the image pickup means, and the magnification m ₁ of the imaging optical system of the image pickup means is the magnification m of the image pickup means. A microscope imaging device that is higher than ₂ .