JP2010145531A - Focus detecting device - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a focus detecting device capable of optimizing the incident position of a luminous flux in accordance with the pupil diameter of an objective lens without controlling laser light, even when replacing the objective lens with another objective lens having different pupil diameter, and also performing the focusing by detecting the reflected light amount from a surface of a focusing object, while eliminating an unfavorable influence caused by the reflected light from a surface of a non-focusing object. <P>SOLUTION: In the focus detecting device for focusing by projecting the focus detecting luminous flux on a sample surface through the objective lens 7, a luminous flux incident position adjusting means 4 including inclined surfaces 4a2 and 4b1 arranged facing each other, comprising optical members 4a and 4b capable of adjusting a distance between the inclined surfaces, and by adjusting the distance between the inclined surfaces, capable of adjusting the incident position of the focus detecting luminous flux at the position of the pupil 10 of the objective lens to the predetermined position in the pupil in accordance with the pupil diameter of the objective lens, is provided. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a focus detection apparatus, and more particularly to a focus detection apparatus for automatically performing focusing on a desired surface of a transparent substrate such as a microplate or a slide glass in a microscopic observation apparatus.

  In the field of biotechnology, statistical analysis results for a large number of cells are frequently used in order to clarify the reactions of various living cells under various conditions. For this purpose, a device called a flow cytometer has been used in the past. However, in recent years, images of a large number of cells are acquired by a microscopic observation device, and a statistical analysis result is obtained by analyzing the acquired images. The method is now being used.

By the way, in order to actually test the reaction of living cells under various conditions, it is necessary to analyze a very large number of living cells cultured in a container such as a microplate.
For this reason, as a microscope observation apparatus for acquiring a cell image, an inverted microscope type optical arrangement for observing from below a container such as a microplate is adopted, and a series of image acquisition such as field position change, focusing, imaging, etc. What automates the operation of is desired.

  When automating a microscopic observation apparatus, an automatic focusing method is particularly important. In general, the automatic focusing method is roughly divided into an active method for detecting a focus by irradiating an object (subject) with illumination light such as infrared light, and detecting the amount of reflected light, etc., and a lens. There are two passive methods for detecting a focus using captured images.

  However, in order to clarify the reaction of various living cells under various conditions, a microscopic observation apparatus that acquires cell images needs to acquire a large number of images, and thus has a rapid operation speed. An active focusing method is often employed. As a focus detection apparatus employing such an active method, for example, there is one described in Patent Document 1 below.

By the way, in such an active type focus detection apparatus, a laser is generally used as a light source as described in Patent Document 1, but in order to achieve high focusing accuracy. Generally, the laser beam diameter is made to coincide with the pupil diameter of the objective lens. However, if the laser beam diameter is optimized for a low-magnification objective lens with a large pupil diameter, the laser beam will be vignetted by the pupil of the objective lens when switching to a high-magnification objective lens with a small pupil diameter. This makes it difficult to perform focus detection.
On the other hand, if the laser beam diameter is optimized for a high-magnification objective lens with a small pupil diameter, the accuracy of focus detection deteriorates when switching to a low-magnification objective lens with a large pupil diameter.

Patent Document 1 relates to laser power control of a conventional focus detection light source, and adjusts the laser beam diameter to an objective lens having a large pupil diameter, and sets the laser power of the focus detection light source through a control circuit with different magnifications. It is described that focus detection is performed by changing each lens to obtain a sufficient amount of received light.
However, with the method of changing the power of the focus detection light source via the control circuit, the power of the laser beam becomes too large when an objective lens with a small pupil diameter is used, and the output is high when the objective lens is removed. There is a possibility that a safety problem such as the output of the laser beam to the outside occurs.

However, Patent Document 1 includes a diaphragm filter assembly in which an ND filter and an aperture stop having a different diameter are combined between a laser light source and an objective lens, and the ND filter is mounted with the power of the laser light source kept constant. A configuration is disclosed in which the amount of laser light is adjusted, and the aperture stop is selected so that the diameter of the laser beam matches the pupil diameter of the objective lens.
JP 2007-163738 A

  Further, in a focus detection apparatus adopting an active method, in order to focus on one surface of a transparent substrate, when the reflected light from one surface is detected by a light detector, from the other surface The reflected light is also detected by the photodetector and becomes noise, which adversely affects the focus detection.

  However, in the configuration in which the diameter of the laser beam is matched with the pupil diameter of the objective lens through the aperture stop as in the focus detection device described in Patent Document 1, reflected light from the other surface is also detected by the photodetector. It is impossible to solve the problem of causing noise and adversely affecting the focus detection.

  The present invention has been made in view of the above problems, and even when switching to an objective lens having a different pupil diameter, the laser beam diameter is optimized to match the pupil diameter of the objective lens without dimming the laser beam. In addition, even when switching to an objective lens with a large pupil diameter, the object of focusing on the transparent substrate is eliminated while eliminating the adverse effects of reflected light from the surface on the side that is not the object of focusing on the transparent substrate. An object of the present invention is to provide a focus detection device capable of detecting the amount of reflected light from the surface on the side with high detection sensitivity and performing focusing on the surface on the side to be focused. .

  In order to achieve the above object, a focus detection apparatus according to the present invention projects a focus detection light beam onto a sample surface through an objective lens, and in a focus detection apparatus that performs focusing, tilts arranged in a state of facing each other. It is composed of two optical members having a surface and the interval between the inclined surfaces can be adjusted, and the focus detection at the pupil position of the objective lens by adjusting the interval between the inclined surfaces of the two optical members. A light beam incident position adjusting means capable of adjusting the incident position of the light beam to a predetermined position in the pupil according to the pupil diameter of the objective lens is provided.

  Further, in the focus detection device of the present invention, the light beam incident position adjusting means separates the focus detection light beam symmetrically with respect to the optical axis, and adjusts the interval between the inclined surfaces of the two optical members. It is preferable to adjust the interval between the two separated light beams.

  In the focus detection apparatus of the present invention, the two optical members include a first prism configured with an inclined surface recessed in a V shape symmetric with respect to the optical axis on the incident side and the incident side on the incident side, It is preferable that the first prism includes an inclined surface that protrudes in a V shape that is symmetric with respect to the optical axis and coincides with the V shape of the inclined surface of the first prism, and a second prism that has a flat exit surface.

  In the focus detection apparatus of the present invention, the light beam incident position adjusting means shifts the focus detection light beam in a direction perpendicular to the optical axis by adjusting the interval between the inclined surfaces of the two optical members. Is preferred.

  In the focus detection apparatus of the present invention, it is preferable that the two optical members are composed of two prisms having the same apex angle that are arranged upside down.

  In the focus detection apparatus of the present invention, the light beam incident position adjusting means deforms the focus detection light beam into a concentric circle centered on the optical axis, and adjusts the interval between the inclined surfaces of the two optical members. It is preferable to adjust the diameter of the deformed concentric light beam.

  In the focus detection apparatus of the present invention, the two optical members include a first conical lens formed of an inclined surface recessed in a conical shape symmetric with respect to the optical axis on the incident side, and the incident side on the incident side. The inclined surface of the first conical lens preferably has a concave conical shape protruding in a conical shape symmetrical to the optical axis, and a second conical lens having a flat exit surface.

  Further, in the focus detection device of the present invention, the focus detection device emits illumination light for generating a focus signal for the objective lens and a transparent substrate, and irradiates through the objective lens, and A mask means having a first light shielding portion for shielding a light beam passing through one region of the illumination light beam divided by a first virtual plane along the optical axis of the illumination light, and reflected by the transparent substrate; And a photodetector having two light receiving portions arranged symmetrically across the second virtual plane along the optical axis of the emitted light, from the transparent substrate detected through the two light receiving portions, respectively. The objective lens is focused on the first or second surface of the transparent substrate based on the amount of the reflected light, and the light beam incident position adjusting means is arranged on the first or second surface of the transparent substrate. Near the surface of the objective. When the focal point of the lens is located, the reflected light from the one surface is incident on the two light receiving portions and the reflected light from the other surface is divided in the second virtual plane in one region. The incident position of the light beam for focus detection at the pupil position of the objective lens can be adjusted to a predetermined position in the pupil of the objective lens so as to pass through an area outside the light receiving unit arranged in the area. Is preferred.

  According to the present invention, even when switching to an objective lens having a different pupil diameter, the laser beam diameter can be optimized according to the pupil diameter of the objective lens without dimming the laser beam, and an objective with a large pupil diameter can be obtained. When switching to a lens, the amount of reflected light from the surface of the transparent substrate that is the focus target side is eliminated while eliminating the adverse effects of the reflected light from the surface that is not the focus target surface of the transparent substrate. Is detected with high detection sensitivity, and a focus detection device capable of focusing on the surface on the side to be focused is obtained.

Prior to the embodiment, the background for deriving the present invention will be described.
Prior to the present invention, the applicant of the present invention, from the surface of the transparent substrate that is the object of focusing, while eliminating the adverse effects of the reflected light from the surface of the transparent substrate that is not the object of focusing. The focus detection device of the invention described in Japanese Patent Application No. 2007-316857 is conceived for the purpose of detecting the amount of reflected light with high detection sensitivity and focusing on the surface of the focus target. I put it out.
Therefore, the focus detection device of the invention described in Japanese Patent Application No. 2007-316857 will be described.

  FIG. 9 is an explanatory diagram showing a schematic configuration of a microscopic observation apparatus including a focus detection apparatus according to an embodiment of the invention described in Japanese Patent Application No. 2007-316857, and FIG. 10 is a focus detection apparatus provided in the microscopic observation apparatus of FIG. FIG. 11 is an explanatory diagram showing the shapes of the first and second light shielding portions constituting the mask means in FIG. 5 and the regions not shielded and shielded by the first and second light shielding portions in the illumination light beam (for focus detection). Is the light reflected from the surface of the transparent substrate that is the target of focusing on the illumination light (for focus detection) in the region that is not shielded by the first and second light shielding portions shown in FIG. It is explanatory drawing which shows the positional relationship with the two light-receiving parts of the light reflected by the surface of the side which is not made into object. FIG. 12 shows the light reflected by the surface on the side to be focused and the focus target when the focus is set on the surface of the transparent substrate in the focus detection apparatus of FIG. It is explanatory drawing which shows notionally the path | route of the light reflected on the surface of the side which is not. FIG. 13 shows the focus detection apparatus in the state shown in FIG. 12, in the vicinity of the two light receiving portions of the light reflected by the respective surfaces of the transparent substrate on the side that is the focus detection target and the side that is not the focus detection target. It is explanatory drawing which shows an incident state. FIG. 14 shows the light reflected by the surface on the side to be focused and the focal point when the focal point is aligned with the surface of the transparent substrate opposite to the object side in the focus detection apparatus of FIG. It is explanatory drawing which shows notionally the path | route of the light reflected on the surface of the side which is not the object of alignment. FIG. 15 shows the focus detection apparatus in the state shown in FIG. 14 in the vicinity of the two light receiving portions of the light reflected by the respective surfaces of the transparent substrate on the side that is the focus detection target and the side that is not the focus detection target. It is explanatory drawing which shows an incident state. For convenience of explanation, FIGS. 12 and 14 show the point light source and mask means in the present invention and the positions of the photodetectors interchanged. Further, a disturbance light shielding member described later is also omitted.

A microscope observation apparatus provided with the focus detection apparatus in FIG. 9 includes a microscope main body 51, an epi-illumination apparatus 52, and a focus detection apparatus 53.
The microscope main body 51 includes an XY stage 55 on which the test object 54 is placed. Below the XY stage 55, an objective lens 56, half mirrors 57 and 70, an imaging lens 58, and a CCD camera 59 are provided. The XY stage 55 is configured to be movable in a horizontal plane (in a plane perpendicular to the paper surface). The objective lens 56 is configured to be movable in the optical axis direction via the drive unit 60.
The test object 54 uses a microplate 54a made of a light-transmitting member such as polystyrene having a plurality of wells as a container, and the cells cultured in the culture solution 54b are placed on the bottom surface 54a1 of the microplate 54a. The surface 54a11 is fixed by a known technique. The bottom surface portion 54a1 of the microplate 54a corresponds to the transparent substrate in the present invention. The cell used for the test object 54 may be a cell stained by an appropriate method according to the item to be analyzed. Moreover, although the plastic thing was illustrated here as a container to be used, you may be the glass bottom plate which affixed glass on the bottom face.

The epi-illumination device 52 is provided below the XY stage 55 and includes a light source 61 and a lens 62. The light source 61 is composed of a white LED.
The focus detection device 53 includes an objective lens 56, a half mirror 57, and a focus detection unit 53 ′. The focus detection unit 53 ′ is disposed on the reflected light path of the half mirror 57 in the microscope main body 51, and λ / 4. A plate 65, a lens 63, a polarizing beam splitter 64, a masking unit 66, a focusing point light source 67 disposed on the transmitted light path of the polarizing beam splitter 64, and a reflected light path of the polarizing beam splitter 64 An ambient light shielding member 68 and a photodetector 69 are provided. The point light source 67 for focusing is configured by a laser diode so as to emit illumination light for generating a focus signal to the transparent substrate (the bottom surface portion 54a1 of the microplate 54a) of the test object 54. It has become. The disturbance light shielding member 68 passes through a mask means 66 described later, irradiates the test object 54 through the objective lens 56, is reflected by the test object 54, and travels toward the light detector 69 side. Is provided in one region when divided by a second imaginary plane along the optical axis X2 (in FIG. 9, a plane perpendicular to the paper surface including the optical axis X2) so as to shield ambient light that is off the axis. The disturbance light shielding member 68 may be omitted in the case where disturbance light outside a predetermined optical path is hardly generated.
The photodetector 69 is composed of a two-divided photodiode. The two-divided photodiode has two light receiving portions 69a and 69b. The two light receiving portions 69a and 69b are arranged symmetrically with a virtual plane along the optical axis X2 (a plane perpendicular to the paper surface including the optical axis X2 in FIG. 9) interposed therebetween.

As shown in FIG. 10, the mask means 66 has a first light shielding part 66a and a second light shielding part 66b.
When the first light shielding portion 66a is divided by a first virtual plane along the optical axis X1 (a plane perpendicular to the paper surface including the optical axis X1 in FIG. 9) of the luminous flux of the illumination light emitted from the point light source 67. It is formed in a shape and size that shields the light beam passing through one of the regions. In the present embodiment, the first light shielding portion 66a has a rectangular shape, but may have any shape as long as it can shield the light beam on one side of the first light shielding portion 66a.
The second light shielding portion 66b is formed in a similar shape (in this case, a rectangle) to one light receiving portion in the photodetector 69, and the first or second surface of the transparent substrate (the bottom surface portion 54a1 of the microplate 54a). Of the (surface 54a11 or surface 54a12 of the bottom surface portion 54a1), when the focal point of the objective lens 56 is located near one surface, the reflected light from one surface enters the two light receiving portions 69a and 69b and the other As shown in FIG. 11, for example, the light reflected from the surface of the light receiving unit disposed in one region when divided by the second virtual plane (the light receiving unit 69a in the example of FIG. 11). A part of the light flux of the illumination light emitted from the point light source 67 is shielded so as to pass through the other area when divided by the first virtual plane along the optical axis X1 so as to pass through the area outside Is constructed sea urchin.

  In the microscopic observation apparatus including the focus detection apparatus of the present embodiment configured as described above, first, the light emitted from the white LED that is the light source 61 of the epi-illumination apparatus 52 is converted into the lens 62, the half mirror 70, and the half mirror. 57, the object 54 placed on the XY stage 55 is illuminated via the objective lens 56. The light from the test object 54 at this time is guided to the CCD camera 59 via the objective lens 56, the half mirror 57, the half mirror 70, and the lens 58. Thereby, the test object 54 can be imaged via the CCD camera 59.

  In the focus detection device 53, a part of the light emitted from the laser diode 67 is shielded through the light shielding member 66. On the other hand, the light passing through without being blocked by the light blocking member 66 is guided to the test object 54 via the polarization beam splitter 64, the lens 63, the λ / 4 plate 65, the half mirror 57, and the objective lens 58. The reflected light from the test object 54 at this time is guided to the light receiving element 69 side via the objective lens 58, the half mirror 57, the λ / 4 plate 65, the lens 63, and the polarization beam splitter 64.

  Here, in the focus detection device 53, the state in which the reflected light from the bottom surface portion 54a1 of the microplate 54a enters the light receiving element 69 is as follows in relation to the two surface positions of the bottom surface portion 54a1 and the focal position of the objective lens 58. It changes as follows.

First, when the focal position of the objective lens 58 is aligned with the surface 54a11 in contact with the culture solution 54b and the cells in the bottom surface portion 54a1 (hereinafter, the surface on which the objective lens 58 is focused is referred to as a target surface), the target surface 54a11. When the focus of the objective lens 58 is located in the vicinity, the optical path of the focus detection device 53 is traced as shown in FIG.
That is, of the light emitted from the laser diode 67 and reflected by the bottom surface portion 54a1 of the microplate 54a, the light reflected by the target surface 54a11 passes through the objective lens 58 and then is a two-divided photodetector as the light receiving element 69. The image is formed at the upper image formation point 71. On the other hand, the light reflected by the surface 54 a 12 facing the target surface 54 a 11 forms an image at an image point 73 farther from the image point 71.

  Therefore, as shown in FIG. 13, the light reflected by the target surface 54a11 is condensed in a state almost close to a point, enters the two-divided photodetector, and is received by both light receiving portions 69a and 69b of the two-divided photodetector. A uniform amount of light is detected. On the other hand, the light reflected by the surface 54a12 travels toward the two-divided photodetector without being imaged. However, the light emitted from the point light source 67 and reflected by the surface 54a12 can enter the two-divided photodetector. Is shielded by the second light shield 66b of the mask means 66 and is not reflected by the surface 54a12. For this reason, all of the light reflected by the surface 54a12 passes through a position outside the two-divided photodetector.

On the other hand, in the case where the focal position of the objective lens 58 is adjusted to the surface 54a12 opposite to the side in contact with the culture solution 54b and the cells in the bottom surface portion 54a1, the focus of the objective lens 58 is positioned in the vicinity of the target surface 54a12. 14, the optical path of the focus detection device 53 is traced.
That is, of the light emitted from the laser diode 67 and reflected by the bottom surface portion 54a1 of the microplate 54a, the light reflected by the target surface 54a12 passes through the objective lens 58 and then is a two-divided photodetector as the light receiving element 69. The image is formed at the upper image formation point 73 ′. On the other hand, the light reflected by the surface 54a11 facing the target surface 54a12 forms an image at an image forming point 71 ′ closer to the image forming point 73 ′.

  Therefore, as shown in FIG. 15, the light reflected by the target surface 54a12 is condensed in a state substantially close to a point, enters the two-divided photodetector, and is received by both light receiving portions 69a and 69b of the two-divided photodetector. A uniform amount of light is detected. On the other hand, the light reflected by the surface 54a11 is directed toward the two-divided photodetector, but the light in the region that can be incident on the two-divided photodetector out of the light emitted from the point light source 67 and reflected by the surface 54a11 is masked. The light is shielded by the second light shielding part 66b of the means 66 and is not reflected by the surface 54a11. For this reason, all of the light reflected by the surface 54a11 passes through a position away from the two-divided photodetector.

  In this way, the two-divided photodetector 69 that receives only the reflected light from the target surface of the test object 54 captures the light quantity information of the reflected light and outputs it as a focusing signal to a control unit (not shown). And this control part calculates the evaluation function of (AB) / (A + B) based on this focusing signal. An example of the change in the amount of light detected by the two-divided photodetector in the focus detection apparatus is shown in the graph of FIG. FIG. 17 is a graph showing an example of the evaluation function calculated value based on the data of FIG. In the graph of FIG. 17, the horizontal axis represents the relative distance between the test object 54 and the objective lens, and the vertical axis represents the focusing evaluation value. In focusing, the objective lens 58 is moved up and down via the drive unit 60 so that the evaluation function calculation value becomes zero. When the objective lens 58 moves to a position where the evaluation function calculation value becomes 0, the focal point of the objective lens 58 is positioned on the target surface of the test object 54, and focusing is achieved.

  As described above, according to the focus detection apparatus of FIG. 9, the light emitted from the focusing light source 67 is applied to the bottom surface of the sample container through the mask means 66, and the reflected light from the surface on the non-target side is excluded. Therefore, only the light emitted from the point light source 67 and reflected from the target surface can be detected by the light receiving element. For this reason, the reflected light from the surface on the non-target side becomes disturbance light, and high-precision focusing can be performed without adversely affecting the focusing signal.

Here, in the focus detection apparatus of FIG. 9, the applicant of the present invention is directed to mask means in which light in a region that can be incident on the two-divided photodetector out of reflected light from the surface on the non-target side that is emitted from the point light source 67 is masked. Attention is paid to the point where the second light shielding part 66b in FIG.
That is, in the invention shown in FIG. 9, the amount of light transmitted through the mask means 66 is reduced by the amount of light shielded by the second light shield 66b, compared to the case where the second light shield 66b is not provided. By the way, the light passing through the region shielded by the second light shield 66b is light that affects the focusing as disturbance light when the second light shield 66b is not provided. Here, if the light passing through the region shielded by the second light shielding portion 66b can be used as the light to be reflected from the target surface, the reflected light from the surface on the non-target side can be used. In addition to reducing the intensity, the intensity of the reflected light from the target surface incident on the two-divided photodetector can be increased, so that more accurate focusing can be performed.

However, the present applicant adjusts the position of the light beam from the point light source 67 so that the light beam from the point light source 67 passes through a position that is out of the area shielded by the mask means 66 in the focus detection apparatus of FIG. Then, as in the case where the second light shielding portion 66b is provided in the mask means 66, an effect of eliminating the adverse influence of disturbance light on the focusing signal can be obtained, and the light flux from the point light source 67 can be effectively used. In addition, it has been found that the intensity incident on the two-divided photodetector is increased, and more accurate focusing can be performed.
In addition, since the pupil diameter of the objective lens is generally larger as the magnification becomes lower and becomes smaller as the magnification becomes higher, the applicant can adjust the position of the light beam from the illumination light source. When the objective lens with a small magnification and a low magnification objective lens with a large pupil diameter are switched, the light beam from the point light source does not enter the area where the light is shielded via the mask means 66 in the focus detection device of FIG. It came to derive | lead-out this invention which can adjust position so that it may pass.

First Embodiment FIG. 1 is an explanatory diagram showing a schematic configuration of a focus detection apparatus according to a first embodiment of the present invention. FIG. 2 shows an illumination light beam not shielded by the mask means in the focus detection apparatus of FIG. 1 through the light beam incident position adjusting means, with the incident position at the pupil position of the objective lens being a predetermined position in the pupil corresponding to the pupil diameter of the objective lens. FIG. 4 is an explanatory diagram showing a state adjusted to (a), (a) is a view seen from above, and (b) is a view seen from the side. FIG. 3 is an exploded perspective view showing the configuration of the light beam incident position adjusting means used in the focus detection apparatus of FIG. FIG. 4 is a diagram for explaining the principle of adjusting the light beam incident position by the light beam incident position adjusting means. FIG. 5 is an explanatory view showing a numerical example of the light beam incident position adjustment by the light beam incident position adjusting means. FIG. 6 shows a case in which a marginal ray of a light beam having a diameter of 2 mm is incident at a position 4 mm from the optical axis at the pupil position of the objective lens with a refractive index n = 1.51 of the prism constituting the light beam incident position adjusting means. 5 is a graph showing the relationship between the interval between the inclined surfaces of the prisms constituting the light beam incident position adjusting means and the angle formed by the optical axis and the inclined surface of the prisms. For convenience of explanation, FIG. 1 shows the light beam incident position adjusting means rotated 90 degrees around the optical axis.

The focus detection apparatus of the first embodiment includes a focus detection light source 1, a collimator lens 2, a mask means 3, a light beam incident position adjustment means 4, a polarization beam splitter 5, a λ / 4 plate 6, and an objective lens. 7, an imaging lens 8 disposed on the reflected light path of the polarization beam splitter 5, and a photodetector 9. In FIG. 1, 10 is an entrance pupil of the objective lens, and S is a transparent substrate having a surface to be detected.
The focus detection light source 1 is composed of a point light source such as a laser diode, for example, and is focused on a target surface S1 of a transparent substrate (for example, a surface portion or a bottom surface portion of a microplate) S as a test object. Illumination light for generating a signal is emitted.
The collimator lens 2 converts the light from the focus detection light source 1 into a parallel light beam.
The mask means 3 passes through one region when the illumination light beam emitted from the point light source 1 is divided by a virtual plane along the optical axis X1 (a plane perpendicular to the paper surface including the optical axis X1 in FIG. 1). It is formed in a shape and size that can block the light beam. In the present embodiment, the mask means 3 is formed in a rectangular shape. It should be noted that any shape is possible as long as the light flux in this one region can be shielded.
The polarization beam splitter 5 transmits one of linearly polarized light components of incident light, either S-polarized light or P-polarized light, and reflects the other linearly polarized light component.
The λ / 4 plate 6 is disposed on the transmission optical path of the polarizing beam splitter 5, converts one linearly polarized light from the polarizing beam splitter 5 into circularly polarized light, and converts the circularly polarized light from the objective lens 7 into linearly polarized light. Is converted into the other linearly polarized light.
The photodetector 9 is composed of a two-divided photodiode. The two-divided photodiode has two light receiving portions 9a and 9b. The two light receiving parts are arranged symmetrically with a virtual plane along the optical axis X2 (a plane perpendicular to the paper surface including the optical axis X2 in FIG. 1) interposed therebetween.

The light beam position adjusting means 4 is composed of a first prism 4a and a second prism 4b. The first prism 4a is composed of a plane 4a1 on the incident side and an inclined surface 4a2 that is recessed in a V shape symmetrical on the output side with respect to the optical axis X1. The second prism 4b is configured with an inclined surface 4b1 protruding in a V shape symmetrical to the optical axis X1 coincident with the V shape of the inclined surface 4a2 of the first prism 4a on the incident side, and a flat surface 4b2 on the output side. .
The first prism 4a and the second prism 4b are arranged in a state where the inclined surface 4a2 and the inclined surface 4b1 face each other, and are configured such that the interval between the inclined surfaces 4a2 and 4b1 can be adjusted.
The first prism 4a and the second prism 4b separate the focus detection light beam symmetrically with respect to the optical axis X1 and adjust the distance between the inclined surfaces 4a2 and 4b1 to thereby separate the two separated light beams. Configured to adjust.

Then, the light beam incident position adjusting means 4 adjusts the distance between the first prism 4 a and the second prism 4 b, thereby determining the incident position of the focus detection light beam at the position of the pupil 10 of the objective lens 7. It can be adjusted to a predetermined position in the pupil 10 according to the diameter of the pupil 10.
Here, when the light beam incident position adjusting means 4 is switched to an objective lens having a large pupil diameter, of the two surfaces on the transparent substrate (for example, the surface portion or the bottom surface portion of the microplate) S as the test object. When the focal point of the objective lens 7 is located in the vicinity of the target surface S1, the reflected light from the target surface S1 is incident on the two light receiving portions of the two-divided photodiode, and the reflected light from the other surface is on the second virtual plane. The focus detection light beam at the position of the pupil 10 of the objective lens 7 is placed at a predetermined position in the pupil 10 of the objective lens 7 so as to pass through an area outside the light receiving unit arranged in the one area when divided. The incident position can be adjusted (FIG. 2 (a)).

  Here, the principle of the light beam incident position adjustment by the light beam incident position adjusting means 4 will be described from the relationship between the shift amount of the separated light beams depending on the distance between the first prism 4a and the second prism 4b shown in FIG. . For convenience of description, the upper part of the light beam shown in FIG. 2A from the optical axis X1 will be described.

As shown in FIG. 4, the surface interval between the first prism 4a and the second prism 4b is d, the light ray height of the marginal light beam from the optical axis X1 of the light beam incident on the first prism 4a is h ₁ , The angle formed between the exit surface (inclined surface) 4a2 of the first prism 4a and the optical axis X1 is θ, and the angle formed between the straight line perpendicular to the exit surface (inclined surface) 4a2 of the first prism 4a and the optical axis is α, The light is refracted by the straight line perpendicular to the exit surface (inclined surface) 4a2 of the first prism 4a and the entrance surface (inclined surface) 4b1 of the second prism 4b and the exit surface (inclined surface) 4a2 of the first prism 4a. Β is an angle formed with the marginal light beam to be emitted, γ is an angle formed between the marginal light beam refracted by the emission surface (inclined surface) 4a2 of the first prism 4a and the optical axis X1, and the emission surface of the first prism 4a. (Inclined surface) 4a2 and incident surfaces of the second prism 4b ( The length between the line perpendicular to the (slope) 4b1 and the intersection of the exit surface (inclined surface) 4a2 of the first prism 4a and the entrance surface (inclined surface) 4b1 of the second prism 4b is l, the first prism The length of the marginal ray refracted by the exit surface (inclined surface) 4a2 of 4a to the entrance surface (inclined surface) 4b1 of the second prism is l ₁ , and the lengths of the first prism 4a and the second prism 4b If the refractive index is n,
α = π / 2−θ
β = sin ⁻¹ [nsin (π / 2−θ)]
l = dsinθ
l / l ₁ = cosβ Therefore l ₁ = l / cosβ
γ = β-α
= Β- (π / 2-θ)
= Β + θ− (π / 2)
It can be expressed as.

Further, the light beam incident on the first prism 4a at the intersection point of the marginal light ray refracted and emitted from the emission surface (inclined surface) 4a2 of the first prism 4a with the incident surface (inclined surface) 4b1 of the second prism. When the vertical distance from the optical axis X1 until the marginal ray (extension) of the h _2,
h ₂ = l ₁ sin γ
= L ₁ sin [β + θ− (π / 2)]
= (L / cosβ) sin [β + θ− (π / 2)]
= (Dsinθ / cosβ) sin [β + θ− (π / 2)]
Therefore,
d = h ₂ (cos β / sin θ) sin [β + θ− (π / 2)]
It becomes.

FIG. 5 shows a numerical example of the light beam incident position adjustment by the light beam incident position adjusting means 4.
In the example of FIG. 5, the refractive index n = 1.51 of the first prism 4a and the second prism 4b, and the angle formed by the exit surface (inclined surface) 4a2 of the tilted first prism 4a and the optical axis X1 is 60. Degree. When the distance between the exit surface (inclined surface) 4a2 of the first prism 4a and the entrance surface 4b1 of the second prism 4b is 0.74 mm, a marginal ray of a light beam having a diameter of 2 mm is positioned at the pupil 10 of the objective lens 7. Can be incident at a position of 4 mm from the optical axis.
Therefore, when the diameter of the pupil 10 of the objective lens 7 is 10 mm, the interval between the inclined surfaces 4b2 and 4b1 of the prisms 4a and 4b of the light beam input position adjusting means 4 configured as shown in FIG. 5 is adjusted. Thus, the incident light beam can be adjusted to a suitable position that does not cause noise.

  The angles of the inclined surfaces 4b2 and 4b1 formed on the prisms 4a and 4b constituting the light beam incident position adjusting unit 4 with respect to the optical axis X1 are not limited to the example in FIG. For example, as shown in FIG. 6, the pupil of the objective lens 7 is adjusted by adjusting the distance between the inclined surfaces of the prisms 4a and 4b according to the angles of the inclined surfaces 4a2 and 4b1 of the prisms 4a and 4b with respect to the optical axis X1. It is possible to adjust the position of the light beam to a predetermined position in the area 10.

  In the focus detection apparatus of the first embodiment configured as described above, first, light emitted from the focus detection light source 1 is converted into a parallel light beam by the collimator lens 2. Of the converted luminous flux, the luminous flux passing through one region when divided by the first virtual plane along the optical axis X <b> 1 is shielded through the mask means 3. On the other hand, the light beam that has passed through without being shielded by the mask means 3 is, as shown in FIGS. 2 (a) and 2 (b), the position of the pupil 10 of the objective lens 7 via the light beam incident position adjusting means 4. Is adjusted so that the focus detection light beam at is incident on a predetermined position in the pupil 10 corresponding to the diameter of the pupil 10 of the objective lens 7. The light whose position has been adjusted enters the polarization beam splitter 5. One linearly polarized light transmitted through the polarizing beam splitter 5 is converted into circularly polarized light by the λ / 4 plate 6 and illuminates the test object through the objective lens 7. Then, the light beam reflected by the test object passes through the objective lens 7, is converted into the other linearly polarized light by the λ / 4 plate 6, is reflected by the polarization beam splitter 5, passes through the imaging lens 8, and passes to the photodetector 9. Led.

  Here, in the focus detection apparatus of the first embodiment, the light beam incidence adjusting means 4 has the inclined surfaces 4a2 and 4b1 arranged in a state of facing each other, and the interval between the inclined surfaces 4a2 and 4b1 can be adjusted. The first prism 4a and the second prism 4b are configured, and the position of the pupil 10 of the objective lens 7 is adjusted by adjusting the interval between the inclined surfaces 4a2 and 4b1 of the first prism 4a and the second prism 4b. The incident position of the focus detection light beam at is adjustable to a predetermined position in the pupil 10 corresponding to the diameter of the pupil 10 of the objective lens 7.

  Therefore, by adjusting the distance between the inclined surfaces 4a2 and 4b1 of the first prism 4a and the second prism 4b, for example, in the vicinity of one surface of the first or second surface of the transparent substrate that is the test object When the focal point of the objective lens 7 is located, the reflected light from one surface is incident on the two light receiving portions in the photodetector 9 and the reflected light from the other surface is divided by the second virtual plane. Of the focus detection light beam in the pupil 10 at the position of the pupil 10 of the objective lens 7 at a predetermined position in the pupil 10 of the objective lens 7 so as to pass through a region outside the light receiving portion arranged in the region in one of the regions. The incident position can be adjusted.

  For this reason, according to the focus detection apparatus of the first embodiment, the laser beam diameter is optimized to match the pupil diameter of the objective lens without dimming the laser light even when switching to an objective lens having a different pupil diameter. In addition, when switching to an objective lens having a large pupil diameter, the influence of the reflected light from the surface that is not the object of focusing on the transparent substrate is eliminated as much as possible, and the object of focusing on the transparent substrate is It is possible to detect the amount of reflected light from the surface on the side with high detection sensitivity and perform focusing on the surface on the side to be focused.

Second Embodiment FIG. 7 is an explanatory view of a schematic configuration of a focus detection apparatus according to a second embodiment of the present invention as seen from the side, and (a) shows a light beam incident position when an objective lens having a large pupil diameter is used. The figure which shows the adjustment state of the light beam incident position by the adjusting means, (b) is the light reflected on the surface of the transparent substrate that is the object of focusing and the light reflected on the surface of the side that is not the object of focusing It is a figure which shows the positional relationship with the 2 division | segmentation photodetector in the state of (a) regarding the light which was made, (c) is light beam incidence by the light beam incident position adjustment means at the time of using an objective lens with a small pupil diameter It is a figure which shows the adjustment state of a position.
In the focus detection apparatus of the second embodiment, the light beam incident position adjusting means 4 is composed of two prisms 4a ′ and 4b ′ having the same apex angle, which are arranged upside down.
The first prism 4a ′ is configured with a flat surface 4a1 ′ on the incident side and an inclined surface 4a2 ′ on the output side.
The second prism 4b ′ is configured with an inclined surface 4b1 ′ parallel to the inclined surface 4a2 ′ on the incident side and a flat surface 4b2 ′ on the output side.
The first prism 4a ′ and the second prism 4b ′ are configured to shift the focus detection light beam in a direction perpendicular to the optical axis X1 by adjusting the interval between the inclined surfaces 4a2 ′ and 4b1 ′. Has been.
Then, the light beam incident position adjusting means 4 adjusts the distance between the first prism 4 a and the second prism 4 b, thereby determining the incident position of the focus detection light beam at the position of the pupil 10 of the objective lens 7. It can be adjusted to a predetermined position in the pupil 10 according to the diameter of the pupil 10.
Other configurations are substantially the same as those of the focus detection apparatus of the first embodiment.

In the focus detection apparatus of the second embodiment configured as described above, the focus detection light beam incident on the prism 4a ′ is shifted in the direction away from the optical axis X1 as the distance between the prism 4a ′ and the prism 4b ′ increases. As the distance between the prism 4a ′ and the prism 4b ′ decreases, the focus detection light beam incident on the prism 4a ′ shifts in a direction approaching the optical axis X1.
Therefore, when an objective lens having a large pupil diameter is used, as shown in FIG. 7A, the interval between the prism 4a ′ and the prism 4b ′ is widened, and the pupil 10 of the objective lens 7 for the focus detection light beam. The incident position at the position is adjusted to a predetermined position (a suitable position that does not cause noise) in the pupil 10 away from the optical axis X1.
At this time, as shown in FIG. 7 (b), the light reflected by the surface of the transparent substrate that is the object of focusing is condensed in a state almost close to a point and is incident on the two-divided photodetector. In addition, an equal amount of light is detected by both light receiving portions of the two-divided photodetector. On the other hand, the light reflected by the surface that is not the object of focusing is directed to the two-divided photodetector without being imaged. Of this light, the light in the region that can enter the two-divided photodetector is , All of which pass through a position off the two-divided photodetector.
On the other hand, when an objective lens having a small pupil diameter is used, as shown in FIG. 7C, the distance between the prism 4a ′ and the prism 4b ′ is reduced, and the pupil 10 of the objective lens 7 for the focus detection light beam. The incident position at the position is adjusted so as to enter the pupil 10.

Therefore, according to the focus detection device of the second embodiment, similarly to the focus detection device of the first embodiment, the laser beam diameter can be adjusted without dimming the laser light even when switching to an objective lens having a different pupil diameter. Can be optimized according to the pupil diameter of the objective lens, and even when switching to an objective lens with a large pupil diameter, the amount of reflected light from the surface on the focus target side is detected with high detection sensitivity. The focusing can be performed on the surface on the side to be focused.
In addition, according to the focus detection apparatus of the second embodiment, only one type of prism is required to form the incident light beam position adjusting means, and the shape is simple. Therefore, the configuration can be simplified and the cost can be reduced.

Third Embodiment FIG. 8A is an explanatory view showing a schematic configuration of a focus detection apparatus according to a third embodiment of the present invention, and FIG. 8B is reflected by the surface of the transparent substrate on the side to be focused. It is a figure which shows the positional relationship with the 2 division | segmentation photodetector which is a photodetector in a focus detection apparatus regarding the light reflected on the surface of the side which is not the object of focusing with the collected light.
In the focus detection apparatus according to the third embodiment, the light beam incident position adjusting means 4 includes a first conical lens having a plane 4a1 ″ on the incident side and an inclined surface 4a2 ″ that is recessed in a conical shape symmetrical on the output side with respect to the optical axis X1. 4 a ″, the incident side is configured by the inclined surface 4 b 1 ″ protruding in a conical shape symmetrical to the optical axis X 11, which coincides with the concave conical shape of the inclined surface 4 a 2 ″ of the first conical lens 4 a ″, and the output side is configured by the plane 4 b 2 ″. Second conical lens 4b ″.
The light beam incident position adjusting means 4 transforms the focus detection light beam into a concentric shape with the optical axis as the center, and increases the interval between the inclined surfaces 4a2 ″ and 4b1 ″ of the conical lenses 4a ″ and 4b ″. The deformed light flux is shifted in a direction away from the optical axis X1.
Other configurations are substantially the same as those of the focus detection apparatus of the first embodiment.
Therefore, as shown in FIG. 8 (b), the light reflected by the surface of the transparent substrate that is the object of focusing is condensed in a state that is almost close to a point and enters the two-divided photodetector. An equal amount of light is detected by both light receiving portions of the two-divided photodetector. On the other hand, the light reflected by the surface that is not the object of focusing is directed to the two-divided photodetector without being imaged. Of this light, the light in the region that can enter the two-divided photodetector is , All of which pass through a position off the two-divided photodetector.

According to the focus detection device of the third embodiment, as shown in FIG. 8, since the entire area of the outer periphery in the pupil 10 of the objective lens 7 can be used as an optical path through which the illumination light for detection passes, the sensitivity of automatic focus detection is good. Become.
Other functions and effects are substantially the same as those of the focus detection apparatus of the first embodiment.

In the focus detection apparatus of each of the above embodiments, when an objective lens having an extremely small pupil diameter is used, the focus detection light beam is incident on a position including substantially the entire range of the pupil 10 at the position of the pupil 10, and noise and The incident light also enters a region near the optical axis.
Therefore, also in the focus detection apparatus of each of the above embodiments, the mask means 66 (FIG. 10) shown in the focus detection apparatus of FIG. 9 can be switched to the mask means 3 together with the mask means 3, and the pupil diameter is small. When an objective lens is used, it is preferable to switch from the mask means 3 to the mask means 66. In this way, when an objective lens having a small pupil diameter is used, highly accurate focusing can be performed without adversely affecting the focusing signal.

In addition, in the focus detection apparatus of each embodiment described above, the prism is used as the two optical members constituting the light beam incident position adjusting unit 4, but a DOE may be used instead of the prism.
By doing so, the optical member constituting the light beam incident position adjusting means 4 can be reduced, and the arrangement space of the light beam incident position adjusting means 4 can be reduced.

  The present invention is useful, for example, in a field where it is required to obtain a statistical analysis result by acquiring images of a large number of cells using an automated microscope observation apparatus and analyzing the acquired images.

It is explanatory drawing which shows schematic structure of the focus detection apparatus concerning 1st embodiment of this invention. The state in which the illumination light beam not shielded by the mask means in the focus detection apparatus of FIG. 1 is adjusted to a predetermined position according to the pupil diameter of the objective lens through the light beam incident position adjustment means. It is explanatory drawing shown, (a) is the figure seen from upper direction, (b) is the figure seen from the side. It is a disassembled perspective view which shows the structure of the light beam incident position adjustment means used for the focus detection apparatus of FIG. It is principle explanatory drawing of light beam incident position adjustment by a light beam incident position adjustment means. It is explanatory drawing which shows an example of the light beam adjustment by a light beam incident position adjustment means. Incidence of light beam when the refractive index n of the prism constituting the light beam incident position adjusting means is n = 1.51 and a marginal light beam having a diameter of 2 mm is incident at a position 4 mm from the optical axis at the pupil position of the objective lens. It is a graph which shows the relationship between the space | interval between the inclined surfaces of the prism which comprises a position adjustment means, and the angle which an optical axis and the inclined surface of a prism make. FIG. 7 is an explanatory view of a schematic configuration of a focus detection apparatus according to a second embodiment of the present invention viewed from the side, (a) is a diagram showing a state in which the position of an incident light beam is shifted in parallel via a light beam incident position adjustment unit; (b) relates to light reflected from the surface of the transparent substrate that is the object of focusing and light reflected from the surface of the side that is not the object of focusing. FIG. 5C is a diagram showing a positional relationship with the split photodetector, and FIG. 5C is a diagram showing a state where the position of the incident light beam is not shifted via the light beam incident position adjusting means. (a) is an explanatory view showing a schematic configuration of the focus detection apparatus of the third embodiment of the present invention, (b) is a focus of the light reflected by the surface of the transparent substrate that is the focus target side It is a figure which shows the positional relationship with the 2 division | segmentation photodetector which is a photodetector in a focus detection apparatus regarding the light reflected by the surface of the side which is not object. It is explanatory drawing which shows schematic structure of the microscope observation apparatus provided with the focus detection apparatus concerning one Embodiment of invention described in Japanese Patent Application No. 2007-316857. The shape of the first and second light shielding parts constituting the mask means in the focus detection apparatus provided in the microscopic observation apparatus of FIG. 9, and the region not shielded by the first and second light shielding parts in the illumination light beam (for focus detection) It is explanatory drawing which shows the area | region shielded. With respect to the illumination light (for focus detection) in the area not shielded by the first and second light shielding portions shown in FIG. 10, the light reflected from the surface of the transparent substrate on the side to be focused, It is explanatory drawing which shows the positional relationship with the two light-receiving part of the light reflected by the surface of the side which is not object. In the focus detection apparatus of FIG. 9, when the focus is set on the surface of the transparent substrate to be tested, the light reflected by the surface on the focus target side is not the focus target. It is explanatory drawing which shows notionally the path | route of the light reflected on the surface of the side. In the focus detection apparatus in the state shown in FIG. 12, the incident states in the vicinity of the two light receiving portions of the light reflected by the respective surfaces of the transparent substrate on the side that is the focus detection target and the side that is not the focus detection target are shown. It is explanatory drawing shown. In the focus detection apparatus of FIG. 9, when the focus is set on the surface of the transparent substrate opposite to the test object side, the light reflected by the surface on the focus target side and the focus target It is explanatory drawing which shows notionally the path | route of the light reflected on the surface of the side which is not. In the focus detection apparatus in the state shown in FIG. 14, the incident states in the vicinity of the two light receiving portions of the light reflected by the respective surfaces of the transparent substrate on the side that is the focus detection target and the side that is not the focus detection target are shown. It is explanatory drawing shown. It is a graph which shows an example of the change of the light reception amount according to the focusing state detected by two light-receiving parts in the focus detection apparatus of FIG. It is a graph which shows an example of the evaluation function calculation value calculated based on the light-receiving amount detected by the two light-receiving parts shown in FIG.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Focus detection light source 2 Collimating lens 3 Mask means 4 Light beam incident position adjustment means 4a, 4a '1st prism 4a1, 4a1', 4a1 "plane 4a2 Inclined surface 4a2 'inclined to V shape symmetrical to an optical axis Surface 4a2 "Inclined surface 4b, 4b 'recessed in a conical shape symmetrical to the optical axis Second prism 4b1 Projected into a V shape symmetrical to the optical axis that matches the V shape of the inclined surface 4a2 of the first prism 4a Inclined surface 4b1 ′ Inclined surfaces 4b2 and 4b2 ′ parallel to the inclined surface 4a2 ′ 5 Polarizing beam splitter 6 λ / 4 plate 7 Objective lens 8 Imaging lens 9 Photo detectors 9a and 9b Light receiving unit 10 Entrance pupil 51 Microscope main body 52 Epi-illumination device 53 Focus detection device 53 ′ Automatic focus detection unit 54 Test object 54a Microplate 54a1 Bottom surface portion 54a11, 54a12 Bottom surface portion 4a1 Surface 54b Culture medium 55 XY stage 56 Objective lens 57, 70 Half mirror 58 Imaging lens 59 CCD camera 60 Driving unit 61 Light source 62, 63 Lens 64 Polarizing beam splitter 65 λ / 4 plate 66 Mask means 66a First light shielding unit 66b Second light shielding portion 67 Point light source 68 Disturbing light shielding member 69 Photo detectors 69a and 69b Light receiving portion S Transparent substrate having surface to be detected S1 Target surface

Claims (8)

In a focus detection device that performs focus adjustment by projecting a focus detection light beam onto a sample surface via an objective lens,
The optical system includes two optical members having inclined surfaces arranged in opposition to each other and capable of adjusting the interval between the inclined surfaces. By adjusting the interval between the inclined surfaces of the two optical members, the objective A focus detection apparatus comprising: a light beam incident position adjusting unit capable of adjusting an incident position of the focus detection light beam at a pupil position of a lens to a predetermined position in the pupil according to a pupil diameter of the objective lens.   The light beam incident position adjusting means separates the focus detection light beam symmetrically with respect to an optical axis, and adjusts an interval between the inclined surfaces of the two optical members, thereby adjusting an interval between the two separated light beams. The focus detection apparatus according to claim 1.   The two optical members are a first prism composed of an inclined surface recessed in a V-shape symmetric with respect to the optical axis on the incident side and an incident side, and a V-shape with an incident side inclined on the inclined surface of the first prism. 3. The focus detection apparatus according to claim 2, wherein the focus detection apparatus includes an inclined surface that projects in a V shape symmetrical to the optical axis, and a second prism having a flat exit surface.   The light beam incident position adjusting means shifts the focus detection light beam in a direction perpendicular to the optical axis by adjusting an interval between inclined surfaces of the two optical members. Focus detection device.   5. The focus detection apparatus according to claim 4, wherein the two optical members include two prisms having the same apex angle, which are arranged with their tops and bottoms reversed.   The light beam incident position adjusting means deforms the focus detection light beam into a concentric circle centered on the optical axis, and adjusts the interval between the inclined surfaces of the two optical members, thereby changing the deformed concentric light beam. The focus detection apparatus according to claim 1, wherein the diameter of the focus detection apparatus is adjusted.   The two optical members have a first conical lens configured with a concave surface that is concave in a conical shape symmetric with respect to the optical axis on the incident side, and an incident side is concave on the inclined surface of the first conical lens. 7. The focus detection apparatus according to claim 6, comprising an inclined surface that coincides with the conical shape and protrudes in a conical shape symmetrical to the optical axis, and a second conical lens having a flat exit surface. The focus detection device emits illumination light for generating an in-focus signal for the objective lens and the transparent substrate, and irradiates through the objective lens with the point light source and the illumination light of the illumination light. Mask means having a first light-shielding portion that shields a light beam passing through one region when divided by a first virtual plane along the optical axis, and a second along the optical axis of the light reflected by the transparent substrate A light detector having two light receiving portions arranged symmetrically across a virtual plane, and based on the amount of reflected light from the transparent substrate detected through the two light receiving portions, the transparent Focusing the objective lens on the first or second surface of the substrate;
The light beam incident position adjusting means is configured to receive reflected light from the first surface when the focal point of the objective lens is located in the vicinity of one of the first and second surfaces of the transparent substrate. In the pupil of the objective lens that passes through a region outside the light receiving unit disposed in the one region when the reflected light from the other surface is divided by the second virtual plane and incident on the other portion The focus detection apparatus according to claim 1, wherein an incident position of the light beam for focus detection at a pupil position of the objective lens can be adjusted to a predetermined position.

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