Microscope System

Abstract

Provided is a microscope system in which the time between the start of acquisition of an image and the completion of estimation of an object model is short.　A microscope system 1 comprises an incoherent light source 2, a detection optical system 3, and an imaging element 4. The incoherent light source 2 is a light source that emits temporally incoherent light. On a specimen, a plurality of coherent illuminations are performed simultaneously by the light emitted from the incoherent light source. Each coherent illumination is illumination by spatially coherent light, and the direction in which the specimen is irradiated with a light flux differs according to each coherent illumination. In a pupil plane of the detection optical system, the respective light fluxes of the coherent illuminations pass through first regions different from each other. Each of the first regions satisfies condition 1, and at least one interval among intervals between two adjacent first regions satisfies condition 2.　(1): LS < PS×10-3 (2): 0.05×T < d

Description

[Technical field]

[0001] The present invention relates to a microscope system. [Background technology]

[0002] There is a reconstruction method that reproduces an actual object with an object model on a computer. In this reconstruction method, the object model on the computer is modified using an optimization method so that the measured image of the object matches the image of the calculated object model. When the image of the object and the image of the object model finally match, the object model on the computer reproduces the actual object.

[0003] The image of the object is captured by the measurement optical system. The image of the object model is calculated by the image calculation method. Therefore, two important aspects of this reconstruction method are the measurement optical system and the image calculation method.

[0004] The measurement optical system may be, for example, the optical system of a microscope. In the optical system of a microscope, a halogen lamp or an LED is used to acquire an image of a specimen. The halogen lamp and the LED are incoherent light sources.

[0005] Illumination using an incoherent light source is classified into incoherent illumination, coherent illumination, and partial coherent illumination depending on the illumination conditions. These types of illumination will be described below.

[0006] In microscopes, Kohler illumination is used. In Kohler illumination, the light source is placed on the focal plane of a condenser lens, or an image of the light source is formed on the focal plane of a condenser lens. The light from each point of the light source is converted into parallel rays by the condenser lens. Thus, the specimen is illuminated with a parallel beam of light.

[0007] Changing the size of the light source changes the spatial coherence of the illumination light at the specimen plane, which in turn changes the imaging properties.

[0008] Even if the light source is an incoherent light source, if the size of the light source is made very small, the light source can be considered as a point light source. Illumination in which light from a point light source is irradiated onto a specimen is called coherent illumination.

[0009] Image processing methods can be classified into linear and nonlinear. Linear processing considers only a single scattering in the sample. Nonlinear processing considers not only a single scattering but also multiple scattering.

[0010] Linear calculations use the first-order Born approximation. In the Born approximation, even if scattering occurs more than once in the sample, the scattering is ignored. In linear calculations, the sample information and the output information are determined in a one-to-one relationship. Therefore, the output information can be calculated analytically. The output information is, for example, an image of the sample.

[0011] Linear operations will be explained using the example of imaging in a microscope. When the specimen information (transmittance distribution of the specimen) O and the output information (image intensity distribution) I can be regarded as a linear system of the convolution of the point spread function PSF of the optical system, the output information I is expressed by the following formula. I=PSF*O Here, * denotes convolution.

[0012] In linear calculation, the calculation time is short, but the calculation accuracy is low because scattering more than twice is ignored. The image of the object model reconstructed using linear calculation is obtained by deconvolving the image of the measured sample with the point spread function.

[0013] Nonlinear calculation is a calculation method that takes into account the occurrence of multiple scattering in the sample. One of the nonlinear calculations is the beam propagation method. In the beam propagation method, the object model is replaced with multiple thin layers. Then, the wavefront change as the light passes through each layer is calculated sequentially to calculate the image of the object model.

[0014] The beam propagation method can calculate the image of an object model more accurately than linear calculations.

[0015] A method for restoring the refractive index distribution of a specimen by optimization calculation is disclosed in Non-Patent Document 1. This method uses the beam propagation method. In addition, spatially coherent illumination is used to acquire the image.

[0016] By estimating the object model from the images, it is possible to estimate the 3D optical properties, e.g. the refractive index distribution of the object. [Prior art documents] [Non-patent literature]

[0017] [Non-Patent Document 1] "High-resolution 3D refractive index microscopy of multiple-scattering samples from intensity images" Optica, Vol. 6, No. 9, pp.1211-1219(2019) Summary of the Invention [Problem to be solved by the invention]

[0018] In Non-Patent Document 1, illumination is performed with a single light beam. In this case, in order to illuminate the specimen from multiple directions, it is necessary to change the illumination angle. Therefore, it takes time to capture an image. As a result, it takes a long time from the start of image capture to the completion of object model estimation.

[0019] The present invention has been made in consideration of the above problems, and has an object to provide a microscope system in which the time from the start of image acquisition to the completion of object model estimation is short. [Means for solving the problem]

[0020] In order to solve the above-mentioned problems and achieve the object, a microscope system according to at least some embodiments of the present invention includes: The system includes an incoherent light source, a detection optical system, and an image sensor, An incoherent light source is a light source that emits light that is not coherent in time. The detection optical system is an optical system that forms an optical image of the sample, The imaging element receives an optical image of the specimen formed by the detection optical system, In the specimen, multiple coherent illuminations are performed simultaneously using light emitted from incoherent sources. Coherent illumination is illumination with light that is spatially coherent. The direction in which the light beam is irradiated onto the specimen varies with each type of coherent illumination. At the pupil plane of the detection optical system, the light beams of the coherent illumination pass through different first regions, Each of the first regions satisfies the following condition (1): At least one of the intervals between two adjacent first regions is characterized in that it satisfies the following condition (2). LS <PS×10 -3 (1) 0.05×T <d (2) Where: LS is the area of ​​the first region (unit: mm 2 ), PS is the pupil area of ​​the detection optics (unit: mm 2 ), d is the distance between two adjacent first regions (unit: mm), T is the pupil diameter of the detection optics (unit: mm), It is.

[0021] In accordance with at least some embodiments of the present invention, a microscope system includes: The system includes an incoherent light source, an illumination optical system, a detection optical system, and an image sensor, An incoherent light source is a light source that emits light that is not coherent in time. The detection optical system is an optical system that forms an optical image of the sample, The imaging element receives an optical image of the specimen formed by the detection optical system, In the specimen, multiple coherent illuminations are performed simultaneously using light emitted from incoherent sources. Coherent illumination is illumination with light that is spatially coherent. The direction in which the light beam is irradiated onto the specimen varies with each type of coherent illumination. On a pupil plane of the illumination optical system, the light beams of the coherent illuminations are located in different second regions, Each of the second regions satisfies the following condition (6): At least one of the intervals between two adjacent second regions is characterized in that it satisfies the following condition (7). LS' <PS’×10 -3 (6) 0.05×T' <d’ (7) Where: LS' is the area of ​​the second region (unit: mm 2 ), PS' is the pupil area of ​​the illumination optical system (unit: mm 2 ), d' is the distance between two adjacent second regions (unit: mm), T' is the pupil diameter of the illumination optical system (unit: mm), It is. Effect of the Invention

[0022] According to the present invention, it is possible to provide a microscope system in which the time from the start of image acquisition to the completion of object model estimation is short. [Brief description of the drawings]

[0023] [Figure 1] 1A and 1B are diagrams illustrating a microscope system according to the present embodiment and a captured image. [Diagram 2] FIG. 2 is a diagram showing a pupil plane of a detection optical system. [Diagram 3] FIG. 2 is a diagram illustrating a pupil plane of a first example of a detection optical system. [Figure 4] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in a second example. [Diagram 5] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in a third example. [Figure 6] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in a fourth example. [Figure 7] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in a fifth example. [Figure 8] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in a sixth example. [Figure 9] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in a seventh example. [Figure 10] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in an eighth example. [Figure 11] FIG. 13 is a diagram illustrating a pupil plane of a detection optical system in a ninth example. [Figure 12] FIG. 1 is a diagram illustrating a microscope system according to an embodiment of the present invention. [Figure 13] FIG. 2 is a diagram showing a pupil plane of an illumination optical system. [Figure 14] FIG. 1 is a diagram illustrating a microscope system according to an embodiment of the present invention. [Figure 15] FIG. [Figure 16] FIG. 1 is a diagram illustrating a microscope system according to an embodiment of the present invention. [Figure 17] 1 is a flowchart of a first simulation. [Figure 18] FIG. 1 is a diagram showing an optical system used in a simulation. [Figure 19] FIG. 13 shows an image of an estimated specimen. [Figure 20] FIG. 13 is a diagram illustrating correction of a wavefront. [Figure 21] FIG. 1 shows the gradient of a specimen. [Figure 22] FIG. 1 shows the gradient of a specimen. [Diagram 23] FIG. 1 is a diagram illustrating a microscope system according to an embodiment of the present invention. [Figure 24] FIG. [Diagram 25] 13 is a flowchart of a second simulation. [Figure 26] 13 is a flowchart of a second simulation. [Figure 27] FIG. 1 is a diagram showing an optical system used in a simulation. [Figure 28]FIG. 2 is a diagram showing wavefronts in each layer. [Figure 29] 2A and 2B are diagrams illustrating a wavefront at a position where a photographed image is acquired and a wavefront at an imaging plane. [Diagram 30] FIG. 13 shows an image of an estimated specimen. [Diagram 31] FIG. 13 is a diagram illustrating correction of a wavefront. [Diagram 32] FIG. 1 illustrates the gradient of a sample and the propagation of a wavefront. [Diagram 33] FIG. 1 illustrates the gradient of a sample and the propagation of a wavefront. [Diagram 34] FIG. 1 shows the gradient of a specimen. [Diagram 35] FIG. 1 shows the gradient of a specimen. [Diagram 36] FIG. 11 is a diagram showing the results of a simulation of the first example. [Figure 37] FIG. 1 shows an aperture material and a reconstructed putative specimen. [Figure 38] FIG. 13 is a diagram showing images of an aperture member and a specimen in the measurement of the second example. [Figure 39] FIG. 13 is a diagram showing an estimated specimen of the second example. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0024] Prior to the description of the examples, the effects of the embodiments according to certain aspects of the present invention will be described. In addition, when specifically describing the effects of the present embodiment, specific examples will be shown. However, as in the case of the examples described later, the exemplified aspects are merely a part of the aspects included in the present invention, and there are many variations in the aspects. Therefore, the present invention is not limited to the exemplified aspects.

[0025] In the following description, a specimen having a thin thickness will be referred to as a "thin specimen", and a specimen having a thick thickness will be referred to as a "thick specimen".

[0026] The microscope system of this embodiment includes an incoherent light source, a detection optical system, and an image sensor. The incoherent light source is a light source that emits light that is not coherent in time. The detection optical system is an optical system that forms an optical image of a specimen, and the image sensor receives the optical image of the specimen formed by the detection optical system. A plurality of coherent illuminations are simultaneously performed on the specimen by light emitted from the incoherent light source, and the coherent illumination is illumination by light that is spatially coherent. The direction in which the light beam is irradiated onto the specimen differs for each of the coherent illuminations. On the pupil plane of the detection optical system, the respective light beams of the coherent illuminations pass through different first regions. The first regions are regions of light beams that pass through the pupil plane of the detection optical system, and each of the first regions satisfies the following condition (1), and at least one of the intervals between two adjacent first regions satisfies the following condition (2). LS <PS×10 -3 (1) 0.05×T <d (2) Where: LS is the area of ​​the first region (unit: mm 2 ), PS is the pupil area of ​​the detection optics (unit: mm 2 ), d is the distance between two adjacent first regions (unit: mm), T is the pupil diameter of the detection optics (unit: mm), It is.

[0027] 1A and 1B are diagrams showing a microscope system and a captured image according to this embodiment, respectively. Fig. 1(a) is a diagram showing the microscope system, and Fig. 1(b) is a diagram showing the captured image.

[0028] As shown in FIG. 1( a ), a microscope system 1 includes an incoherent light source 2 , a detection optical system 3 , and an image sensor 4 .

[0029] The incoherent light source 2 is a light source that emits light that is not coherent in time. Coherent illumination is performed on the specimen 5 by the light emitted from the incoherent light source. Coherent illumination is illumination by light that is spatially coherent.

[0030] In coherent illumination of the specimen 5, a plurality of light beams are simultaneously irradiated onto the specimen 5. Furthermore, each of the plurality of light beams is irradiated onto the specimen 5 from a different direction.

[0031] FIG. 1(a) shows light beams L1 and L2. Light beams L1 and L2 are light beams that are irradiated onto a specimen 5 and are emitted from an incoherent light source 2. Light beams L1 and L2 are independent of each other and are irradiated onto the specimen 5 simultaneously. The direction of light beam L1 relative to the specimen 5 is different from the direction of light beam L2 relative to the specimen 5. Light beams L1 and L2 are each a light beam of coherent illumination light.

[0032] The specimen 5 is a thin specimen. In FIG. 1, the focal position Fo of the detection optical system 3 is located inside the specimen 5.

[0033] The detection optical system 3 is an optical system that forms an optical image 5' of the specimen 5. Light emitted from the specimen 5 is condensed onto an image-forming plane IP by the detection optical system 3. An optical image 5' is formed on the image-forming plane IP.

[0034] The imaging surface of the image sensor 4 is located on the image plane IP. The image sensor 4 receives an optical image 5' of the specimen 5 formed by the detection optical system 3. The image sensor 4 captures the optical image 5'. As a result, a captured image I shown in FIG. 1(b) is obtained. mea (r) is obtained, where r indicates the two-dimensional coordinate (x, y).

[0035] Since the specimen 5 is a thin specimen, one photographed image is acquired. Therefore, the detection optical system 3 and the image sensor 4 do not move in the optical axis direction. Moreover, the specimen 5 does not move in the optical axis direction either.

[0036] The light beam emitted from the specimen 5 reaches the pupil position Pu of the detection optical system 3. A plurality of light beams are simultaneously irradiated onto the specimen 5. Thus, a plurality of light beams simultaneously reach the pupil position Pu. Furthermore, each of the plurality of light beams is irradiated onto the specimen 5 from a different direction. Thus, at the pupil position Pu, the plurality of light beams pass through different regions.

[0037] 2 is a diagram showing a pupil plane of the detection optical system. The first region is a region of a light beam passing through the pupil plane of the detection optical system.

[0038] 2, the number of first regions on the pupil plane 10 of the detection optical system 3 is eight. For example, the first region 11 is a region through which the light beam L1 passes through the pupil plane 10. The first region 12 is a region through which the light beam L2 passes through the pupil plane 10.

[0039] In the microscope system of this embodiment, each of the first areas satisfies the following condition (1). LS <PS×10 -3 (1) Where: LS is the area of ​​the first region (unit: mm 2 ), PS is the pupil area of ​​the detection optics (unit: mm 2 ), It is.

[0040] Each of the multiple light beams is an independent light beam. Therefore, the light beams are emitted from different positions in the light source. The light beams emitted from the light source are irradiated onto a specimen. In this case, it is preferable that the light beams emitted from one emission position are irradiated onto the specimen with one wavefront.

[0041] In the microscope system of this embodiment, the captured image I mea (r) can be obtained. mea (r) can be used, for example, to estimate the refractive index distribution, which will be described later.

[0042] In the estimation of the refractive index distribution, a calculated image is calculated from the estimated specimen using the beam propagation method for partially coherent imaging. The calculated image is the captured image I mea The parameters of the estimated sample (refractive index distribution) are changed using the gradient descent method or other methods so as to approach (r).

[0043] In the beam propagation method for partially coherent imaging, the calculation time is proportional to the number of wavefronts irradiated onto the specimen. If the upper limit of condition (1) is exceeded, multiple wavefronts will be irradiated onto the specimen by the light beam emitted from one emission position. Therefore, the calculation time becomes too long.

[0044] In the microscope system of this embodiment, at least one of the intervals between two adjacent first regions satisfies the following condition (2). 0.05×T <d (2) Where: d is the distance between two adjacent first regions (unit: mm), T is the pupil diameter of the detection optics (unit: mm), It is.

[0045] An optical image is formed by the light beam in one first region, and an optical image is formed by the light beam in the other first region. Because the angles of incidence on the specimen are different for the two light beams, the two optical images are also different.

[0046] If the lower limit of condition (2) is exceeded, one first region and the other first region become too close. In this case, the difference between the two optical images becomes small. When the difference becomes small, the information obtained from the two optical images becomes almost the same. As a result, for example, in estimating the refractive index distribution, the reconstruction performance does not improve despite the long calculation time.

[0047] In the microscope system of this embodiment, it is preferable that half of the first regions satisfy the condition (2).

[0048] When half of the first regions satisfy the condition (2), different pieces of information can be obtained from two adjacent optical images, and a large amount of different pieces of information can be obtained. When estimating the refractive index distribution, it can be estimated with relatively high accuracy.

[0049] In the microscope system of this embodiment, it is preferable to satisfy the following condition (3).

number

[0050] The left side of condition (3) is the total area obtained by adding up the areas of the first regions. By satisfying condition (3), the number of wavefronts irradiated onto the sample can be reduced, and the calculation time can be shortened.

[0051] In the microscope system of this embodiment, some of the first regions are located within a first annular region, and the first annular region is preferably a region with a radius of 50% or more of the pupil region of the detection optical system.

[0052] Fig. 3 is a diagram showing the pupil plane of the detection optical system of the first example. Fig. 3 shows a first region on the pupil plane 10 of the detection optical system. The first region 20 and the first region 21 are located on a circumference. The first region 20 is located outside the circumference on which the first region 21 is located. It is sufficient if either the first region 20 or the first region 21 is present.

[0053] 3, one circle is drawn with a dashed line. The radius of the circle is 50% of the radius of the pupil 10. The area outside the circle is the first annular area 30. The first annular area 30 is an area whose radius is 50% or more.

[0054] In the first example, the first region 20 and the first region 21 are located in the first annular region 30. The first example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the first example makes it possible to estimate a relatively fine structure.

[0055] The calculation time will be shorter if there are many first regions that satisfy condition (2). The calculation time will be shorter even if some of the first regions do not satisfy condition (2). It is desirable for more than half of the first regions to satisfy condition (2).

[0056] In the microscope system of this embodiment, it is preferable that some of the first regions are arranged so as to form double circles within the first annular region.

[0057] 3, in the first example, the first region 20 and the first region 21 form a double circle in the first annular region 30. Therefore, a large amount of mutually different information can be acquired. When estimating the refractive index distribution, a relatively fine structure can be estimated.

[0058] In the microscope system of this embodiment, some of the first regions are located within the second annular region, and the second annular region is preferably an area having a radius of 70% to 90% of the pupil region of the detection optical system.

[0059] Fig. 4 is a diagram showing a pupil plane of the detection optical system of the second example. Fig. 4 shows a first region on the pupil plane 10 of the detection optical system. The first region 20 and the first region 21 are located on a circumference. The first region 20 is located outside the circumference on which the first region 21 is located. It is possible that only the first region 20 is present and the first region 21 is not required.

[0060] In Fig. 4, two circles are drawn with dashed lines. The radius of the inner circle is 70% of the radius of the pupil 10. The radius of the outer circle is 90% of the radius of the pupil 10. The region between the two circles is the second annular region 31. The second annular region 31 is the region whose radius is from 70% to 90%.

[0061] In the second example, the first region 20 is located in the second annular region 31. The second example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the second example makes it possible to estimate a relatively fine structure. It is not necessary for some of the first regions to satisfy condition (2). It is desirable for more than half of the first regions to satisfy condition (2).

[0062] In the microscope system of this embodiment, some of the first regions are located within a third annular region, and the third annular region is preferably an area having a radius of 50% to 70% of the pupil region of the detection optical system.

[0063] Fig. 5 is a diagram showing a pupil plane of a detection optical system of a third example. Fig. 5 shows a first region on a pupil plane 10 of the detection optical system. The first region 20 and the first region 21 are located on a circumference. The first region 20 is located outside the circumference on which the first region 21 is located. Only the first region 21 is present, and the first region 20 may not be present.

[0064] 5, two circles are drawn with dashed lines. The radius of the inner circle is 50% of the radius of the pupil 10. The radius of the outer circle is 70% of the radius of the pupil 10. The region between the two circles is the third annular region 32. The third annular region 32 is the region whose radius is from 50% to 70%.

[0065] In the third example, the first region 21 is located in the third annular region 32. The third example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the third example makes it possible to estimate a relatively fine structure. It is not necessary for some of the first regions to satisfy condition (2). It is desirable for more than half of the first regions to satisfy condition (2).

[0066] In the microscope system of this embodiment, some of the first regions are located within the first circular region, and the first circular region is preferably a region of the pupil region of the detection optical system that is closer to the center than the first annular region.

[0067] Fig. 6 is a diagram showing a pupil plane of a detection optical system of a fourth example. Fig. 6 shows a first region on a pupil plane 10 of the detection optical system. The first region 20 and the first region 40 are located on a circumference. The first region 20 is located outside the circumference on which the first region 40 is located.

[0068] 6, one circle is drawn with a dashed line. The area outside the circle is the first circular area 30. The area inside the circle is the first circular area 50. The first circular area 50 is an area closer to the center than the first circular area.

[0069] In the fourth example, the first region 40 is located in the first circular region 50. The fourth example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the fourth example can estimate not only a relatively fine structure but also a relatively coarse structure. Some of the first regions do not need to satisfy condition (2). It is desirable that more than half of the first regions satisfy condition (2).

[0070] In the microscope system of this embodiment, it is preferable that some of the first regions are arranged so as to form a circle within the first circular region.

[0071] 6, in the fourth example, a circle is formed in the first circular region 50 by the first region 40. When estimating the refractive index distribution, a relatively coarse structure can be estimated.

[0072] In the microscope system of this embodiment, some of the first regions are located within the second circular region, and the second circular region is preferably a region having a radius of 50% or less of the pupil region of the detection optical system.

[0073] Fig. 7 is a diagram showing the pupil plane of the detection optical system of the fifth example. Fig. 7 shows the first region on the pupil plane 10 of the detection optical system. The first region 40 and the first region 41 are located on a circumference. The first region 41 is located outside the circumference on which the first region 40 is located. It is sufficient to have either the first region 40 or the first region 41.

[0074] 7, one circle is drawn with a dashed line. The radius of the circle is 50% of the radius of the pupil 10. The region inside the circle is a second circular region 51. The second circular region 51 is a region whose radius is 50% or less.

[0075] In the fifth example, the first region 40 and the first region 41 are located in the second circular region 51. The fifth example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the fifth example can estimate a relatively coarse structure. It is not necessary for some of the first regions to satisfy condition (2). It is desirable for more than half of the first regions to satisfy condition (2).

[0076] In the microscope system of this embodiment, it is preferable that some of the first regions are arranged so as to form a circle within the second circular region.

[0077] 7, in the fifth example, the first region 40 and the first region 41 form a circle in the second circular region 51. Therefore, a large amount of mutually different information can be acquired. When estimating the refractive index distribution, a relatively coarse structure can be estimated.

[0078] In the microscope system of this embodiment, some of the first regions are located within a fourth annular region, and the fourth annular region is preferably a region with a radius of 30% to 50% of the pupil region of the detection optical system.

[0079] Fig. 8 is a diagram showing a pupil plane of a detection optical system of a sixth example. Fig. 8 shows a first region on the pupil plane 10 of the detection optical system. The first region 40 and the first region 41 are located on a circumference. The first region 41 is located outside the circumference on which the first region 40 is located. It is possible that only the first region 41 is present and the first region 40 is not required.

[0080] 8, two circles are drawn with dashed lines. The radius of the inner circle is 30% of the radius of the pupil 10. The radius of the outer circle is 50% of the radius of the pupil 10. The region between the two circles is the fourth annular region 52. The fourth annular region 52 is a region whose radius is from 30% to 50%.

[0081] In the sixth example, the first region 41 is located in the fourth annular region 52. The sixth example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the sixth example makes it possible to estimate a relatively coarse structure. It is not necessary for some of the first regions to satisfy condition (2). It is desirable for more than half of the first regions to satisfy condition (2).

[0082] In the microscope system of this embodiment, some of the first regions are located within a third circular region, and the third circular region is preferably a region having a radius of 30% or less of the pupil region of the detection optical system.

[0083] Fig. 9 is a diagram showing a pupil plane of a detection optical system of a seventh example. Fig. 9 shows a first region on the pupil plane 10 of the detection optical system. The first region 40 and the first region 41 are located on a circumference. The first region 41 is located outside the circumference on which the first region 40 is located. It is possible that only the first region 40 is present and the first region 41 is not required.

[0084] 9, one circle is drawn by a dashed line. The radius of the circle is 30% of the radius of the pupil 10. The region inside the circle is a third circular region 53. The third circular region 53 is a region whose radius is 30% or less.

[0085] In the seventh example, the first region 40 is located in the third circular region 53. The seventh example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the seventh example can estimate a relatively coarse structure. It is not necessary for some of the first regions to satisfy condition (2). It is desirable for more than half of the first regions to satisfy condition (2).

[0086] In the microscope system of this embodiment, when the pupil of the detection optical system is divided into four sectors with equal central angles, it is preferable that any of the first regions is located in all four sectors.

[0087] Fig. 10 is a diagram showing the pupil plane of the detection optical system of Example 8. Fig. 10 shows first regions on the pupil plane 10 of the detection optical system. The first regions 60 are positioned in a grid pattern.

[0088] 10, two perpendicular straight lines are drawn. The two straight lines divide the pupil plane 10 into a first sector region 70, a second sector region 71, a third sector region 72, and a fourth sector region 74. Of the four sector regions, the central angle in one sector region is equal to the central angle in the other sector regions.

[0089] In the eighth example, the first region 60 is located in all of the first sector region 70, the second sector region 71, the third sector region 72, and the fourth sector region 74. The eighth example satisfies conditions (1), (2), and (3). When estimating the refractive index distribution, the eighth example can estimate not only a relatively fine structure but also a relatively coarse structure. Some of the first regions do not need to satisfy condition (2). It is desirable that more than half of the first regions satisfy condition (2).

[0090] In the microscope system of this embodiment, some of the first regions are preferably paired across the center of the pupil of the detection optical system.

[0091] Fig. 11 is a diagram showing the pupil plane of the detection optical system of the ninth example. Fig. 11 shows a first region on the pupil plane 10 of the detection optical system. The first region 80 and the first region 81 are located on a circumference.

[0092] In the ninth example, first region 80 and first region 81 form a pair on either side of the center C of the pupil of the detection optical system. In the ninth example, conditions (1), (2), and (3) are satisfied. When estimating the refractive index distribution, the ninth example makes it possible to estimate a relatively coarse structure. It is not necessary for some of the first regions to satisfy condition (2). It is desirable for more than half of the first regions to satisfy condition (2).

[0093] Moreover, in the ninth example, the number of first regions is smaller than in the seventh example. When estimating the refractive index distribution, it can be estimated in a shorter time than the estimation time in the seventh example.

[0094] In the microscope system of this embodiment, it is preferable that each of the first areas satisfies the following condition (4). PS x 10 -6 <LS (4) Where: LS is the area of ​​the first region (unit: mm 2 ), PS is the pupil area of ​​the detection optics (unit: mm 2 ), It is.

[0095] If the lower limit of condition (4) is exceeded, the optical image becomes dark. In this case, the captured image I mea The signal-to-noise ratio (SN) in (r) deteriorates. mea (r) can be used to estimate the refractive index distribution, for example. mea As the signal-to-noise ratio (r) deteriorates, the estimation accuracy deteriorates.

[0096] In the microscope system of this embodiment, it is preferable that at least one of the intervals between two adjacent first regions satisfies the following condition (5). d<0.5×T (5) Where: d is the distance between two adjacent first regions (unit: mm), T is the pupil diameter of the detection optics (unit: mm) It is.

[0097] If the upper limit of the condition (5) is exceeded, the number of first regions will be reduced, resulting in a dark optical image.

[0098] It is desirable that the first region that satisfies condition (2) also satisfies condition (5).

[0099] It is preferable that the microscope system of this embodiment satisfies the following condition (A). 4≦n≦100 (A) Where: n is the number of first regions, It is.

[0100] If the lower limit of condition (A) is exceeded, the number of first regions is reduced, resulting in a dark optical image. If the upper limit of condition (A) is exceeded, for example, the time required to estimate the refractive index distribution is increased.

[0101] The microscope system of this embodiment includes an incoherent light source, an illumination optical system, a detection optical system, and an image sensor. The incoherent light source is a light source that emits light that is not coherent in time. The detection optical system is an optical system that forms an optical image of the specimen, and the image sensor receives the optical image of the specimen formed by the detection optical system. A plurality of coherent illuminations are simultaneously performed on the specimen by light emitted from the incoherent light source, and the coherent illumination is illumination by light that is spatially coherent. The direction in which the light beam is irradiated onto the specimen differs for each of the coherent illuminations, and on the pupil plane of the illumination optical system, the light beams of each of the coherent illuminations are located in different second regions. Each of the second regions satisfies the following condition (6), and at least one of the intervals between two adjacent second regions satisfies the following condition (7). LS' <PS’×10 -3 (6) 0.05×T' <d’ (7) Where: LS' is the area of ​​the second region (unit: mm 2 ), PS' is the pupil area of ​​the illumination optical system (unit: mm 2 ), d' is the distance between two adjacent second regions (unit: mm), T' is the pupil diameter of the illumination optical system (unit: mm), It is.

[0102] Fig. 12 is a diagram showing the microscope system of this embodiment. The same components as those in Fig. 1 are given the same reference numbers and will not be described.

[0103] The microscope system 90 includes an incoherent light source 2 , an illumination optical system 91 , a detection optical system 3 , and an image sensor 4 .

[0104] When the incoherent light source 2 is placed at a position away from the pupil plane PI of the illumination optical system 91, the light emitted from the incoherent light source 2 passes through the pupil plane PI. When the incoherent light source 2 is placed at the position of the pupil plane PI, the light is emitted from the pupil plane PI. The light emitted from the incoherent light source 2 passes through the illumination optical system 91 and is irradiated onto the sample 5.

[0105] Fig. 13 is a diagram showing the pupil plane of the illumination optical system. In Fig. 13, the number of second regions on the pupil plane 100 of the illumination optical system 91 is eight. On the pupil plane 100, the multiple lights are located in different regions. The positions of the second regions are different from each other. The second regions indicate the positions of the multiple lights.

[0106] For example, the second region 101 is a region where the light flux L1 passes through the pupil plane 100, or a region where the light flux L1 is generated. The second region 102 is a region where the light flux L2 passes through the pupil plane 100, or a region where the light flux L2 is generated. The light flux L1 and the light flux L2 are each a light flux of coherent illumination light.

[0107] In the microscope system of this embodiment, each of the second areas satisfies the following condition (6). LS' <PS’×10 -3 (6) Where: LS' is the area of ​​the second region (unit: mm 2 ), PS' is the pupil area of ​​the illumination optical system (unit: mm 2 ), It is.

[0108] The technical meaning of condition (6) is the same as that of condition (1).

[0109] In the microscope system of this embodiment, at least one of the intervals between two adjacent second regions satisfies the following condition (7). 0.05×T' <d’ (7) Where: d' is the distance between two adjacent second regions (unit: mm), T' is the pupil diameter of the illumination optical system (unit: mm), l It is.

[0110] The technical meaning of condition (7) is the same as the technical meaning of condition (2).

[0111] In the microscope system of this embodiment, it is preferable that the detection optical system includes an objective lens and an imaging lens, the illumination optical system includes a condenser lens, the area of ​​the second region is expressed by the following equation (8), and the diameter of the pupil of the illumination optical system is expressed by the following equation (9). PS' = (FLcd × NA) 2 ×π (8) T' = FLcd × NA (9) Where: FLcd is the focal length of the condenser lens (unit: mm), NA is the numerical aperture of the objective lens, It is.

[0112] It is preferable that the microscope system of this embodiment further comprises an aperture member. Each of the multiple light beams is emitted from each of multiple independent regions on a predetermined surface. The predetermined surface is a surface that is perpendicular to the optical axis of the detection optical system and is located on the opposite side of the detection optical system with respect to the specimen. The aperture member is disposed on the predetermined surface and comprises multiple independent transmission regions. The transmission regions are regions that transmit light, and each of the transmission regions corresponds to each of the second regions.

[0113] Fig. 14 is a diagram showing a microscope system according to this embodiment. The same components as those in Fig. 12 are given the same reference numbers and will not be described.

[0114] The microscope system 110 includes an incoherent light source 2 , an illumination optical system 91 , an aperture member 111 , a detection optical system 3 , and an image sensor 4 .

[0115] In the microscope system 110, each of the multiple light beams is emitted from each of multiple regions on a predetermined plane. Each of the multiple regions is independent of each other. The predetermined plane is a plane that is perpendicular to the optical axis of the detection optical system 3 and is located on the opposite side of the detection optical system 3 with respect to the specimen 5. In the microscope system 110, the pupil plane PI of the illumination optical system 91 is the predetermined plane.

[0116] The aperture member 111 is disposed on a predetermined plane. In the microscope system 110, the aperture member 111 is disposed on a pupil plane PI.

[0117] The microscope system 110 may include a processor 112. By including the processor 112, for example, the refractive index distribution of the sample 5 can be estimated.

[0118] 15 is a diagram showing an aperture member. The aperture member 111 has a plurality of independent transmission regions 112. The plurality of transmission regions 112 are independent from each other. The transmission regions 112 are surrounded by a light blocking region 113.

[0119] The transmissive region 112 is a region that transmits light. When light passes through the transmissive region 112, a light beam L1 is irradiated onto the sample 5. When light passes through another transmissive region 112, a light beam L2 is irradiated onto the sample 5.

[0120] The transmission regions 112 are regions of light at the pupil plane PI. Each of the transmission regions 112 corresponds to each of the second regions.

[0121] In the aperture member 111, the transmission regions 112 are located on the circumference of four circles. The four circles are referred to as a first circle, a second circle, a third circle, and a fourth circle from the outer edge toward the center of the aperture member.

[0122] A transmitting region 112a is located on the circumference of a first circle. A transmitting region 112b is located on the circumference of a second circle. A transmitting region 112c is located on the circumference of a third circle. A transmitting region 112d is located on the circumference of a fourth circle. When the refractive index distribution is estimated using the aperture member 111, the estimation can be performed with high accuracy.

[0123] A transmissive region is provided in the center of the aperture member 111. Providing a transmissive region in the center can facilitate positioning of the aperture member 111. However, the transmissive region in the center is not necessarily required.

[0124] A specific example of an aperture member will now be described. The second region is located on a pupil plane PI, and the first region is located on a pupil plane Pu. The pupil plane PI can be conjugate with the pupil plane Pu. In this case, the second region is conjugate with the first region. Since the second region is a transmissive region of the aperture member, the first region can be regarded as a transmissive region of the aperture member. As a result, FIGS. 3 to 11 can be regarded as showing specific examples of aperture members.

[0125] Fig. 3 shows a first example of an aperture member. In Fig. 3, first region 20 and first region 21 represent the transmission regions of the aperture member. Some of the aperture members are located in a region with a radius of 50% or more of the pupil region of the illumination optical system. Some of the transmission regions of the aperture member are arranged to form double circles within the region with a radius of 50% or more.

[0126] The first example satisfies conditions (6) and (7). When estimating the refractive index distribution, the first example makes it possible to estimate a relatively fine structure.

[0127] A second example of the aperture member is shown in Fig. 4. In Fig. 4, some of the transmission regions of the aperture member are located in a region with a radius of 70% to 90% of the pupil region of the illumination optical system.

[0128] The second example satisfies conditions (6) and (7). When estimating the refractive index distribution, the second example makes it possible to estimate a relatively fine structure.

[0129] A third example of an aperture member is shown in Fig. 5. In Fig. 5, some of the transmission regions of the aperture member are located in a region with a radius of 50% to 70% of the pupil region of the illumination optical system.

[0130] The third example satisfies conditions (6) and (7). When estimating the refractive index distribution, the third example makes it possible to estimate a relatively fine structure.

[0131] Fig. 6 shows a fourth example of the aperture member. In Fig. 6, some of the transmission regions of the aperture member are located in a region on the central side of the pupil region of the illumination optical system with respect to a region having a radius of 50% or more.

[0132] The fourth example satisfies the conditions (6) and (7). When estimating the refractive index distribution, in the fourth example, not only a relatively fine structure but also a relatively coarse structure can be estimated.

[0133] Fig. 7 shows a fifth example of the aperture member. In Fig. 7, some of the transmission regions of the aperture member are located in a region with a radius of 50% or less of the pupil region of the illumination optical system.

[0134] The fifth example satisfies conditions (6) and (7). When estimating the refractive index distribution, the fifth example can estimate a relatively coarse structure.

[0135] Fig. 8 shows a sixth example of the aperture member. In Fig. 8, some of the transmission regions of the aperture member are located in a region with a radius of 30% to 50% of the pupil region of the illumination optical system.

[0136] The sixth example satisfies conditions (6) and (7). When estimating the refractive index distribution, the sixth example makes it possible to estimate a relatively coarse structure.

[0137] Fig. 9 shows a seventh example of the aperture member. In Fig. 9, some of the transmission regions of the aperture member are located in an area with a radius of 30% or less of the pupil region of the illumination optical system.

[0138] The seventh example satisfies the conditions (6) and (7). When estimating the refractive index distribution, the seventh example can estimate a relatively coarse structure.

[0139] Fig. 10 shows an eighth example of an aperture member. In Fig. 10, when the pupil of the illumination optical system is divided into four sectors with equal central angles, any of the transmission regions of the aperture member is located in each of the four sectors.

[0140] The eighth example satisfies the conditions (6) and (7). When estimating the refractive index distribution, in the eighth example, not only a relatively fine structure but also a relatively coarse structure can be estimated.

[0141] Fig. 11 shows a ninth example of the aperture member. In Fig. 11, some of the transmission regions of the aperture member are paired on either side of the center of the pupil of the illumination optical system.

[0142] The ninth example satisfies the conditions (6) and (7). When estimating the refractive index distribution, the ninth example can estimate a relatively coarse structure. Furthermore, the ninth example has a smaller number of transmission regions in the aperture member than the seventh example. When estimating the refractive index distribution, the estimation can be performed in a shorter time than the estimation time in the seventh example.

[0143] In the microscope system of this embodiment, it is preferable that each of the multiple light beams is emitted from each of multiple independent regions on a predetermined plane. The predetermined plane is a plane perpendicular to the optical axis of the detection optical system and is located on the opposite side of the detection optical system with respect to the specimen. A plurality of incoherent light sources are arranged on the predetermined plane, and each of the incoherent light sources corresponds to each of the second regions.

[0144] Fig. 16 is a diagram showing a microscope system according to this embodiment. The same components as those in Fig. 12 are given the same reference numbers and will not be described.

[0145] The microscope system 120 includes an incoherent light source 121 , an illumination optical system 122 , a detection optical system 3 , and an image sensor 4 .

[0146] In the microscope system 120, each of the multiple light beams is emitted from each of multiple regions on a predetermined plane. Each of the multiple regions is independent of each other. The predetermined plane is a plane that is perpendicular to the optical axis of the detection optical system 3 and is located on the opposite side of the detection optical system 3 with respect to the specimen 5. In the microscope system 120, the pupil plane PI of the illumination optical system 122 is the predetermined plane.

[0147] The incoherent light source 121 is disposed on a predetermined plane. In the microscope system 120, the incoherent light source 121 is disposed on a pupil plane PI.

[0148] The incoherent light source 121 includes a plurality of independent light emitting regions 121a. Each of the plurality of light emitting regions 121a is independent from each other.

[0149] The light emitting region 121a is a region that emits light. When light is emitted from the light emitting region 121a, a light beam L1 is irradiated onto the specimen 5. When light is emitted from another light emitting region 121a, a light beam L2 is irradiated onto the specimen 5.

[0150] The light emitting regions 121a are regions of light on the pupil plane PI. Each of the light emitting regions 121a corresponds to each of the second regions.

[0151] If the transmission region 112 shown in Fig. 15 is regarded as the light emitting region 121a, Fig. 15 can be regarded as a diagram showing an incoherent light source. Also, Figs. 3 to 11 can be regarded as specific examples of an incoherent light source. Therefore, the effects produced by the aperture members of the first to ninth examples are also produced when an incoherent light source is used.

[0152] The microscope system of this embodiment preferably further includes a processor, which determines a wavefront passing through an estimated specimen that is a model of the specimen for each light beam by forward propagation calculation, calculates an intensity distribution at an imaging position of the detection optical system corresponding to the wavefront for each light beam, generates a calculated image by adding up the intensity distributions for each light beam, and performs optimization processing to reduce the difference between the calculated image and a measurement image output from the image sensor, thereby reconstructing the estimated specimen.

[0153] The microscope system of this embodiment includes a processor. By including the processor, the microscope system of this embodiment can reconstruct the estimated specimen. In reconstructing the estimated specimen, for example, the refractive index distribution of the specimen is estimated.

[0154] In reconstructing the estimated specimen, the wavefront passing through the estimated specimen, which is a model of the specimen, is found for each light beam by forward propagation calculation. For each light beam, the intensity distribution at the imaging position of the detection optical system corresponding to the wavefront is calculated. A calculated image is generated by adding up the intensity distributions for each light beam. An optimization process is performed to reduce the difference between the calculated image and the measured image output from the image sensor.

[0155] The reconstruction of the putative specimen is described below.

[0156] In a microscope system 110 shown in Fig. 14, a specimen 5 is positioned between an illumination optical system 91 and a detection optical system 3. In a microscope system 120 shown in Fig. 16, a specimen 5 is positioned between an illumination optical system 122 and a detection optical system 3. The illumination optical system and the detection optical system form a measurement optical system.

[0157] Light beams are simultaneously incident on the specimen 5 from a plurality of directions. The specimen 5 is illuminated by the light beams simultaneously incident from a plurality of directions. In the microscope system of the embodiment, the illumination by the respective light beams is coherent illumination.

[0158] The detection optical system 3 forms an optical image 5' of the specimen 5. The optical image 5' is captured by the image sensor 4 to obtain a captured image I shown in FIG. mea (r) is obtained.

[0159] Photograph I mea (r) is input to the processor. The processor mea Using (r), a reconstruction of the estimated sample is performed. In the reconstruction, a simulation is performed.

[0160] In the first simulation, the specimen is a thin specimen. In the second simulation, the specimen is a thick specimen.

[0161] The first simulation will be described below. The optical system used in the first simulation is the measurement optical system of the microscope system 110 shown in FIG.

[0162] 14, the specimen 5 is a thin specimen. The focal position Fo of the detection optical system 3 is located inside the specimen 5. For example, the distance between the focal position Fo and the surface 5a of the specimen 5 is Δz1.

[0163] Since the specimen 5 is a thin specimen, one photographed image is acquired. Therefore, the detection optical system 3 and the image sensor 4 do not move in the optical axis direction. Moreover, the specimen 5 does not move in the optical axis direction either.

[0164] 17 is a flowchart of the first simulation. Before explaining the flowchart, an estimated sample and a wavefront will be explained.

[0165] FIG. 18 is a diagram showing an optical system used in the simulation. The optical system used in the simulation is mea The measurement optical system is the same as that used to acquire (r). In the simulation, an estimated specimen 130 is used instead of the specimen 5.

[0166] FIG. 18 shows the estimated specimen 130 and the wavefront f in m (r), amplitude transmittance T s (r), wavefront g out m (r) Wavefront u at the acquisition position of the captured image m(r), and the wavefront u at the image plane img m (r) is shown.

[0167] In the microscope system 110, an aperture member 111 is disposed at a pupil plane PI. The second region is a transmission region of the aperture member 111. Since light exits from the transmission region, the second region can be considered as a light source.

[0168] In Figure 18, the first light source is LS The second regions can be disposed at the pupil positions of the illumination optical system 91.

[0169] Returning to Fig. 17, the simulation will be described. The simulation includes a step of estimating an estimated specimen, a step of calculating an image of the estimated specimen, a step of optimizing the refractive index distribution of the estimated specimen, a step of updating the estimated specimen, and a step of reconstructing and outputting the structure of the estimated specimen.

[0170] In step S10, the number of second regions N LS is set. In the microscope system 110, the transmission region of the aperture member is located on the pupil plane of the illumination optical system. The transmission region is the second region. Therefore, in step S10, N LS The number of transparent areas is set to

[0171] Step S20 is a step of estimating an estimated specimen. One photographed image is acquired for the specimen 5. The estimated specimen 130 is a thin specimen, and can be regarded as one thin layer. Therefore, the initial value of the amplitude transmittance is set once.

[0172] In step S20, the amplitude transmittance T s The initial value is set to (r).

[0173] In order to calculate the image of the estimated specimen 130, information on the estimated specimen 130, for example, the refractive index distribution, is required. The estimated specimen 130 is a specimen that models the specimen 5. Therefore, it is preferable to use the refractive index distribution of the specimen 5 for the refractive index distribution of the estimated specimen 130.

[0174] However, the refractive index distribution of the specimen 5 is mea Therefore, the refractive index distribution of the estimated sample 130 must be estimated.

[0175] As shown in equation (10), the refractive index distribution n s (r) is the amplitude transmittance T s (r). Therefore, in step S20, the amplitude transmittance T s Set the initial value of (r).

number

number

[0176] Amplitude transmittance T s (r) value of the captured image I mea If it can be estimated from (r), the estimated value may be used as the initial value. s If the value of (r) can be estimated, the estimated value can be set as the initial value. If the initial value cannot be estimated, e.g., T s Let (r) = 1.

[0177] In step S30, the value of a variable m is initialized. Steps S41, S42, S43, S44, and S45 described below are executed for all light sources. The variable m represents the number of times these steps have been executed.

[0178] Steps S40 and S50 are steps for calculating the image of the estimated specimen. The number of images of the estimated specimen is the same as the number of captured images. Since the number of captured images is 1, the number of images of the estimated specimen is also 1.

[0179] Step S40 includes step S41, step S42, step S43, step S44, step S45, step S46, and step S47.

[0180] In step S41, a wavefront f in m (r) is calculated. in m (r) is the Nth light source from the first light source LS It represents the wavefronts of light emitted from the first light source through the second light source.

[0181] The pupil of the illumination optical system contains N LS If the first wavefront where the first light source is located is the wavefront emitted from each light source, then the wavefront f in m (r) represents the first wave front.

[0182] As described above, each of the second regions can be regarded as a point light source. In Fig. 18, the illumination light Lm is emitted from the m-th light source. The illumination light Lm is incident on the estimated specimen 130.

[0183] In this case, the wavefront f in m (r) is expressed by equations (11) and (12).

number

number

[0184] In step S42, the wavefront g emitted from the estimated specimen 130 out m (r) is calculated. For thin specimens, the wavefront g out m (r) is expressed by equation (13).

number

[0185] wave surface g out m (r) is the wavefront f in m (r) is the wavefront after passing through the estimated sample 130. in m Since (r) represents the first wavefront, wavefront g out m (r) represents the second wave front.

[0186] Since the estimated sample 130 is a thin sample, the wavefront f in m From (r), wavefront g out m (r) can be calculated directly.

[0187] In step S43, the wavefront u m The acquisition position of the photographed image is the focal position Fo of the detection optical system 3 on the specimen side when the photographed image is acquired.

[0188] Wavefront u m (r) is expressed by equation (14).

number

[0189] In step S60 described later, the residual is calculated. The photographed image and the image of the estimated specimen are used to calculate the residual. In order to calculate the image of the estimated specimen, it is necessary to obtain the wavefront at the acquisition position of the photographed image.

[0190] As described above, the distance between the focal position Fo and the surface 5a is Δz1. If the sign of the distance measured in the light traveling direction is positive, the acquisition position of the captured image is a position that is −Δz1 away from the surface 5a.

[0191] Therefore, in the optical system used in the simulation, the acquisition position of the photographed image is a position −Δz1 away from the surface 130a of the estimated specimen 130. In this case, the wavefront at the acquisition position of the photographed image is a wavefront at a position −Δz1 away from the surface 130a.

[0192] The wavefront u in Eq. (14) m (r) is the wavefront g out m (r) is a wavefront that has propagated by Δz1 in the direction opposite to the traveling direction of the light. This wavefront is at a position that is -Δz1 away from the surface 130a. Therefore, the wavefront u in Equation (14) m (r) represents the wavefront at the position where the captured image was taken.

[0193] Strictly speaking, the position where the photographed image is captured and the position of the surface 5a are different. However, since the specimen 5 is a thin specimen, the value of Δz1 is very small. Therefore, the position where the photographed image is captured and the position of the surface 5a can be considered to be substantially the same.

[0194] The estimated specimen 130 is also a thin specimen. Therefore, the position of the surface 130a and the position −Δz1 away from the surface 130a can be considered to be almost the same. That is, the wavefront g out m Position of (r) and wavefront u m The position of (r) can be considered to be almost the same. In this case, the wavefront u m Instead of (r), wavefront g out m (r) can also be used.

[0195] In step S44, the wavefront u img m (r) is calculated. m (r) propagates to the image plane IP. At this time, it passes through the detection optical system 3. The detection optical system 3 forms a Fourier optical system. Therefore, as shown in equation (15), the wavefront u img m (r) is the wavefront u m It can be calculated using (r) and the pupil function P(u) of the detection optical system.

number

[0196] In step S45, the wavefront u img m (r) is squared. Wavefront u img m (r) represents the amplitude of light. Therefore, the wavefront u img m The light intensity is calculated by squaring (r).

[0197] |u img m (r)| 2represents the light intensity distribution on the image plane IP. If the first intensity distribution is the light intensity distribution at the image position of the detection optical system, |u img m (r)| 2 represents a first light intensity distribution at the imaging position of the detection optical system.

[0198] Wavefront f in m (r), wavefront g out m (r), wave surface u m (r), and wavefront u img m (r) represents the illumination light emitted from the mth light source, that is, the wavefront generated by the illumination light emitted from one light source.

[0199] Image of putative specimen I est (r) is generated by the illumination light emitted from all light sources. Therefore, for all light sources, the wavefront f in m (r), wavefront g out m (r), wave surface u m (r), and wavefront u img m (r) must be sought.

[0200] In step S46, the value of the variable m is set to the number N of the second regions. LS If the result of the determination is NO, step S47 is executed. If the result of the determination is YES, step S50 is executed.

[0201] (If the result is NO: m ≠ N LS ) If the determination result is NO, in step S47, the value of the variable m is incremented by 1. When step S47 is completed, the process returns to step S41.

[0202] In step S47, the value of the variable m is incremented by 1. Therefore, for another light source, the wavefront f in m (r) is calculated, and in step S42, the wavefront gout m (r) is calculated, and in step S43, the wavefront u m (r) is calculated, and in step S44, the wavefront u img m (r) is calculated, and in step S45, |u img m (r)| 2 is calculated.

[0203] Steps S41, S42, S43, S44, and S45 are carried out to calculate |u img m (r)| 2 This is repeated until is found.

[0204] (If the answer is YES: m=N LS ) If the result of the determination is YES, in step S50, |u img m (r)| 2 The result is the estimated specimen image I est (r) is calculated. Image of estimated specimen I est (r) is expressed by equation (16).

number

[0205] FIG. 19 is a diagram showing an image of the estimated specimen. Image I of Estimated Specimen est (r) is the wavefront u for all light sources. img m As shown in Fig. 19, the wavefront u img m (r) is calculated, and the wavefront u img m (r) to |u img m (r)| 2 is calculated |u img m (r)| 2 are all added together. The result is the image I of the estimated specimen. est (r) is calculated.

[0206] In step S60, the residual is calculated. The residual is expressed by equation (17). As shown in equation (17), the residual is calculated by subtracting the captured image I mea (r) and image of presumed specimen I est (r) is calculated.

number

[0207] Equation (17) represents the norm of a matrix. The norm is expressed by equation (18).

number

[0208] In step S70, the residual is compared with a threshold value. If the result of the determination is NO, step S80 is executed. If the result of the determination is YES, step S110 is executed.

[0209] (If the result is NO: Residual ≧ Threshold)

[0210] In step S80, the value of a variable m is initialized. Steps S91 and S92 described below are executed for all light sources. The variable m indicates the number of times these steps have been executed.

[0211] Step S90 is a step for optimizing the refractive index distribution of the estimated specimen.

[0212] Step S90 includes step S91, step S92, step S93, and step S94.

[0213] In step S91, the wavefront u' m (r) is calculated. m In the calculation of (r), the captured image I mea (r) and image of presumed specimen I est (r) is used. Also, the wavefront u' m(r) is the wavefront at the position where the captured image is taken.

[0214] Wavefront u' m (r) is expressed by equation (19).

number

[0215] Fig. 20 is a diagram showing correction of a wavefront. Fig. 20(a) is a diagram showing a wavefront before correction emerging from an estimated specimen. Fig. 20(b) is a diagram showing a wavefront before correction at the acquisition position of a photographed image. Fig. 20(c) is a diagram showing a wavefront after correction at the acquisition position of a photographed image. Fig. 20(d) is a diagram showing a wavefront after correction emerging from an estimated specimen.

[0216] As shown in Figure 19, image I of the estimated specimen est (r) is the wavefront u img m In addition, as shown in FIG. 19 and FIG. 20(b), the wavefront u img m (r) is the wavefront u m Calculated based on (r).

[0217] As shown in FIG. 20(a), the wavefront u m To calculate (r), the amplitude transmittance T s (r) is used. The amplitude transmittance T s (r) is the estimated amplitude transmittance. When step S90 is executed for the first time, this amplitude transmittance T s (r) is different from the amplitude transmittance of sample 5.

[0218] Amplitude transmittance T s The larger the difference between (r) and the amplitude transmittance of specimen 5, the larger the estimated specimen image I est (r) and image I mea The difference with (r) also becomes large. Therefore, the estimated specimen image I est (r) and image I mea The difference between (r) and (r) is the amplitude transmittance T sThis can be considered to reflect the difference between the amplitude transmittance of (r) and that of specimen 5.

[0219] Therefore, as shown in equation (11), the estimated sample image I est (r) and image I mea Using (r), the wavefront u m As a result, as shown in FIG. 20(c), the wavefront after correction, that is, the wavefront u' m (r) is obtained.

[0220] Wavefront u' m By using (r), a new amplitude transmittance T s (r) can be calculated. m (r) is the wavefront u m (r) is different. Therefore, the new amplitude transmittance T s (r) is the wavefront u m This differs from the amplitude transmittance when (r) was calculated.

[0221] Thus, the wavefront u' m Using (r), the amplitude transmittance T s (r) can be calculated. However, as shown in FIG. 20(a), the amplitude transmittance T s To calculate (r), the wavefront g out m (r) is necessary.

[0222] As shown in Fig. 20(a) and (c), the wavefront u' m The position of (r) and the wavefront g out m The position of (r) is different. Therefore, the amplitude transmittance T s In order to calculate (r), the wavefront g' is out m (r) is necessary.

[0223] wave surface g' out m (r) is expressed by equation (20). m (r) is the wavefront after correction, so the wavefront g' out m (r) is also the wavefront after correction.

number

[0224] As described above, the capture position of the captured image is a position that is -Δz1 away from the surface 130a. In other words, the position of the surface 130a is a position that is Δz1 away from the capture position of the captured image. Therefore, the wavefront at the position of the surface 130a is a wavefront at a position that is Δz1 away from the capture position of the captured image.

[0225] Wavefront g' in Eq. (20) out m (r) is the wavefront u' m (r) is the wavefront that has propagated Δz1 in the direction of light travel. This wavefront is at a position Δz1 away from the position where the shadow image was captured. Therefore, the wavefront g' in equation (20) out m (r) represents the wavefront at the location of surface 130a.

[0226] The wavefront at the surface 130a is f in m (r) is the wavefront after passing through the estimated specimen 130. As described above, f in m (r) represents the first wavefront. If the second wavefront is the wavefront after the first wavefront passes through the estimated sample, then the wavefront g' out m (r) represents the second wave front.

[0227] As described above, the value of Δz1 is very small. In addition, the estimated specimen 130 is a thin specimen. Therefore, the acquisition position of the photographed image and the position separated by Δz1 from the acquisition position of the photographed image can be considered to be substantially the same. That is, the wavefront u' m The position of (r) and the wavefront g out m The position of (r) can be considered to be almost the same. In this case, the wavefront g' out m Instead of (r), the wavefront u' m (r) can also be used.

[0228] In step S92, the gradient ΔT s m (r) is calculated. The gradient of the sample, ΔT s m is expressed by equation (21). The gradient of the sample ΔT s m To calculate (r), for example, the gradient descent method can be used.

number

[0229] As shown in FIG. 20(a), the wavefront g out m To calculate (r), the amplitude transmittance T s (r) is used. The amplitude transmittance T s (r) is the estimated amplitude transmittance. Therefore, this amplitude transmittance T s (r) is different from the amplitude transmittance of sample 5.

[0230] Amplitude transmittance T s The greater the difference between the amplitude transmittance of (r) and that of the specimen 5, the greater the wavefront g out m (r) and wavefront g' out m The difference with (r) also becomes large. Therefore, the wavefront g out m (r) and wavefront g' out m The difference between (r) and (r) is the amplitude transmittance T s This can be considered to reflect the difference between the amplitude transmittance of (r) and that of specimen 5.

[0231] Wavefront f in m (r), amplitude transmittance T s (r), wavefront g out m (r), and wavefront g'out m (r) is known. Then, as shown in equation (21), the wavefront f in m (r), amplitude transmittance T s (r), wavefront g out m (r), and wavefront g' out m (r) Using the gradient ΔT of the specimen s m (r) can be calculated.

[0232] FIG. 21 is a diagram showing the gradient of the specimen.

[0233] The gradient ΔT of the sample obtained in step S92 s m (r) represents the gradient of the sample in the illumination light emitted from a single light source. The gradient of the sample ΔT s m (r) is determined by the illumination light emitted from all light sources. Therefore, for all light sources, the gradient of the specimen ΔT s m (r) must be sought.

[0234] In step S93, the value of the variable m is set to the number of light sources N LS If the result of the determination is NO, step S94 is executed. If the result of the determination is YES, step S100 is executed.

[0235] (If the result is NO: m ≠ N LS ) If the determination result is NO, in step S94, the value of the variable m is incremented by 1. When step S94 is completed, the process returns to step S91.

[0236] In step S94, the value of the variable m is incremented by 1. Therefore, for another light source, the wavefront u' is calculated in step S91. m (r) is calculated, and in step S92, the gradient of the sample ΔT s m (r) is calculated.

[0237] Steps S91 and S92 are carried out to calculate the gradient ΔT of the specimen for all light sources. s m This is repeated until (r) is found.

[0238] FIG. 22 shows the gradient of the specimen. In FIG. 22, the gradient of the specimen ΔT s m (r) is required.

[0239] (If the answer is YES: m=N LS ) If the result of the determination is YES, in step S100, the amplitude transmittance T s (r) is updated. Step S100 is a step of updating the estimation samples.

[0240] Updated amplitude transmission T s (r) is expressed by equation (22).

number

[0241] Furthermore, when the sample 5 is considered as a perfect phase object with no absorption, the amplitude transmittance T s (r) can be further updated.

number

[0242] After step S100 is completed, the process returns to step S30. s At (r), steps S30 to S100 are executed.

[0243] By repeatedly executing steps S30 to S100, the updated amplitude transmittance T s(r) gradually approaches the amplitude transmittance of the sample 5. In other words, the residual becomes smaller. Eventually, the residual becomes smaller than the threshold value.

[0244] (If the result is YES: Residual < Threshold) In step 110, the estimated refractive index distribution of the specimen is calculated. The obtained amplitude transmittance T s (r) is the same as or approximately the same as the amplitude transmittance of the sample 5. The obtained amplitude transmittance T s The refractive index distribution n(r) can be calculated from (r) and equation (1).

[0245] By using the refractive index distribution n(r) obtained in step S110, the structure of the estimated specimen can be reconstructed. The reconstructed structure of the estimated specimen can be output to, for example, a display device. The estimated specimen 130 is a thin specimen. In the first simulation, the structure of the thin specimen can be reconstructed.

[0246] As described above, the amplitude transmittance T obtained in step S110 s (r) is the same as or approximately the same as the amplitude transmittance of the sample 5. In this case, the refractive index distribution n(r) can also be considered to be the same as or approximately the same as the refractive index distribution of the sample 5. Therefore, the structure of the reconstructed estimated sample 130 can be considered to be the same as or approximately the same as the structure of the sample 5.

[0247] In the first simulation, steps S40, S50, and S90 are repeatedly executed. As a result, the amplitude transmittance T s (r) is updated. As described above, steps S40 and S50 are steps for calculating the image of the estimated specimen. Step S90 is a step for optimizing the refractive index distribution of the estimated specimen.

[0248] Amplitude transmittance T s (r) represents the estimated specimen. Therefore, the step of calculating the image of the estimated specimen and the step of optimizing the refractive index distribution of the estimated specimen are repeatedly performed, thereby updating the estimated specimen.

[0249] The second simulation will now be described. The optical system used in the second simulation is the measurement optical system of the microscope system 120 shown in FIG.

[0250] Fig. 23 is a diagram showing a microscope system according to this embodiment. The same components as those in Fig. 16 are given the same reference numbers and will not be described.

[0251] The sample 140 is a thick sample. Light beams are simultaneously incident on the sample 140 from a plurality of directions. In FIG. 23, light beams L1 and L2 are shown.

[0252] The light emitted from the specimen 140 is collected on an image plane IP by the detection optical system 3. An optical image 140' is formed on the image plane IP. The optical image 140' is an optical image of the specimen 140.

[0253] The microscope system 120 includes a moving stage 141. The moving stage 141 moves in the direction of an optical axis AX.

[0254] As described above, the captured images are used to optimize the refractive index distribution of the estimated specimen. Since the specimen 140 is a thick specimen, multiple captured images are acquired. In order to acquire multiple captured images, the specimen 140 is fixed, and the focal position of the detection optical system 3 is moved by the moving stage 141.

[0255] The detection optical system 3 has, for example, an infinity corrected objective lens and an imaging lens. In this case, the focal position of the detection optical system 3 can be moved by moving the objective lens. The detection optical system 3 and the image sensor 4 may be fixed, and the specimen 140 may be moved.

[0256] The following describes a case where four captured images are acquired.

[0257] Fig. 24 is a diagram showing captured images. Fig. 24(a) is a diagram showing an image captured at a first position. Fig. 24(b) is a diagram showing an image captured at a second position. Fig. 24(c) is a diagram showing an image captured at a third position. Fig. 24(d) is a diagram showing an image captured at a fourth position.

[0258] The focal position Fo of the specimen 140 is changed by changing the distance between the detection optical system 3 and the specimen 140. Here, the focal position Fo of the specimen 140 is changed four times. As a result, the following four captured images are acquired. Photograph I mea1 (r): Image at a distance of 3×Δz from surface 140a. Photograph I mea2 (r): Image at a distance of 2×Δz from surface 140a. Photograph I mea3 (r): Image at a distance Δz from surface 140a. Photograph I mea4 (r): Image of surface 140a.

[0259] Photograph I mea1 (r), Image I mea2 (r), Image I mea3 (r) and the captured image I mea4 (r) is input to the processor, where the four captured images are used to reconstruct the estimated specimen. The reconstruction is performed through simulation.

[0260] 25 and 26 are flowcharts of the second simulation. The same processes as those in the first flowchart are given the same numbers and will not be described. Before describing the flowchart, the estimated sample and the wavefront will be described.

[0261] Fig. 27 is a diagram showing an optical system used in the simulation. The same components as in Fig. 16 are given the same reference numbers and their explanations are omitted.

[0262] The optical system used in the simulation is mea1(r), Image I mea2 (r), Image I mea3 (r) and the captured image I mea4 The measurement optical system is the same as that used to acquire (r). In the simulation, an estimated specimen 150 is used instead of the specimen 140.

[0263] Figure 27 shows the estimated specimen 150, wavefront f in m (r), amplitude transmittance T z (r), and wavefront g out m (r) is shown.

[0264] When the estimated sample is a thin sample, the wavefront f in m From (r), wavefront g out m (r) can be calculated directly. However, if the estimated specimen is a thick specimen, the wavefront f in m (r) to wavefront g out m It is difficult to calculate (r) directly.

[0265] The estimated specimen 150 is a thick specimen. Therefore, the estimated specimen 150 is replaced with a number of thin layers along the optical axis direction. Then, for each thin layer, the wavefronts on both sides of the layer are calculated.

[0266] FIG. 28 is a diagram showing the wavefront in each layer. The calculation of the wavefront will be described later. The number of layers can be the same as the number of acquired images. However, the number of layers may be greater than the number of acquired images. In FIG. 28, the number of layers is the same as the number of acquired images.

[0267] In FIG. 28, the position Z=1 is the position of the first layer, the position Z=2 is the position of the second layer, the position Z=3 is the position of the third layer, and the position Z=4 is the position of the fourth layer.

[0268] Returning to FIG. 25 and FIG. 26, the simulation will be described.

[0269] In step S10, the number of second regions N LS In the microscope system 120, an incoherent light source is located on the pupil plane of the illumination optical system. Therefore, in step S10, N LS The number of light sources is set to

[0270] In step S200, the number of layers N IM is set. The estimated specimen 150 is a thick specimen. Therefore, as described above, we replace the estimated specimen 150 with multiple thin layers. The number of layers N IM represents the number of thin layers.

[0271] Images are taken at multiple positions in the specimen 140. The number of layers N IM can be equal to the number of positions at which images were taken. If the focal position Fo relative to the specimen 140 is changed four times, N IM =4.

[0272] 1 to N IM The numbers up to indicate the position of the thin layer. For example, N IM For =4, the number 1 represents the position of the first layer, the number 2 represents the position of the second layer, the number 3 represents the position of the third layer, and the number 4 represents the position of the fourth layer.

[0273] The image of the estimated specimen is calculated by simulation. Therefore, the number of layers N IM can be freely set. For example, the number of layers N IM can be greater than the number of positions at which captured images were taken.

[0274] For example, N IM When = 7, the number of thin layers is seven. In this case, seven images of the estimated specimen are calculated. In the simulation, the photographed images and the images of the estimated specimen in the thin layers are used, as described below. Therefore, the seven positions where the images of the estimated specimen are calculated include the four positions where the photographed images were obtained.

[0275] The relationship between the seven positions and the captured images can be, for example, as follows:

[0276] The number 1 represents the position of the first layer. At this position, the captured image I mea1 (r) is acquired. Also, at this position, the image of the estimated specimen in the first layer is calculated. Therefore, the image of the estimated specimen in the first layer and the captured image I mea1 (r) will be used in the steps described below.

[0277] The number 2 represents the location of the second layer. No images were taken at this location.

[0278] The number 3 represents the position of the third layer. At this position, the captured image I mea2 (r) is acquired. Also, at this position, the image of the estimated specimen in the third layer is calculated. Therefore, the image of the estimated specimen in the third layer and the captured image I mea2 (r) will be used in the steps described below.

[0279] The number 4 represents the location of the fourth layer. No images were taken at this location.

[0280] The number 5 represents the position of the fifth layer. At this position, the image I mea3 (r) is acquired. Also, at this position, the image of the estimated specimen in the fifth layer is calculated. Therefore, the image of the estimated specimen in the fifth layer and the captured image I mea3 (r) will be used in the steps described below.

[0281] The number 6 represents the location of the sixth layer. No images were taken at this location.

[0282] The number 7 represents the position of the seventh layer. At this position, the image I mea4 (r) is acquired. Also, at this position, the image of the estimated specimen in the seventh layer is calculated. Therefore, the image of the estimated specimen in the seventh layer and the captured image I mea4 (r) will be used in the steps described below.

[0283] In step S210, the number of corrections N CR is set.

[0284] In step S220, the value of a variable z is initialized. Step S231, which will be described later, is executed for all acquisition positions. The variable z indicates the number of times that step S231 has been executed.

[0285] Step S230 is a step of estimating an estimated specimen. Four photographed images are acquired for the specimen 140. As described above, the estimated specimen 150 is replaced with four thin layers. Therefore, the initial value of the amplitude transmittance is set four times.

[0286] Step S230 includes step S231, step S232, and step S233.

[0287] In step S231, the amplitude transmittance T z The initial value is set to (r).

[0288] The transport of intensity equation may be used to set the initial values. The transport of intensity equation is disclosed in, for example, the following document: MR Teague, “Deterministic phase retrieval: a Greens function solution,” J. Opt. Soc. Am. 73, 1434-1441 (1983)

[0289] The intensity transport equation at the focal position Z0 is expressed by equation (24).

number

[0290] Using equation (24), the phase distribution φ of the specimen is calculated from the focused image and two defocused images. Z0 (r) can be easily found.

[0291] However, if the difference in light intensity at the same point of two defocused images is zero or very small, the phase cannot be measured. Even with partial coherent illumination, if the numerical aperture of the illumination light is close to the numerical aperture of the objective lens, the difference in light intensity will be zero or very small. Therefore, in such cases, it is difficult to set the initial value using the transport of intensity equation.

[0292] As mentioned above, the phase distribution φ Z0 (r) is calculated from a focused image and two defocused images. The focused image is acquired, for example, by moving the objective lens in the optical axis direction at regular intervals. In this case, multiple focused images are acquired discretely along the optical axis. Therefore, the two defocused images are also acquired discretely.

[0293] The phase distribution φ expressed by equation (24) Z0 (r) is the phase distribution in the plane perpendicular to the optical axis. Since the focused image and the two defocused images are acquired discretely, the phase distribution φ Z0 The planes representing (r) are also located discretely along the optical axis.

[0294] As shown in equation (25), the phase distribution φ z (r) is the amplitude transmittance T s (r). In this way, the amplitude transmittance T z An initial value can be set for (r).

number

[0295] Phase distribution φ obtained from the transport of intensity equation Z0 is the phase distribution φ z (r) can be used. The initial value can be set using the transport of intensity equation. If it is difficult to estimate the initial value, e.g., T z It is also acceptable to set (r)=1.

[0296] In step S232, the value of the variable z is set to the number N of acquisition positions. IM If the result of the determination is NO, step S233 is executed. If the result of the determination is YES, step S30 is executed.

[0297] (If the result is NO: z ≠ N IM ) If the determination result is NO, in step S233, the value of the variable z is incremented by 1. When step S233 is completed, the process returns to step S231.

[0298] In step S233, the value of the variable z is incremented by 1. Therefore, for another acquisition position, the amplitude transmittance T z The initial value is set to (r).

[0299] Step S231 is repeated until the initial values ​​are set for all acquisition positions.

[0300] (If the result is YES: z=N IM ) In step S30, the value of a variable m is initialized. Steps S240, S41, S42, S251, and S260 described below are executed for all light sources. The variable m represents the number of times these steps have been executed.

[0301] In step S240, function I estz The value of (r) is initialized. I estz (r) represents the image of the putative specimen 30. As mentioned above, the image of the putative specimen 150 has been displaced into four thin layers. Thus, I estz(r) shows the image of a thin layer.

[0302] Steps S250 and S270 are steps for calculating the images of the estimated specimen. The number of images of the estimated specimen is the same as the number of captured images. Since the number of captured images is four, the number of images of the estimated specimen is also four.

[0303] Step S250 includes step S41, step S42, step S251, step S252, step S253, and step S260.

[0304] In step S41, a wavefront f in m (r) is calculated. Wavefront f in m (r) is expressed by the above equations (11) and (12).

[0305] In step S42, the wavefront g emerging from the estimated specimen 150 is out m (r) is calculated. Wavefront g out m (r) is the wavefront f in m (r). The estimated specimen 150 is displaced into four thin layers. Thus, the wavefront is calculated at each of the thin layers.

[0306] In FIG. 28, the position Z=1 is the position of the first layer, the position Z=2 is the position of the second layer, the position Z=3 is the position of the third layer, and the position Z=4 is the position of the fourth layer.

[0307] The four thin layers are spaced equally apart. The distance between two adjacent layers is Δz. The wavefront propagates between the two layers. Therefore, Δz represents the propagation distance.

[0308] Wavefront f in the first layer 1 m (r) is expressed by equation (26) and equation (12).

number

[0309] The position of the first layer coincides with the position of the surface 150b of the estimated specimen 150. The surface 150b has a wavefront f in m (r) is incident. Therefore, the wavefront f 1 m (r) is the wavefront f in m In FIG. 28, the wavefront f 1 m Instead of (r), the wavefront f in m (r) is shown.

[0310] Wavefront g at the first layer 1 m (r) is expressed by equation (27).

number

[0311] Wavefront f in the second layer 2 m (r) is the wavefront g 1 m (r) is the wavefront when it propagates by Δz. Wavefront f 2 m (r) is expressed by equation (28).

number

[0312] Wavefront g at the second layer 2m (r) is expressed by equation (29).

number

[0313] Wavefront f at the third layer 3 m (r) is the wavefront g 2 m (r) is the wavefront when it propagates by Δz. The wavefront f at the third layer 3 m (r) is expressed by equation (30).

number

[0314] Wavefront g at the third layer 3 m (r) is expressed by equation (31).

number

[0315] Wavefront f at the fourth layer 4 m (r) is the wavefront g 3 m (r) is the wavefront when it propagates by Δz. The wavefront f at the fourth layer 4 m (r) is expressed by equation (32). By setting ΔD=Δz in equation (21), the wavefront f 4 m (r) can be calculated.

number

[0316] Wavefront g at the fourth layer 4 m (r) is expressed by equation (33).

number

[0317] The position of the fourth layer coincides with the position of the surface 150a of the estimated specimen 150. From the surface 150a, the wavefront g out m (r) is emitted. Therefore, the wavefront g 4 m (r) is the wavefront g out m In FIG. 28, the wavefront g 4 m Instead of (r), wavefront g out m (r) is shown.

[0318] As described above, when the estimated specimen is thick, it is replaced with multiple thin layers and the wavefront propagating between the two layers is obtained, so that the wavefront g out m (r) can be calculated.

[0319] In step S251, the value of a variable z is initialized. Steps S261, S262, and S263 described below are executed for all acquisition positions. The variable z indicates the number of times these steps have been executed.

[0320] Step S260 includes step S261, step S262, step S263, step S264, and step S265.

[0321] In step S261, a wavefront u at the acquisition position of the captured image is z m (r) is calculated. z m(r) is expressed by equation (34).

number

[0322] In step S262, the wavefront u imgz m (r) is calculated. imgz m (r) is expressed by equation (35).

number

[0323] In step S263, the wavefront u imgz m (r) is squared. Wavefront u imgz m (r) represents the amplitude of light. Therefore, the wavefront u imgz m The light intensity is calculated by squaring (r).

[0324] |u imgz m (r)| 2 represents the light intensity distribution on the image plane IP. If the first intensity distribution is the light intensity distribution at the image position of the detection optical system, |u imgz m (r)| 2 represents a first light intensity distribution at the imaging position of the detection optical system.

[0325] In step S264, the value of the variable z is set to the number N of acquisition positions. IM If the result of the determination is NO, step S265 is executed. If the result of the determination is YES, step S252 is executed.

[0326] (If the result is NO: z ≠ N IM ) If the determination result is NO, in step S265, the value of the variable z is incremented by 1. When step S265 ends, the process returns to step S261.

[0327] In step S265, the value of the variable z is incremented by 1. Therefore, steps S261, S262, and S263 are executed for another acquisition position.

[0328] Steps S261, S262, and S263 are repeated until the initial values ​​are set for all acquisition positions.

[0329] The process in step S250 will be described using layers 1 and 4. Layers 2 and 3 can be considered in the same way as layer 1.

[0330] Fig. 29 is a diagram showing a wavefront at the acquisition position of a captured image and a wavefront at the imaging plane. Fig. 29(a) is a diagram showing two wavefronts at the first layer. Fig. 29(b) is a diagram showing two wavefronts at the fourth layer.

[0331] The image captured at z=1 is the captured image I mea1 (r) Photograph I mea1 (r) is an image at a position that is 3×Δz away from the surface 140a. The first layer is 3×Δz away from the surface 150a. Therefore, the position of the first layer is the same as that of the captured image I mea1 Corresponding to the acquisition position of (r).

[0332] wave surface g out m The exit position of (r) coincides with the surface 150a. As shown in FIG. 29(a), the wavefront g out m The exit position of (r) is different from the position of the first layer. The first layer is the wavefront g out m It is 3×Δz away from the injection position of (r).

[0333] Wavefront u in the first layer 1 m (r) is the wavefront gout m (r) is the wavefront when the light propagates by 3×Δz in the opposite direction to the traveling direction of the light. Therefore, by setting ΔD=-3×Δz in step S261, the wavefront u 1 m (r) can be calculated.

[0334] Wavefront u 1 m After (r) is calculated, in step S262, the wavefront u on the image plane is calculated from equation (35). img1 m (r) is calculated.

[0335] Furthermore, in step S263, the light intensity |u img1 (r)| 2 is calculated.

[0336] The image captured at z=2 is image I mea2 (r) Photograph I mea2 (r) is an image at a position at a distance of 2×Δz from the surface 140a. The second layer is separated from the surface 150a by 2×Δz. Therefore, the position of the second layer is located in the captured image I mea2 Corresponding to the acquisition position of (r).

[0337] wave surface g out m The exit position of (r) is different from the position of the second layer. The second layer is the wavefront g out m It is 2×Δz away from the injection position of (r).

[0338] Wavefront u at the second layer 2 m (r) is the wavefront g out m (r) is the wavefront when the light propagates by 2×Δz in the opposite direction to the traveling direction. Therefore, by setting ΔD=-2×Δz in step S261, the wavefront u 2 m (r) can be calculated.

[0339] Wavefront u 2m Once (r) is calculated, in step S262, the wavefront u img2 m (r) is calculated.

[0340] Furthermore, in step S263, the light intensity |u img2 (r)| 2 is calculated.

[0341] The image captured at z=3 is image I mea3 (r) Photograph I mea3 (r) is an image at a position at a distance Δz from the surface 140a. The third layer is separated from the surface 150a by Δz. Therefore, the position of the third layer is located in the captured image I mea3 Corresponding to the acquisition position of (r).

[0342] wave surface g out m The exit position of (r) is different from that of the third layer. The third layer is the wavefront g out m It is Δz away from the injection position of (r).

[0343] Wavefront u at the third layer 3 m (r) is the wavefront g out m (r) is the wavefront when the light propagates by Δz in the opposite direction to the traveling direction. Therefore, by setting ΔD=Δz in step S261, the wavefront u 3 m (r) can be calculated.

[0344] Wavefront u 3 m Once (r) is calculated, in step S262, the wavefront u img3 m (r) is calculated.

[0345] Furthermore, in step S263, the light intensity |u img3 (r)| 2 is calculated.

[0346] The image captured at z=4 is image I mea4 (r) Photograph I mea4 (r) is an image of the surface 140a. The fourth layer coincides with the surface 150a. Therefore, the position of the fourth layer is the same as that of the captured image I mea4 Corresponding to the acquisition position of (r). wave surface g out m The exit position of the wavefront g(r) is the surface 150a. out m The injection position of (r) is the same as that of the fourth layer. Wavefront u at the fourth layer 4 m (r) is the wavefront g out m (r) The same. Wavefront g out m (r) is the wavefront u 4 m It can be replaced with (r). Wavefront u 4 m Once (r) is calculated, in step S262, the wavefront u img4 m

[0347] wave surface g out m The exit position of the wavefront g(r) is the surface 150a. out m The injection position of (r) is the same as that of the fourth layer.

[0348] Wavefront u at the fourth layer 4 m (r) is the wavefront g out m (r) The same. Wavefront g out m (r) is the wavefront u 4 m It can be replaced with (r).

[0349] Wavefront u 4 m Once (r) is calculated, in step S262, the wavefront u img4m (r) is calculated.

[0350] Furthermore, in step S263, the light intensity |u img4 (r)| 2 is calculated.

[0351] Wavefront u z m (r) and wavefront u imgz m (r) represents the illumination light emitted from the mth light source, that is, the wavefront generated by the illumination light emitted from one light source.

[0352] Image of putative specimen I estz (r) is generated by the illumination light emitted from all light sources at the acquisition position, so the wavefront must be calculated for all light sources.

[0353] (If the result is YES: z=N IM ) Step S242 is executed.

[0354] Wavefront f in m (r), wavefront g out m (r), wave surface u z m (r), and wavefront u imgz m (r) represents the illumination light emitted from the mth light source, that is, the wavefront generated by the illumination light emitted from one light source.

[0355] Image of putative specimen I estz (r) is generated by the illumination light emitted from all light sources. Therefore, for all light sources, the wavefront f in m (r), wavefront g out m (r), wave surface u z m (r), and wavefront u imgz m (r) must be sought.

[0356] In step S252, the value of the variable m is set to the number of light sources N LS If the result of the determination is NO, step S253 is executed. If the result of the determination is YES, step S270 is executed.

[0357] (If the result is NO: m ≠ N LS ) If the determination result is NO, in step S253, the value of the variable m is incremented by 1. After step S253 is completed, the process returns to step S41.

[0358] In step S253, the value of the variable m is incremented by 1. Therefore, for another light source, the wavefront f in m (r) is calculated, and in step S42, the wavefront g out m (r) is calculated, and in step S261, the wavefront u z m (r) is calculated, and in step S262, the wavefront u imgz m (r) is calculated, and in step S263, |u imgz m (r)| 2 is calculated.

[0359] Steps S41, S42, S251, and S260 are performed to calculate |u imgz m (r)| 2 This is repeated until is found.

[0360] (If the answer is YES: m=N LS ) If the result of the determination is YES, in step S270, |u imgz m (r)| 2 The result is the estimated specimen image I estz (r) is calculated. Image of estimated specimen I estz (r) is expressed by equation (36).

number

[0361] Fig. 30 is a diagram showing an image of an estimated specimen, Fig. 30(a) is a diagram showing an image of an estimated specimen in the first layer, and Fig. 30(b) is a diagram showing an image of an estimated specimen in the fourth layer.

[0362] Image of putative specimen I est1 (r) is the wavefront u for all light sources. img1 m (r) is the image obtained. Image I of the estimated specimen est4 (r) is the wavefront u for all light sources. img4 m This is the image that is obtained when (r) is

[0363] As shown in FIG. 30(a), for each light source, the wavefront u img1 m (r) is calculated, and the wavefront u img1 m (r) to |u img1 m (r)| 2 is calculated, and |u img1 m (r)| 2 are all added together. As a result, the estimated specimen image I est1 (r) is calculated.

[0364] As shown in FIG. 30(b), for each light source, the wavefront u img4 m (r) is calculated, and the wavefront u img4 m (r) to |u img4 m (r)| 2 is calculated, and |u img4 m (r)| 2 are all added together. As a result, the estimated specimen image I est4 (r) is calculated.

[0365] (If the answer is YES: m=N LS ) In step S280, the residual is calculated. The residual is expressed by equation (37). As shown in equation (37), the residual is calculated by subtracting the captured image I meaz (r) and image of presumed specimen I estz (r) is calculated.

number

[0366] As mentioned above, the number of captured images is 4, and the number of estimated specimen images is also 4. Therefore, I mea1 (r) and I est1 From (r), the residuals in the first layer are calculated. mea2 (r) and I est2 From (r), the residuals at the second layer are calculated. mea3 (r) and I est3 From (r), the residuals at the third layer are calculated. mea4 (r) and I est4 From (r), the residuals at the fourth layer are calculated.

[0367] The residual used in step 70 is calculated from the residual at the first layer, the residual at the second layer, the residual at the third layer, and the residual at the fourth layer.

[0368] In step S70, the residual is compared with a threshold value. If the result of the determination is NO, step S290 is executed. If the result of the determination is YES, step S110 is executed.

[0369] (If the result is NO: Residual ≧ Threshold) In step S290, the value of a variable L is initialized. Steps S301, S302, S303, S304, and S310 described below are executed the number of times set in step S210. The variable L indicates the number of times these steps have been executed.

[0370] Step S300 includes step S301, step S302, step S303, step S304, step S305, step S306, and step S310.

[0371] In step S301, 1 to N IM In step S311 described later, a corrected wavefront is calculated. In the calculation of the corrected wavefront, one captured image and one estimated sample image are used.

[0372] As described above, in step S270, multiple estimated specimen images are calculated. The image of the estimated specimen used to calculate the corrected wavefront is one. Therefore, the image of the estimated specimen used to calculate the corrected wavefront is selected from the multiple estimated specimen images.

[0373] N IM is the number of layers. N IM If = 4, then in step S301, one number is randomly selected from the numbers 1 to 4.

[0374] For example, if the selected number is 1, the number 1 represents the first layer. The image of the estimated specimen in the first layer corresponds to the image captured at the first acquisition position. Therefore, the image captured at the first acquisition position and the image of the estimated specimen in the first layer are used to calculate the corrected wavefront.

[0375] Furthermore, for example, if the selected number is 4, the selected number represents the fourth layer. The image of the estimated specimen in the fourth layer corresponds to the image captured at the fourth acquisition position. Therefore, the image captured at the fourth acquisition position and the image of the estimated specimen in the fourth layer are used to calculate the corrected wavefront.

[0376] In step S302, the value selected in step S301 is input to the variable zL. As described above, in step S301, 1 to N IM One number is randomly selected from the numbers up to 1. For example, if the selected number is 1, then 1 is input to the variable zL in step S302.

[0377] In step S303, the value of a variable m is initialized. Steps S311, S312, and S313 described below are executed for all light sources. The variable m represents the number of times these steps have been executed.

[0378] Step S310 is a step for optimizing the refractive index distribution of the estimated specimen.

[0379] Step S310 includes step S311, step S312, step S313, step S314, and step S315.

[0380] In step S311, the wavefront u' zL m (r) is calculated. zL m (r) is the wavefront at the position of the layer indicated by the value of the variable zL.

[0381] Wavefront u' zL m In the calculation of (r), the captured image I meazL (r) and image of presumed specimen I estzL (r) is used. Image I meazL (r) is the captured image I meaz The image of the estimated specimen I is taken at the position indicated by the value of the variable zL. estzL (r) is an image of the putative specimen I estz is an image of an estimated sample at a position indicated by the value of variable zL.

[0382] Wavefront u' zL m (r) is expressed by equation (38).

number

[0383] Fig. 31 is a diagram showing correction of a wavefront. Fig. 31(a) is a diagram showing a wavefront before correction emerging from an estimated specimen. Fig. 31(b) is a diagram showing a wavefront before correction at the acquisition position of a photographed image. Fig. 31(c) is a diagram showing a wavefront after correction at the acquisition position of a photographed image. Fig. 31(d) is a diagram showing a wavefront after correction emerging from an estimated specimen.

[0384] The case where the number selected in step S301 is 1, that is, zL=1, will be described.

[0385] As shown in FIG. 30(a), the image I of the estimated specimen est1 (r) is the wavefront u img1 m In addition, as shown in FIG. 30(a) and FIG. 31(b), the wavefront u img1 m (r) is the wavefront u 1 m Calculated based on (r).

[0386] As shown in FIG. 31(a), the wavefront u 1 m To calculate (r), the amplitude transmittance T z (r) is used. The amplitude transmittance T z (r) is the estimated amplitude transmittance. When step S300 is executed for the first time, this amplitude transmittance T z (r) is different from the amplitude transmittance of the sample 140.

[0387] Amplitude transmittance T z The larger the difference between (r) and the amplitude transmittance of the specimen 140, the larger the estimated specimen image I estz (r) and image I meaz The difference with (r) also becomes large. Therefore, the estimated specimen image I estz (r) and image I meaz The difference between (r) and (r) is the amplitude transmittance T z This can be considered to reflect the difference between (r) and the amplitude transmittance of the sample 140.

[0388] As mentioned above, zL = 1. Therefore, in equation (38), zL = 1 is set, and the estimated sample image Iest1 (r) and image I mea1 Using (r), the wavefront u 1 m As a result, as shown in FIG. 31(c), the corrected wavefront, i.e., the wavefront u' 1 m (r) is obtained.

[0389] Wavefront u' 1 m Using (r), we can calculate the new amplitude transmittance. 1 m (r) is the wavefront u 1 m (r). Therefore, the new amplitude transmittance is 1 m This differs from the amplitude transmittance when (r) was calculated.

[0390] In step S312, the corrected wavefront g' out m,zL (r) is calculated. Wavefront g' out m,zL (r) is the wavefront u' zL m (r) is the wavefront when it propagates by ΔD. Wavefront g' out m,zL (r) is expressed by equation (39).

number

[0391] As mentioned above, the wavefront u' 1 m Using (r), the amplitude transmittance T z (r) can be calculated. However, as shown in FIG. 31, the amplitude transmittance T z To calculate (r), the wavefront g out m The wavefront at position (r) is required.

[0392] As shown in Figures 31(a) and 31(c), the wavefront u' 1 m The position of (r) and the wavefront g outm The position of (r) is different. Therefore, the amplitude transmittance T z In order to calculate (r), the wavefront g' is out m,1 (r) is necessary.

[0393] wave surface g' out m,1 (r) is the wavefront u' 1 m (r) is the wavefront when it propagates by 3×Δz. In equation (39), by setting ΔD=3×Δz and zL=1, the wavefront g' out m,1 (r) can be calculated.

[0394] In step S313, the gradient ΔT z m,zL (r) is calculated. ΔT z m,zL (r) is the gradient of the specimen when illuminated by the mth light source and corrected with the photographed image and the estimated specimen image at the position of the layer indicated by the value of the variable zL.

[0395] Sample gradient ΔT z m,zL is expressed by equation (40). The gradient of the sample ΔT z m,zL To calculate (r), for example, the gradient descent method can be used.

number

[0396] As described above, the estimated specimen 150 is displaced into a number of thin layers. Thus, for each thin layer, the gradient of the specimen, ΔT z m,zL (r) needs to be calculated.

[0397] Figure 32 shows the gradient of the specimen and the propagation of the wavefront. If the estimated specimen 150 is replaced by four thin layers, i.e. N IM = 4. Also assume that zL = 1. Fig. 32(a) is a diagram showing the gradient of the sample. Fig. 32(b) is a diagram showing the propagation of the wavefront.

[0398] wave surface g out m To calculate (r), the amplitude transmittance T 4 (r) is used. The amplitude transmittance T 4 (r) is the estimated amplitude transmittance. Therefore, this amplitude transmittance T 4 (r) is different from the amplitude transmittance of the sample 140.

[0399] Amplitude transmittance T 4 The greater the difference between (r) and the amplitude transmittance of the sample 140, the greater the wavefront g out m (r) and wavefront g' out m,1 The difference with (r) also becomes large. Therefore, the wavefront g out m (r) and wavefront g' out m,1 The difference between (r) and (r) is the amplitude transmittance T 4 This can be considered to reflect the difference between (r) and the amplitude transmittance of the sample 140.

[0400] Wavefront f 4 m (r), amplitude transmittance T 4 (r), wavefront g out m (r), and wavefront g' out m,1 (r) is known. Therefore, by setting z = 4 and zL = 1 in equation (40), the gradient ΔT 4 m,1 (r) can be calculated.

[0401] g 4 m (r) and wavefront g out m (r) is the same, so g 4m Instead of (r), wavefront g out m (r) can be used. Also, g' 4 m,1 (r) is g' out m,1 Since it is the same as (r), g' 4 m,1 Instead of (r), g' out m,1 Just use (r).

[0402] Next, the gradient of the specimen ΔT 3 m,1 Calculate (r). The gradient of the sample ΔT 3 m,1 To calculate (r), the wavefront g 3 m The wavefront at the position (r) is required. To calculate this wavefront, the wavefront f' is calculated as shown in FIG. 4 m,1 (r) is necessary.

[0403] Wavefront f' 4 m,1 (r) can be calculated by setting z = 4 and zL = 1 in equation (41).

number

[0404] Next, the calculated wavefront f' 4 m,1 Using (r), wavefront g 3 m,1 The wavefront at position (r) is calculated.

[0405] Figure 33 shows the sample gradient and the propagation of the wavefront. Figure 33(a) shows the sample gradient and the propagation of the wavefront. Figure 33(b) shows the wavefront propagation.

[0406] As shown in FIG. 33(a), the wavefront g' 3 m,1 (r) is the wavefront f' 4 m,1(r) is the wavefront when it propagates by Δz. This is the propagation of the wavefront from the fourth layer to the third layer.

[0407] As described above, the propagation of the wavefront from the third layer to the fourth layer is expressed by equation (32). Therefore, by changing equation (32) as follows, the wavefront g' 3 m,1 (r) can be calculated. Wavefront f 4 m (r) is the wavefront g' 3 m,1 Replace with (r). wave surface g 3 m (r) is the wavefront f' 4 m,1 Replace with (r). Let ΔD = -Δz.

[0408] Wavefront f 3 m (r), amplitude transmittance T 3 (r), wavefront g 3 m (r), and wavefront g' 3 m,1 (r) is known. Therefore, by setting z = 3 and zL = 1 in equation (40), the gradient ΔT 3 m,1 (r) can be calculated.

[0409] Wavefront f' 3 m,1 (r) can be calculated by setting z = 3 and zL = 1 in equation (41).

[0410] The gradient of the sample for the second and first layers can be calculated in the same manner as for the third layer.

[0411] FIG. 34 is a diagram showing the gradient of the specimen. In FIG. 34, the gradient ΔT 1 m,1 (r), Gradient ΔT of the specimen in the second layer 2 m,1 (r) The gradient ΔT of the specimen in the third layer 3m,1 (r), and the gradient ΔT of the specimen in the fourth layer 4 m,1 (r), has been calculated.

[0412] The gradient ΔT of the sample obtained in step S313 z m,1 (r) is the gradient of the specimen when illuminated by the mth light source and corrected with the captured image at the position of the first layer and the estimated image of the specimen at the position of the first layer. The gradient of the specimen ΔT z m,1 (r) is determined by the illumination light emitted from all light sources. Therefore, for all light sources, the gradient of the specimen ΔT z m,1 (r) must be sought.

[0413] In step S314, the value of the variable m is set to the number of light sources N LS If the result of the determination is NO, step S315 is executed. If the result of the determination is YES, step S304 is executed.

[0414] (If the result is NO: m ≠ N LS ) If the determination result is NO, in step S315, the value of the variable m is incremented by 1. When step S315 ends, the process returns to step S311.

[0415] In step S315, the value of the variable m is incremented by 1. Therefore, for another light source, the wavefront u z ' m,1 (r) is calculated, and in step S312, the wavefront g out ' m,1 (r) is calculated, and in step S313, the gradient ΔT z m,1 (r) is calculated.

[0416] Steps S311, S312, and S313 calculate the gradient ΔT of the sample for all light sources. z m,1 This is repeated until (r) is found.

[0417] FIG. 35 shows the gradient of the specimen. In FIG. 35, the gradient of the specimen ΔT z m.1 (r) is required.

[0418] (If the answer is YES: m=N LS ) If the result of the determination is YES, in step S304, the amplitude transmittance T z (r) is updated. Step S304 is a step of updating the estimation sample.

[0419] Updated amplitude transmission T z (r) is expressed by equation (42).

number

[0420] In step S305, the value of the variable L is set to the number of corrections N CR If the result of the determination is NO, step S306 is executed. If the result of the determination is YES, step S30 is executed.

[0421] (If the result is NO: m ≠ N CR ) If the determination result is NO, in step S306, 1 is added to the value of the variable L. When step S306 ends, the process returns to step S301.

[0422] In step S301, 1 to N IM One of the numbers is randomly selected from the range 1 to 10. Based on the selected number, the estimated specimen image and acquisition position used for correction are determined.

[0423] Then, in step S311, the wavefront u z ' m,1 (r) is calculated, and in step S312, the wavefront g out ' m,1(r) is calculated, and in step S313, the gradient ΔT z m,1 (r) is calculated, and in step S304, the amplitude transmittance T z (r) is updated.

[0424] Steps S301, S302, S303, S304, and S310 are repeated until the set number of corrections have been completed.

[0425] (If the answer is YES: m=N CR ) If the result of the determination is YES, the process returns to step S30. z At (r), steps S30 to S300 are executed.

[0426] By repeatedly executing steps S30 to S300, the updated amplitude transmittance T s (r) gradually approaches the amplitude transmittance of the sample 140. That is, the residual becomes smaller. Eventually, the residual becomes smaller than the threshold value.

[0427] (If the result is YES: Residual < Threshold) In step S110, the estimated refractive index distribution of the specimen is calculated. The obtained amplitude transmittance T z (r) is the same as or approximately the same as the amplitude transmittance of the sample 140. The obtained amplitude transmittance T z From (r) and equation (1), the refractive index distribution n z (r) is found.

[0428] The refractive index distribution n obtained in step S110 z Using (r), the structure of the estimated specimen can be reconstructed. The reconstructed structure of the estimated specimen can be output to, for example, a display device. The estimated specimen 150 is a thick specimen. In the second simulation, the structure of the thick specimen can be used to reconstruct the three-dimensional configuration of the estimated specimen.

[0429] As described above, the amplitude transmittance T obtained in step S110 z (r) is equal to or approximately equal to the amplitude transmittance of the sample 140. In this case, the refractive index profile n z (r) can also be considered to be identical or nearly identical to the refractive index profile of the sample 140. Therefore, the structure of the reconstructed estimated sample 150 can be considered to be identical or nearly identical to the structure of the sample 6.

[0430] In the second simulation, steps S250, S270, and S310 are repeatedly executed. As a result, the amplitude transmittance T z (r) is updated. As described above, steps S250 and S270 are steps for calculating the image of the estimated specimen. Step S310 is a step for optimizing the refractive index distribution of the estimated specimen.

[0431] Amplitude transmittance T z (r) represents the estimated specimen. Therefore, the step of calculating the image of the estimated specimen and the step of optimizing the refractive index distribution of the estimated specimen are repeatedly performed, thereby updating the estimated specimen.

[0432] Figure 36 shows the results of the simulation of the first example. Figure 36(a) shows the specimen. Figure 36(b) shows the aperture member. Figure 36(c) shows an image of the specimen. Figure 36(d) shows the reconstructed estimated specimen.

[0433] The specimen is a photonic crystal fiber (hereafter referred to as "PCF"). In PCF, multiple through holes are formed inside the cladding. The through holes are called the core. The PCF is immersed in liquid. Therefore, the core is filled with liquid.

[0434] The values ​​of the various parameters are as follows: Specimen outer diameter 230μm Core diameter 6μm Core refractive index 1.466 Cladding refractive index 1.462 Refractive index of liquid: 1.466 Objective lens numerical aperture: 1.4 Magnification of objective lens: 60x Objective lens focal length: 3mm Numerical aperture of second region: 0.3, 1.25 Condenser lens focal length 7mm Diameter of the aperture material's transmission area: 0.2 mm Illumination wavelength: 0.7μm

[0435] The numerical aperture of the second region is expressed by the numerical aperture of the condenser lens. As shown in Figure 36(b), the second regions are located on the circumference of two circles. The light beam emitted from the second region located on the inner circumference can be considered to have a numerical aperture of 0.3. The light beam emitted from the second region located on the outer circumference can be considered to have a numerical aperture of 1.25.

[0436] The parameter values ​​in each conditional expression are as follows: Detection optics pupil area (PS') 55.4mm 2 Area of ​​second region (LS') 0.0058mm 2 LS' / PS' 1.0×10 -4 Detection optics pupil diameter (T') 8.4mm Distance between two adjacent second regions (d') 0.47 mm d' / T' 0.056

[0437] Detection optics pupil area (8.4 / 2) 2 ×3.14=55.4 Area of ​​the second region (0.086 / 2) 2 ×3.14=0.0058 Diameter of the second region: 0.2 / 7×3=0.086 Pupil diameter of detection optical system: 2×1.4×3=8.4 Radius to the second inner region: 0.3×3=0.9 Distance between two adjacent second regions 0.9×sin30 / cos15=0.47

[0438] As shown in Figure 36(b), the second regions are located on the circumference of two circles. The radius to the second region located on the inside is the radius of the inner circle.

[0439] Figure 37 shows the aperture material and the reconstructed estimated specimen. Figures 37(a), 37(b), 37(c), and 37(d) show the aperture material. Figures 37(e), 37(f), 37(g), and 37(h) show the reconstructed estimated specimen.

[0440] The estimated specimen was reconstructed using an objective lens with a numerical aperture of 1.4 and an illumination light wavelength of 0.7 μm.

[0441] The correspondence between the opening material in each figure and the reconstructed estimated specimen is as follows: Reconstructed specimen of opening Fig. 37(a) Fig. 37(e) Fig. 37(b) Fig. 37(f) Fig. 37(c) Fig. 37(g) Fig. 37(d) Fig. 37(g)

[0442] In the aperture material shown in Figure 37(a), the numerical aperture of the second region is 0.3 and 0.5. As shown in Figure 37(e), in the reconstructed estimated specimen, the core extends in the Z-axis direction. In the aperture material shown in Figure 37(a), it is difficult to estimate the fine structure.

[0443] In the aperture member shown in Fig. 37(b), the numerical aperture of the second region is 1.05 and 1.25. As shown in Fig. 37(f), there are artifacts in the reconstructed estimated specimen. Thin images of the core are present on the left and right.

[0444] In the aperture member shown in Fig. 37(c), the numerical aperture of the second region is 0.3, 0.5, 1.05, and 1.25. As shown in Fig. 37(g), artifacts are present in the reconstructed estimated specimen. However, the artifacts are reduced compared to Fig. 37(f).

[0445] For the aperture materials shown in Figure 37(d), the numerical apertures of the second region are 0.3, 0.5, 1.05, and 1.25, but two aperture materials are used. As shown in Figure 37(h), there are artifacts in the reconstructed estimated specimen, but the artifacts are the same as those in Figure 37(g).

[0446] Comparing Figure 37(e) and Figure 37(f), the larger the numerical aperture of the second region, the finer the structure is reconstructed. Also, comparing Figure 37(f) and Figure 37(g), the more the number of second regions, the fewer the artifacts. However, if the number of second regions is large, the calculation in reconstruction becomes difficult to converge.

[0447] As shown in Fig. 37(d), by using two aperture members, the ratio of relatively coarse structure estimation and relatively fine structure estimation can be adjusted in the reconstruction, and the reconstruction time can be shortened by making such adjustments.

[0448] Furthermore, when the aperture material shown in FIG. 37(d) is used, it is possible to estimate only a relatively coarse structure, or to estimate only a relatively fine structure, compared to when the aperture material shown in FIG. 37(c) is used.

[0449] Figure 38 shows the aperture member and the image of the specimen in the measurement of the second example. Figure 38(a) shows the aperture member. Figure 38(b) and Figure 38(c) show the images of the specimen.

[0450] The specimen is a lattice-like structure. The specimen is immersed in oil with a refractive index of 1.518. Therefore, the space surrounded by the lattice is filled with oil.

[0451] The values ​​of the various parameters are as follows: Objective lens numerical aperture: 1.42 Magnification of objective lens: 60x Objective lens focal length: 3mm Numerical aperture of the second region: 0.3, 0.45 Condenser lens focal length 7.1mm Diameter of the aperture material's transmission area: 0.2 mm

[0452] The numerical aperture of the second region is expressed by the numerical aperture of the condenser lens. As shown in Fig. 37(a), the second regions are located on the circumference of two circles. The light beam emitted from the second region located on the inner circumference can be considered to have a numerical aperture of 0.3. The light beam emitted from the second region located on the outer circumference can be considered to have a numerical aperture of 0.45.

[0453] The parameter values ​​in each conditional expression are as follows: Detection optics pupil area (PS') 57.0mm 2 Area of ​​second region (LS') 0.0056mm 2 LS' / PS' 9.8×10 -5 Detection optics pupil diameter (T') 8.52mm Distance between two adjacent second regions (d') 0.47 mm d' / T' 0.055

[0454] Detection optics pupil area (8.52 / 2) 2 ×3.14=57.0 Area of ​​the second region (0.0845 / 2) 2 ×3.14=0.0056 Diameter of the second region: 0.2 / 7.1×3=0.0845 Pupil diameter of detection optical system: 2×1.42×3=8.52 Radius to the second inner region: 0.3×3=0.9 Distance between two adjacent second regions 0.9×sin30 / cos15=0.47

[0455] As shown in Figure 37(a), the second regions are located on the circumference of two circles. The radius to the second region located on the inside is the radius of the inner circle.

[0456] Figure 39 shows the results of the second example. Figures 39(a) and 39(b) show the estimation samples with the initial values ​​set. Figures 39(c), 39(d), and 39(e) show the reconstructed estimation samples.

[0457] The correspondence between the initial aperture material and the reconstructed estimated specimen in each figure is as follows: Initialized estimation sample Reconstructed estimation sample Fig. 39(a) Fig. 39(c) Fig. 39(b) Fig. 39(d) None Figure 39(e)

[0458] In Fig. 39(a), the initial values ​​are set so that the contour of the estimated specimen can be distinguished, and in Fig. 39(b), the initial values ​​are set using the transport of intensity equation.

[0459] From a comparison between FIG. 37(c) and FIG. 37(e) and a comparison between FIG. 37(d) and FIG. 37(e), it is clear that setting the initial value enables the estimated sample to be reconstructed with higher accuracy.

[0460] The microscope system 1 shown in Fig. 1(a) may include an illumination optical system. In this case, it is preferable that the microscope system 1 satisfies conditions (6) and (7). The microscope system 1 may also include an aperture member. [Industrial Applicability]

[0461] As described above, the present invention is suitable for a microscope system in which the time from the start of image acquisition to the completion of object model estimation is short. [Explanation of symbols]

[0462] 1. Microscope system 2 Incoherent light source 3. Detection optics 4. Image sensor 5 specimens 5a surface 5' optical image 10 Pupil plane of detection optical system 3 11, 12, 20, 21 1st area 30 First Circular Region 31 Second Circular Region 32 The third circular region 40, 41 1st area 50 First Circle Area 51 Second circular area 52 Fourth Circular Region 53 Third Circle Area 60 1st area 70 First sector area 71 Second sector 72 Third sector 74 First sector area 80, 81 1st area 90 Microscope System 91 Illumination optical system 100 Pupil plane of illumination optical system 101, 102 2nd area 110 Microscope System 111 Opening material 112 processors 113 Shading area 112a, 112b, 112c, 112d transmission area 120 Microscope System 121 Incoherent Light Source 122 Illumination optical system 130 Estimated sample 130a surface 140 specimens 140' optical image 141 Moving Stage 150 Estimated sample 150a surface L1, L2 luminous flux Fo: focal position of the detection optical system Pu pupil position of detection optical system IP image plane C Center of pupil of detection optical system IP image plane PI Pupil plane of illumination optical system

Claims

1. The system includes an incoherent light source, a detection optical system, and an image sensor, The incoherent light source is a light source that emits light that is not coherent in time, The detection optical system is an optical system that forms an optical image of a sample, The imaging element receives an optical image of the sample formed by the detection optical system, A plurality of coherent illuminations are simultaneously performed on the sample by the light emitted from the incoherent light source; The coherent illumination is illumination with light that is spatially coherent; The direction in which the light beam is irradiated onto the sample is different for each of the coherent illuminations, In a pupil plane of the detection optical system, the light beams of the coherent illuminations pass through different first regions, Each of the first regions satisfies the following condition (1): A microscope system, characterized in that at least one of the distances between two adjacent first regions satisfies the following condition (2). LS<PS×10 －３ (1) 0.05×T<d (2) Where: LS is the area of ​​the first region (unit: mm ２ ), PS is the area of ​​the pupil of the detection optical system (unit: mm ２ ), d is the distance between two adjacent first regions (unit: mm), T is the diameter of the pupil of the detection optical system (unit: mm), It is.

2. The microscope system according to claim 1 , wherein half of the first regions satisfy the condition (2).

3. The microscope system according to claim 1, characterized in that the following condition (3) is satisfied: [0010] Where: LSi is the area of ​​the i-th first region (unit: mm ２ ), n is the number of first regions, It is.

4. some of the first regions are located within a first annular region; 2. The microscope system according to claim 1, wherein the first annular region is a region having a radius of 50% or more of a pupil region of the detection optical system.

5. 5. The microscope system according to claim 4, wherein some of the first regions are arranged in the first annular region so as to form a double circle.

6. some of the first regions are located within a second annular region; 5. The microscope system according to claim 4, wherein the second annular region is a region having a radius of 70% to 90% of a pupil region of the detection optical system.

7. some of the first regions are located within a third annular region; 7. The microscope system according to claim 6, wherein the third annular region is a region having a radius of 50% to 70% of a pupil region of the detection optical system.

8. Some of the first regions are located within a first circular region; 5. The microscope system according to claim 4, wherein the first circular area is a pupil area of ​​the detection optical system that is closer to a center than the first annular area.

9. 9. The microscope system according to claim 8, wherein some of the first regions are arranged to form a circle within the first circular region.

10. some of the first regions are located within a second circular region; 2. The microscope system according to claim 1, wherein the second circular area is an area having a radius of 50% or less of a pupil area of ​​the detection optical system.

11. The microscope system according to claim 10 , wherein some of the first regions are arranged in the second circular region so as to form a circle.

12. some of the first regions are located within a fourth annular region; 11. The microscope system according to claim 10, wherein the fourth annular region is a region having a radius of 30% to 50% of a pupil region of the detection optical system.

13. some of the first regions are located within a third circular region; 13. The microscope system according to claim 12, wherein the third circular area is an area having a radius of 30% or less of a pupil area of ​​the detection optical system.

14. 2. The microscope system according to claim 1, wherein when a pupil of the detection optical system is divided into four sectors having equal central angles, any one of the first regions is located in each of the four sectors.

15. 2. The microscope system according to claim 1, wherein some of the first regions are paired on either side of the center of a pupil of the detection optical system.

16. The microscope system according to claim 1 , wherein each of the first regions satisfies the following condition (4): PS×10 －６ <LS (4)

17. 2. The microscope system according to claim 1, wherein at least one of the intervals between two adjacent first regions satisfies condition (2) and the following condition (5). d<0.5×T (5)

18. Further comprising an illumination optical system; on a pupil plane of the illumination optical system, the light beams of the coherent illuminations are located in second regions different from each other, Each of the second regions satisfies the following condition (6): The microscope system according to claim 1 , wherein at least one of the intervals between two adjacent second regions satisfies condition (2) and the following condition (7). LS’<PS’×10 －３ (6) 0.05×T'<d' (7) Where: LS' is the area of ​​the second region (unit: mm ２ ), PS' is the pupil area of ​​the illumination optical system (unit: mm ２ ), d' is the distance between two adjacent second regions (unit: mm), T' is the pupil diameter of the illumination optical system (unit: mm), It is.

19. Further comprising an opening member, The light beams of the respective coherent illuminations are emitted from a plurality of independent regions on a predetermined surface, the predetermined surface is a surface perpendicular to an optical axis of the detection optical system and is located on the opposite side of the detection optical system with respect to the specimen, the aperture member is disposed on the predetermined surface and includes a plurality of independent transmission areas, the transmission areas being areas that transmit light, The microscope system according to claim 1 , wherein each of the transmission regions corresponds to each of the first regions.

20. The light beams of the respective coherent illuminations are emitted from a plurality of independent regions on a predetermined surface, the predetermined surface is a surface perpendicular to an optical axis of the detection optical system and is located on the opposite side of the detection optical system with respect to the specimen, The incoherent light source is arranged in a plurality of parts on the predetermined plane, The microscope system according to claim 1 , wherein each of the incoherent light sources corresponds to each of the first regions.

21. The system includes an incoherent light source, an illumination optical system, a detection optical system, and an image sensor, The incoherent light source is a light source that emits light that is not coherent in time, The detection optical system is an optical system that forms an optical image of a sample, The imaging element receives an optical image of the sample formed by the detection optical system, A plurality of coherent illuminations are simultaneously performed on the sample by the light emitted from the incoherent light source; The coherent illumination is illumination with light that is spatially coherent; The direction in which the light beam is irradiated onto the sample is different for each of the coherent illuminations, on a pupil plane of the illumination optical system, the light beams of the coherent illuminations are located in second regions different from each other, Each of the second regions satisfies the following condition (6): A microscope system, characterized in that at least one of the intervals between two adjacent second regions satisfies the following condition (7). LS’<PS’×10 －３ (6) 0.05×T'<d' (7) Where: LS' is the area of ​​the second region (unit: mm ２ ), PS' is the pupil area of ​​the illumination optical system (unit: mm ２ ), d' is the distance between two adjacent second regions (unit: mm), T' is the pupil diameter of the illumination optical system (unit: mm), It is.

22. the detection optical system includes an objective lens and an imaging lens; the illumination optical system includes a condenser lens; The area of ​​the second region is expressed by the following formula (8):

22. The microscope system according to claim 21, wherein a pupil diameter of the illumination optical system is expressed by the following equation (9): PS’=(FLcd×NA) ２ ×π (8) T'=FLcd×NA (9) Where: FLcd is the focal length of the condenser lens (unit: mm), NA is the numerical aperture of the objective lens, It is.

23. Further comprising an opening member, Each of the plurality of light beams is emitted from each of a plurality of independent regions on a predetermined surface, the predetermined surface is a surface perpendicular to an optical axis of the detection optical system and is located on the opposite side of the detection optical system with respect to the specimen, the aperture member is disposed on the predetermined surface and includes a plurality of independent transmission areas, the transmission areas being areas that transmit light, The microscope system according to claim 21 , wherein each of the transmission regions corresponds to each of the second regions.

24. Each of the plurality of light beams is emitted from each of a plurality of independent regions on a predetermined surface, the predetermined surface is a surface perpendicular to an optical axis of the detection optical system and is located on the opposite side of the detection optical system with respect to the specimen, The incoherent light source is arranged in a plurality of parts on the predetermined plane, The microscope system according to claim 21 , wherein each of the incoherent light sources corresponds to each of the second regions.

25. Further comprising a processor, The processor, A wavefront passing through an estimated sample obtained by modeling the sample is obtained for each of the light beams by a forward propagation calculation; calculating an intensity distribution at an image-forming position of the detection optical system corresponding to the wavefront for each of the light beams; generating a calculated image by adding up the intensity distributions for each of the light beams; 22. The microscope system according to claim 1, wherein the estimated specimen is reconstructed by performing an optimization process that reduces a difference between the calculated image and a measurement image output from the image sensor.

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