JP2015099177A - Structured illumination device and structured illumination microscope device - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a structured illumination device which can control the ratio between components of orders of structured illumination.SOLUTION: A structured illumination device comprises an optical system that forms an interference fringe by a predetermined number of light beams on a sample. The optical system comprises a spatial light modulation part that controls polarization of at least one light beam of the predetermined number of light beams, and a polarization member that extracts, from the light beam whose polarization direction is controlled, a component in a predetermined polarization direction.

Description

  The present invention relates to a structured illumination apparatus and a structured illumination microscope apparatus.

  In recent years, a plurality of microscopy methods exceeding the resolution limit of an optical microscope have been proposed, and they are collectively referred to as a super-resolution optical microscope. One type of super-resolution optical microscope is a structured illumination microscope. This microscope projects a striped stripe pattern (structured illumination) onto the sample or sample surface to be observed (sample surface or sample surface), and the fluorescence (or scattering, etc.) emitted from the sample is excited by the projection. ) Is acquired by the image sensor. In order to construct a super-resolution image, it is necessary to acquire a plurality of images excited by structured illumination with different phases. By analyzing these multiple images, a super-resolution image exceeding the resolution limit of the imaging optical system for observation is obtained. In order to achieve super-resolution in a two-dimensional plane, it is necessary to change the direction of structured illumination.

  By projecting the structured illumination onto the specimen surface, the spatial frequency of the structured illumination and the spatial frequency of the specimen generate moire fringes. This moire fringe includes spatial frequency information of a sample that has been frequency-converted to a low spatial frequency and exceeds the resolution limit of the imaging optical system. If the spatial frequency of the moire fringes is lower than the spatial frequency at the resolution limit of a normal imaging optical system, the information can be detected by the imaging optical system. Therefore, it is possible to achieve super-resolution by acquiring an image including information on the moire fringe, acquiring a plurality of images by changing the phase of structured illumination, and performing arithmetic processing using them. (For example, refer to Patent Document 1).

US Reissue Patent No. 38307 Specification

  However, when the structured illumination described above is used, there is a problem that the ideal ratio between the components of the light components of the respective orders included in the structured illumination differs depending on the specimen.

  The present invention has been made in view of such circumstances, and provides a structured illumination apparatus and a structured illumination microscope apparatus capable of controlling the ratio between components of light components of respective orders included in structured illumination. provide.

  The present invention has been made to solve the above-described problems, and one aspect of the present invention is a structured illumination apparatus including an optical system that forms interference fringes by a predetermined number of light beams on a specimen. The system includes a spatial light modulator that controls polarization of at least one of the predetermined number of light beams, and a polarization member that extracts a component in a predetermined polarization direction from the light beams emitted from the spatial light modulator. A structured lighting device.

  Another aspect of the present invention is a structured illumination microscope apparatus including the structured illumination apparatus described above and an imaging unit that acquires a modulated image that is an image of the sample spatially modulated by the interference fringes.

  According to the present invention, it is possible to control the ratio between the components of the light components of the respective orders included in the structured illumination.

1 is a configuration diagram of a structured illumination microscope apparatus 1 according to an embodiment of the present invention. It is a figure explaining the light beam branching part 14 which concerns on the same embodiment. It is the schematic which shows the structure of the light quantity adjustment part 18 which concerns on the same embodiment. It is a figure which shows the control unit of 18 A of spatial light modulation parts which concern on the embodiment. It is a figure explaining the orientation of the molecule | numerator of the ferroelectric liquid crystal of 18 A of spatial light modulation parts which concern on the embodiment. It is FIG. (1) explaining adjustment of the light quantity by the light quantity adjustment part 18 which concerns on the embodiment. It is FIG. (2) explaining adjustment of the light quantity by the light quantity adjustment part 18 which concerns on the embodiment. It is FIG. (1) explaining the light beam selection part 19 which concerns on the embodiment. It is FIG. (2) explaining the light beam selection part 19 which concerns on the same embodiment. It is a figure explaining the function of the light beam selection part 19 concerning the embodiment. It is a figure explaining light beam selection member 19B concerning the embodiment. It is a figure explaining operation | movement of the translation mechanism 15 of the light beam branching part 14 which concerns on the same embodiment. It is a figure explaining control by control device 43 concerning the embodiment.

[First Embodiment]
Hereinafter, a structured illumination microscope apparatus will be described as an embodiment of the present invention.

  FIG. 1 is a configuration diagram of the structured illumination microscope apparatus 1. The structured illumination microscope apparatus 1 according to the present embodiment is a structured illumination microscope of 3D-SIM (Structured Illumination Microscopy) mode. The structured illumination microscope apparatus 1 according to the present embodiment can control the ratio between the zero-order component and the first-order and second-order components of the optical transfer function (OTF) of the illumination system.

  First, the structure of the structured illumination microscope apparatus 1 will be described.

  As shown in FIG. 1, the structured illumination microscope apparatus 1 includes a laser unit 100, an optical fiber 11, an illumination optical system 10, an imaging optical system 30, an imaging element 42, a control device 43, an image storage / An arithmetic device 44 and an image display device 45 are provided. The illumination optical system 10 is an epi-illumination type, and the sample 2 is illuminated using the objective lens 31 and the dichroic mirror 33 of the imaging optical system 30.

  The laser unit 100 includes a first laser light source 101, a second laser light source 102, shutters 103 and 104, a mirror 105, a dichroic mirror 106, and a lens 107. Each of the first laser light source 101 and the second laser light source 102 is a light source that emits coherent laser light, and the emission wavelengths thereof are different from each other. Here, the wavelength λ1 of the first laser light source 101 is longer than the wavelength λ2 of the second laser light source 102 (λ1> λ2). The first laser light source 101, the second laser light source 102, and the shutters 103 and 104 are driven by the control device 43, respectively.

  The optical fiber 11 is configured by, for example, a polarization-preserving single mode fiber in order to guide the laser light emitted from the laser unit 100. The position of the output end of the optical fiber 11 in the direction of the optical axis O can be adjusted by the position adjusting mechanism 11A. The position adjusting mechanism 11A is driven and controlled by the control device 43. For example, a piezo element or the like is used as the position adjusting mechanism 11A.

  When a polarization plane preserving single mode fiber is used as the optical fiber 11, the polarization plane of the laser light is preserved before and after the optical fiber 11. This is effective for maintaining the quality of polarized light. On the other hand, when a multimode fiber is used as the optical fiber 11, the polarizing plate 13 is essential.

  The illumination optical system 10 includes, in order from the emission end side of the optical fiber 11, a collector lens 12, a polarizing plate 13, a light beam branching unit 14, a condensing lens 17, a light amount adjusting unit 18, and a light beam selecting unit 19. The lens 21, the field stop 22, the field lens 23, the excitation filter 24, the dichroic mirror 33, and the objective lens 31 are arranged.

  The light beam splitting unit 14 includes a translation mechanism 15 and a diffractive optical element (diffraction grating) 16. The light beam selection unit 19 includes a half-wave plate 19A, a light beam selection member 19B, and a rotation mechanism 19C. And are provided. Each of the light beam branching unit 14 and the light beam selecting unit 19 is driven by the control device 43.

  In the imaging optical system 30, an objective lens 31, a dichroic mirror 33, an absorption filter 34, and a second objective lens 35 are arranged in order from the sample 2 side.

  Specimen 2 is, for example, fluorescent cells (cells stained with a fluorescent dye) arranged on parallel flat glass surfaces, or fluorescent living cells (moving cells stained with fluorescent dyes) present in a petri dish. ) And so on. In this cell, both the first fluorescent region excited by light of wavelength λ1 and the second fluorescent region excited by light of wavelength λ2 are expressed.

  The image sensor 42 is a two-dimensional image sensor composed of a CCD, a CMOS, or the like. When the image sensor 42 is driven by the control device 43, the image sensor 42 captures an image formed on the imaging surface 41 and generates an image. This image is taken into the image storage / arithmetic unit 44 via the control unit 43. The frame period (imaging repetition period) of the image sensor 42 depends on the rate-determining among the imaging time of the image sensor 42 (that is, the time required for charge accumulation and charge readout), the time required for switching the direction of interference fringes, and other required times. Determined.

The control device 43 drives and controls the laser unit 100, the position adjustment mechanism 11A, the light beam branching unit 14, the light amount adjusting unit 18, the light beam selecting unit 19, and the image sensor 42. The control device 43 accepts the setting of the ratio C ₁ of the primary component to the zero-order component of the optical transfer function (OTF) of the illumination optical system 10 and the ratio C ₂ of the secondary component to the zero-order component. Based on the ratios C ₁ and C ₂ and the wavelength λ of the laser light emitted from the laser unit 100, a control signal S3 for controlling the light amount adjusting unit 18 is generated. Details of these controls will be described later. Further, the ratios C ₁ and C ₂ are set in advance, and the control device 43 generates the control signal S3 so that the ratios C ₁ and C ₂ are always maintained even when the wavelength λ of the laser light changes. You may make it do.
The image storage / arithmetic unit 44 performs a calculation on the image given via the control unit 43, stores the calculated image in an internal memory (not shown), and sends it to the image display unit 45.

  Next, the behavior of laser light in the structured illumination microscope apparatus 1 will be described. When the laser beam (first laser beam) emitted from the first laser light source 101 enters the mirror 105 through the shutter 103, it is reflected by the mirror 105 and enters the dichroic mirror 106. On the other hand, laser light (second laser light) having a wavelength λ2 emitted from the second laser light source 102 enters the dichroic mirror 106 through the shutter 104 and is integrated with the first laser light. The first laser beam and the second laser beam emitted from the dichroic mirror 106 enter the incident end of the optical fiber 11 through the lens 107. When the control device 43 controls the laser unit 100, the wavelength of the laser light (= use wavelength λ) incident on the incident end of the optical fiber 11 is switched between the long wavelength λ1 and the short wavelength λ2.

  The laser light incident on the incident end of the optical fiber 11 propagates inside the optical fiber 11 and generates a point light source at the output end of the optical fiber 11. The laser light emitted from the point light source is converted into a parallel light beam by the collector lens 12 and is incident on the diffraction grating 16 of the light beam branching section 14 via the polarizing plate 13 and is branched into diffracted light beams of respective orders. These orders of diffracted light beams (hereinafter referred to as “diffracted light beam groups”) are collected by the condenser lens 17 at different positions on the pupil conjugate plane 25.

  Here, the pupil conjugate plane 25 is the focal position (rear focal position) of the lens 17, and the lens 23 and the lens 21 are arranged with respect to the pupil 32 of the objective lens 31 (the position where the ± first-order diffracted light is condensed). It is a conjugate position. The lens 17 is disposed so that the focal position (back focal position) of the lens 17 coincides with the pupil conjugate plane 25. Note that the concept of “conjugate position” includes a position determined by a person skilled in the art in consideration of design-related matters such as aberration and vignetting of the objective lens 17 and the lenses 21 and 23.

  Since the laser light emitted from the optical fiber 11 is basically linearly polarized, the polarizing plate 13 can be omitted, but it is effective for reliably cutting off the extra polarized component. Further, in order to increase the utilization efficiency of the laser light, it is desirable that the axis of the polarizing plate 13 coincides with the polarization direction of the laser light emitted from the optical fiber 11.

  The diffracted light beams of respective orders toward the pupil conjugate plane 25 enter the light amount adjusting unit 18. The light amount adjusting unit 18 adjusts the light amount of the 0th-order diffracted light beam and the light amounts of the 1st-order and −1st-order diffracted light beams in accordance with the control signal S3 from the control device 43. Here, the light amount is an average light amount during which one modulated image is captured by the image sensor 42. Each diffracted light beam whose light amount has been adjusted by the light amount adjusting unit 18 enters a light beam selecting unit 19 disposed in the vicinity of the pupil conjugate plane 25. The light beam selection unit 19 selectively passes a 0th-order diffracted light beam and a pair of diffracted light beams (± 1st-order diffracted light beams) among the incident diffracted light beams of each order.

  The 0th-order diffracted light beam and the ± 1st-order diffracted light beam that have passed through the light beam selection unit 19 form a conjugate plane with the diffraction grating 16 near the field stop 22 by the lens 21. Thereafter, each of the 0th-order diffracted light beam and the ± 1st-order diffracted light beam is converted into convergent light by the field lens 23, further passes through the excitation filter 24, is reflected by the dichroic mirror 33, and is mutually reflected on the pupil plane 32 of the objective lens 31. Concentrate at different positions.

  Each of the 0th-order diffracted light beam and the ± 1st-order diffracted light beam collected on the pupil surface 32 becomes a parallel light beam when exiting from the tip of the objective lens 31 and interferes with each other on the surface of the sample 2 to form interference fringes. Form. This interference fringe is used as structured illumination light.

  When the specimen 2 is illuminated with such structured illumination light, moire fringes corresponding to the difference between the periodic structure of the structured illumination light and the periodic structure of the specimen 2 (of the fluorescent region) appear. In this moire fringe, Since the high frequency structure of the sample 2 is shifted to a lower frequency side than the original frequency, the light (fluorescence) indicating this structure is directed to the objective lens 31 at an angle smaller than the original angle. Therefore, when the specimen 2 is illuminated with structured illumination light, even the high-frequency structural information (of the fluorescent region) of the specimen 2 is transmitted by the objective lens 31.

  When the fluorescence generated in the sample 2 is incident on the objective lens 31, it is converted into parallel light by the objective lens 31, then passes through the dichroic mirror 33 and the barrier filter 34, and passes through the second objective lens 35 and passes through the second imaging lens 42. A modulated image of the sample 2 is formed on the imaging surface 41.

  This modulated image is imaged by the image sensor 42 and taken into the image storage / arithmetic device 44 via the control device 43. Further, the captured modulated image is subjected to a known demodulation operation in the image storage / arithmetic unit 44 to generate a demodulated image (super-resolution image). The super-resolution image is stored in an internal memory (not shown) of the image storage / arithmetic unit 44 and is sent to the image display unit 45. As a known demodulation operation, for example, a method disclosed in US Pat. No. 8,115,806 is used.

  Next, the light beam branching portion 14 will be described in detail. FIG. 2 is a diagram for explaining the light beam branching portion 14, FIG. 2A is a view of the diffraction grating 16 of the light beam branching portion 14 as viewed from the optical axis O direction, and FIG. It is a figure which shows the positional relationship of the condensing point which a 1st-order diffracted light beam forms in a pupil conjugate plane. 2A is a schematic diagram, the structure period of the diffraction grating 16 illustrated in FIG. 2A is not necessarily the same as the actual structure period.

  As shown in FIG. 2A, the diffraction grating 16 is a diffraction grating having periodic structures in a plurality of different directions within a plane orthogonal to the optical axis O of the illumination optical system 10. The material of the diffraction grating 16 is, for example, quartz glass. Here, it is assumed that the diffraction grating 16 has a periodic structure in each of the first direction V1, the second direction V2, and the third direction V3 that are different by 120 °, and the structural period in these three directions is common. .

  Note that the periodic structure of the diffraction grating 16 is either a concentration type periodic structure formed using density (transmittance) or a phase type periodic structure formed using steps (phase difference). However, the phase difference type periodic structure is preferable in that the diffraction efficiency of ± first-order diffracted light is high.

  Such parallel light beams incident on the diffraction grating 16 are divided into a first diffracted light beam group branched in the first direction V1, a second diffracted light beam group branched in the second direction V2, and a third light beam branched in the third direction V3. It is converted into a diffracted light beam group.

  The first diffracted light beam group includes a 0th-order diffracted light beam and a ± 1st-order diffracted light beam, and of these, the ± 1st-order diffracted light beams having common orders travel in a symmetric direction with respect to the optical axis O. Similarly, the second diffracted light beam group includes a 0th-order diffracted light beam and a ± 1st-order diffracted light beam, and of these, the ± 1st-order diffracted light beams having common orders travel in a symmetric direction with respect to the optical axis O. To do. Similarly, the third diffracted light beam group includes a 0th-order diffracted light beam and a ± 1st-order diffracted light beam, and of these, the ± 1st-order diffracted light beams having common orders travel in a symmetric direction with respect to the optical axis O. To do.

  Among these light fluxes, the ± first-order diffracted light flux of the first diffracted light flux group, the ± first-order diffracted light flux of the second diffracted light flux group, and the ± first-order diffracted light flux of the third diffracted light flux group are transmitted by the condenser lens 17 described above. Are condensed at different positions in the pupil conjugate plane. Further, the 0th-order diffracted light beam is condensed at the intersection of the pupil conjugate plane and the optical axis O by the condenser lens 17 described above.

  As shown in FIG. 2B, the condensing points 25d and 25g of the first-order diffracted light beams of the first diffracted light beam group are symmetric with respect to the optical axis O, and the arrangement direction of the condensing points 25d and 25g is This corresponds to the first direction V1.

Here, when the wavelength of the laser light emitted from the optical fiber 11 is λ, the structural period of the diffraction grating 16 is P, and the focal length of the lens 17 is fc, the height from the optical axis O to the condensing points 25d and 25g. D is represented by the following formula.
D∝fcλ / P
Therefore, when the wavelength λ of the laser light is changed, a shift occurs at the positions of the condensing points 25d and 25g.

  The condensing points 25c and 25f of the ± 1st-order diffracted light beams of the second diffracted light beam group are symmetric with respect to the optical axis O, and the arrangement direction of the condensing points 25c and 25f corresponds to the second direction V2. . If the wavelength λ is the same, the height from the optical axis O to the condensing points 25c and 25f of the second diffracted light beam group is from the optical axis O to the condensing points 25d and 25g of the first diffracted light beam group. Is the same as the height of

  Further, the condensing points 25b and 25e of the ± first-order diffracted light beams of the third diffracted light beam group are symmetric with respect to the optical axis O, and the arrangement direction of the condensing points 25b and 25e corresponds to the third direction V3. . If the wavelength λ is the same, the height from the optical axis O to the condensing points 25b and 25e of the third light beam group is from the optical axis O to the condensing points 25d and 25g of the first diffracted light beam group. Same as height.

  In addition, a condensing point here is a gravity center position of the area | region which has an intensity | strength 80% or more of maximum intensity | strength. Therefore, the illumination optical system 10 of the present embodiment does not need to collect the light beam until a complete condensing point is formed.

  In the light beam splitting section 14 described above, the translation mechanism 15 includes a piezo motor or the like. The translation mechanism 15 is a diffraction grating that extends in a direction perpendicular to the optical axis O of the illumination optical system 10 and is non-perpendicular to each of the first direction V1, the second direction V2, and the third direction V3. 16 is translated. When the diffraction grating 16 is translated in this direction, the fringe phase of the structured illumination light shifts (details will be described later).

Next, the light amount adjusting unit 18 will be described in detail. FIG. 3 is a schematic diagram illustrating a configuration of the light amount adjusting unit 18. The light amount adjustment unit 18 includes a spatial light modulation unit 18A and a polarizer 18B. The 0th-order diffracted light beam R ₀ and the ± 1st-order diffracted light beams R ₊₁ and R _{−1 of} the third diffracted light beam group are incident on the spatial light modulator 18A. The spatial light modulator 18A controls the orientation of the ferroelectric liquid crystal (FLC) by a control signal S3 from the control device 43, so that each incident light beam is converted into the first polarized light. The spatial light modulator is controlled to one of the second polarized light. The control signal S3 is obtained by changing the ratio of the time for controlling to the first polarization and the time for controlling to the second polarization while capturing one modulated image by the image sensor 42. To control. The first polarized light and the second polarized light will be described later. In the present embodiment, the spatial light modulator 18A is a transmission type, but may be a reflection type. Since the ferroelectric liquid crystal is used for the spatial light modulator 18A, the alignment state can be changed at a high speed (μs order).

  Each light beam whose polarization is controlled by the spatial light modulator 18A is incident on the polarizer 18B. The polarizer 18B is a polarizer that passes only light polarized in a specific direction, such as a polarizing plate.

  FIG. 4 is a diagram illustrating a control unit of the spatial light modulation unit 18A. In FIG. 4, each of the regions Ab, Ac, Ad, Ae, Af, Ag, and Ao is a unit in which the orientation of the ferroelectric liquid crystal is controlled by the control signal S3. Among these, the region Ao is a region where the 0th-order diffracted light beam is incident. Regions Ad and Ag are regions where the first-order diffracted light beams of the first diffracted light beam group are incident. Regions Ac and Af are regions where the ± first-order diffracted light beams of the second diffracted light beam group are incident. Regions Ab and Ae are regions where the ± first-order diffracted light beams of the third diffracted light beam group are incident.

  FIG. 5 is a diagram for explaining the orientation of the molecules of the ferroelectric liquid crystal in the spatial light modulator 18A. The molecules of the ferroelectric liquid crystal can take two alignment states ST1 and ST2 shown in FIG. The angle θ formed by the major axis of the orientation state ST1 and the major axis of the orientation state ST2 is called a cone angle and has a value determined by the ferroelectric liquid crystal. The ferroelectric liquid crystal can switch between the two alignment states ST1 and ST2 in the order of microseconds.

When the refractive index difference between the major axis and the minor axis of the ferroelectric liquid crystal molecules is Δn, the thickness of the spatial light modulator 18A is d, and the wavelength of the incident light beam is λ, the phase difference provided by the spatial light modulator 18A is given. Δφ is expressed by the following equation.
Δφ = 2πΔnd / λ
In the following FIGS. 6 and 7, for the sake of simplicity of explanation, when the thickness d of the spatial light modulator 18A is Δφ = π, that is, when the spatial light modulator 18A operates as a half-wave plate. Will be described as an example. In the present embodiment, the wavelength λ of the laser emitted from the laser unit 100 is not constant, but the spatial light modulator 18A has a wavelength shorter than the center of the range that λ can take. The thickness d is preferably set so as to operate as a half-wave plate. This is because when the wavelength is short, the wavelength range that operates close to the half-wave plate can be widened within the possible range of λ than when the wavelength is long.

  6 and 7 are diagrams for explaining the adjustment of the light amount by the light amount adjusting unit 18. 6 and 7, Ip represents the direction of polarization of the light beam incident on the spatial light modulator 18A. In FIG. 6, the molecules of the ferroelectric liquid crystal are in the alignment state ST1, and the long axis L1 of the molecules is rotated counterclockwise by θ / 2 from the polarization direction Ip of the incident light beam. . At this time, the spatial light modulator 18A functions as a half-wave plate with the long axis L1 of the molecule being the fast axis. For this reason, the polarization (first polarization) Op1 of the light beam emitted from the spatial light modulator 18A is rotated counterclockwise by θ / 2 times, ie, θ, from the polarization direction Ip of the incident light beam. It has become.

  The first polarized light Op1 is incident on the polarizer 18B. In the present embodiment, the angle θp from the axis Ip of the polarizer 18B is equal to θ. That is, the axis of the polarizer 18B coincides with the direction of the first polarization Op1. For this reason, the first polarized light Op1 is emitted as it is as the polarized light O1 from the polarizer 18B.

  On the other hand, in FIG. 7, the molecules of the ferroelectric liquid crystal are in the alignment state ST2, and the long axis L1 of the molecules is rotated clockwise by θ / 2 from the polarization direction Ip of the incident light beam. Yes. At this time, the spatial light modulator 18A functions as a half-wave plate with the long axis L2 of the molecule being the fast axis. For this reason, the polarization (second polarization) Op2 of the light beam emitted from the spatial light modulator 18A is rotated by θ / 2 times, ie, θ, clockwise from the polarization direction Ip of the incident light beam. ing.

The second polarized light Op2 is incident on the polarizer 18B. In the present embodiment, the axis of the polarizer 18B coincides with the direction of the first polarization Op1. For this reason, the component in the direction of the first polarization Op1 out of the second polarization Op2 is emitted from the polarizer 18B as the polarization O2. Thus, the amount of light emitted from the polarizer 18B is smaller when the spatial light modulator 18A is in the alignment state ST2 than when it is in the alignment state ST1. Therefore, while one modulated image is captured for each of the regions Ab to Ag, one modulated image is captured by controlling the time length in which the orientation state is ST1 and the time length in which the orientation state is ST2. It is possible to control the average amount of light during the period.
Note that although the example in which the polarizer 18B is disposed on the diffractive optical element (diffraction grating) 16 side with respect to the light beam selecting unit 19 described later has been described, it may be disposed on the sample 2 side with respect to the light beam selecting unit 19.

  Next, the light beam selection unit 19 will be described in detail. 8 and 9 are diagrams illustrating the light beam selection unit 19. As shown in FIG. 8, the half-wave plate 19A of the light beam selection unit 19 sets the polarization direction of the incident diffracted light beam of each order, and as shown in FIG. 9, the light beam selection member 19B of the light beam selection unit 19 This is a mask that selectively allows any one of the first to third diffracted light beam groups to pass through the ± 1st order diffracted light beam and the 0th order diffracted light beam.

  Then, the rotation mechanism 19C of the light beam selection unit 19 rotates the light beam selection member 19B around the optical axis O so that the selected ± first-order diffracted light beam is between the first to third diffracted light beam groups. In addition to switching, the half-wave plate 19A is rotated around the optical axis O in conjunction with the light beam selection member 19B, so that the polarization direction when the selected ± 1st-order diffracted light beams enter the sample 2 is S. Keep polarized.

  That is, the light beam selection unit 19 switches the stripe direction of the structured illumination light.

  The rotating mechanism 19C includes, for example, a holding member (not shown) that holds the light beam selection member 19B and can rotate around the optical axis O, and a first (not shown) formed around the holding member. , A second gear (not shown) that meshes with the first gear, and a motor (rotary motor) (not shown) connected to the second gear. When this motor is driven, the second gear rotates, the rotational force is transmitted to the first gear, and the light beam selection member 19B rotates around the optical axis O.

  Hereinafter, the conditions for maintaining the stripe state will be specifically described.

  First, the direction of the fast axis of the half-wave plate 19A is the ± first-order diffracted light beam with respect to the branch direction (any one of the first direction V1 to the third direction V3) of the selected ± first-order diffracted light beam. It is necessary to set so that the polarization direction of Here, the fast axis of the half-wave plate 19A is a direction in which the amount of phase delay when light polarized in the direction of the axis passes through the half-wave plate 19A is minimized. It is.

  The opening pattern of the light beam selection member 19B includes a first opening portion H19A and a second opening portion H19B that individually pass one and the other of the ± first-order diffracted light beams belonging to the same diffracted light beam group. The lengths around the optical axis O of each of the first opening H19A and the second opening H19B are set such that the diffracted light beam linearly polarized in the above-described direction can pass therethrough. Therefore, each shape of the 1st opening part H19A and the 2nd opening part H19B is a shape close | similar to partial ring zone shape.

  Here, as shown in FIG. 3A, the rotational position of the half-wave plate 19 when the direction of the fast axis of the half-wave plate 19 is parallel to the direction of the axis of the polarizing plate 13, The rotation position of the half-wave plate 19 is used as a reference (hereinafter referred to as “first reference position”).

  Further, the rotational position of the light beam selection member 19B when the light beam selection direction of the light beam selection member 19B (= the branching direction of the selected ± first-order diffracted light beam) is perpendicular to the axis direction of the polarizer 18B is selected as the light beam selection. The rotation position of the member 19B is used as a reference (hereinafter referred to as “second reference position”).

  At this time, as shown in FIG. 8B, the amount of rotation of the half-wave plate 19A from the first reference position is controlled to one half of the amount of rotation of the light beam selection member 19B from the second reference position. It should be.

  That is, when the rotation amount of the half-wave plate 19A from the first reference position is θ / 2, the rotation amount of the light beam selection member 19B from the second reference position is set to θ.

  Therefore, the rotation mechanism 19C of the light beam selection unit 19 selects the ± first-order diffracted light beam (the branch direction is the first direction V1) of the first diffracted light beam group, as shown in FIG. When the light beam selection direction of the selection member 19B is rotated rightward from the second reference position by the rotation angle θ1, the fast axis direction of the half-wave plate 19A is rotated rightward from the first reference position. Rotate by angle θ1 / 2.

  At this time, the polarization directions of the diffracted light beams of the respective orders before passing through the half-wave plate 19A are parallel to the axis direction of the polarizer 18B, as indicated by a broken line double arrow in FIG. 9A. On the other hand, the polarization direction of each order diffracted light beam after passing through the half-wave plate 19A rotates to the right by the rotation angle θ1, so the polarization direction of the selected ± 1st order diffracted light beam is As indicated by a solid line double arrow in FIG. 9A, the ± first-order diffracted light beams are perpendicular to the branching direction (first direction V1).

  In other words, the direction of the fast axis of the half-wave plate 19A is a direction according to the branching direction (first direction V1) of the ± first-order diffracted light beam selected by the light beam selection member 19B, and is 1/2 Polarization direction (the axial direction of the polarizer 18B) that the ± 1st-order diffracted light beam incident on the wave plate 19A has, and the polarization direction (first direction) that the ± 1st-order diffracted light beam that exits from the half-wave plate 19A should have. Is set in the bisector direction of the angle formed by (perpendicular to the direction V1).

Further, as shown in FIG. 9B, the rotation mechanism 19C of the light beam selection unit 19 selects the ± first-order diffracted light beam (the branch direction is the second direction V2) of the second diffracted light beam group, as shown in FIG. When the light beam selection direction of the selection member 19 is rotated rightward from the second reference position by the rotation angle θ2, the fast axis direction of the half-wave plate 19A is rotated rightward from the first reference position. Rotate by angle θ2 / 2.
At this time, the polarization directions of the diffracted light beams of the respective orders before passing through the half-wave plate 19A are parallel to the direction of the axis of the polarizer 18B, as indicated by the broken line in FIG. 9B. On the other hand, the polarization directions of the diffracted light beams of the respective orders after passing through the half-wave plate 19A rotate to the right by the rotation angle θ2, so the polarization directions of the selected ± 1st order diffracted light beams are As shown by the solid line double arrow in FIG. 9B, these are perpendicular to the branching direction (second direction V2) of the ± first-order diffracted light beams.

  In other words, the direction of the fast axis of the half-wave plate 19A is a direction according to the branching direction (second direction V2) of the ± first-order diffracted light beam selected by the light beam selection member 19, and is 1/2 The polarization direction (the axial direction of the polarizer 18B) that the ± first-order diffracted light beam incident on the wave plate 19A has, and the polarization direction (second) that the ± first-order diffracted light beam that exits from the half-wave plate 19A should have. Is set in the bisector direction of the angle formed by (perpendicular to the direction V2).

  Further, as shown in FIG. 9C, the rotation mechanism 19C of the light beam selection unit 19 selects the ± first-order diffracted light beam (the branch direction is the third direction V3) of the third diffracted light beam group, as shown in FIG. When the light beam selection direction of the selection member 19B is rotated from the second reference position to the left (seen from the sample side; hereinafter the same) by the rotation angle θ3, the direction of the fast axis of the half-wave plate 19A is The first reference position is rotated to the left by the rotation angle θ3 / 2.

  At this time, the polarization directions of the diffracted light beams of the respective orders before passing through the half-wave plate 19A are parallel to the direction of the axis of the polarizer 18B, as indicated by the broken line in FIG. 9C. On the other hand, the polarization direction of each order diffracted light beam after passing through the half-wave plate 19A rotates to the left by the rotation angle θ3, so the polarization direction of the selected ± 1st order diffracted light beam is As shown by the actual double-headed arrow in FIG. 9C, these are perpendicular to the branching direction (third direction V3) of the ± first-order diffracted light beams.

  In other words, the direction of the fast axis of the half-wave plate 19A is a direction according to the branching direction (third direction V3) of the ± first-order diffracted light beam selected by the light beam selection member 19B, and is 1/2 The polarization direction (the axial direction of the polarizer 18B) that the ± first-order diffracted light beam incident on the wave plate 19A has, and the polarization direction (third) that the ± first-order diffracted light beam that exits from the half-wave plate 19A should have. In the direction of the bisector of the angle formed by (perpendicular to the direction V3).

  Accordingly, the rotation mechanism 19C of the light beam selection unit 19 may be interlocked with the half-wave plate 19A and the light beam selection member 19B with a gear ratio of 2: 1.

  In the above description, the rotatable half-wave plate 19A is used to keep the ± first-order diffracted light beam incident on the sample 2 as S-polarized light. Instead, a fixedly arranged liquid crystal element may be used, and the liquid crystal element may function as the half-wave plate 19A. If the orientation of the liquid crystal element is electrically controlled, the refractive index anisotropy of the liquid crystal element can be controlled, so that the fast axis as a half-wave plate can be rotated around the optical axis O. Incidentally, there are other methods for keeping the ± first-order diffracted light beam incident on the specimen 2 in S-polarized light (described later).

  FIG. 10 is a diagram illustrating the function of the light beam selection unit 19 described above. In FIG. 10, a double arrow line surrounded by a circular frame indicates the polarization direction of the light beam, and a double arrow line surrounded by a square frame indicates the axial direction of the optical element. However, the fast axis of the spatial light modulator 18A is the fast axis in the orientation state ST1.

  Further, as shown in FIG. 11, a plurality of (six in the example shown in FIG. 11) notches H19C are formed on the outer peripheral portion of the light beam selecting member 19B. A timing sensor H19D for detecting the notch H19C is provided. As a result, the rotation mechanism 19C can detect the rotation position of the light beam selector 19 and thus the rotation position of the half-wave plate 19A.

  Next, the translation mechanism 15 of the beam splitter 14 will be described in detail.

  FIG. 12 is a diagram for explaining the operation of the translation mechanism 15 of the light beam branching section 14.

  First, in order to enable the demodulation operation described above, three or more modulated images with the same direction of interference fringes and different phases are necessary for the same sample 2. This is because the modulated image generated by the structured illumination microscope apparatus 1 includes, in the structure of the sample 2, the 0th order modulation component, the + 1st order modulation component, which is the structure information whose spatial frequency is modulated by the structured illumination light, and −1. This is because the next modulation component is included, and in order to make these three unknown parameters known by the demodulation operation, the number of modulated images equal to or greater than the number of these unknown parameters is required.

  Therefore, the translation mechanism 15 of the light beam splitting unit 14 is in the direction perpendicular to the optical axis O of the illumination optical system 10 as described above, as shown in FIG. The diffraction grating 16 is shifted in a non-perpendicular direction (x direction) with respect to all of the first direction V1, the second direction V2, and the third direction V3.

  However, the shift amount L of the diffraction grating 16 necessary for shifting the phase of the interference fringes by a desired shift amount φ is set when the light beam selection direction by the light beam selection unit 19 is the first direction V1 and in the second direction V2. And when in the third direction V3 are not necessarily the same.

  As shown in FIG. 12B, the structural period of each of the first direction V1, the second direction V2, and the third direction V3 of the diffraction grating 16 is P, and the shift direction (x direction) of the diffraction grating 16 and the first The angle formed by the direction V1 is set as θ1, the angle formed by the shift direction (x direction) of the diffraction grating 16 and the second direction V2 is set as θ2, and the shift direction (x direction) of the diffraction grating 16 and the third direction V3. Is set to θ3, the required shift amount L1 of the diffraction grating 16 in the x direction when the light beam selection direction is the first direction V1 is L1 = φ × P / (a × 4π × | cos θ1 | ), And the required shift amount L2 of the diffraction grating 16 in the x direction when the light beam selection direction is the second direction V2 is expressed by L2 = φ × P / (a × 4π × | cos θ2 |), The required shift amount L3 of the diffraction grating 16 in the x direction when the light beam selection direction is the third direction V3 is: 3 = φ × P / (a × 4π × | cosθ3 |) represented by.

  That is, the shift amount L in the x direction of the diffraction grating 16 necessary for setting the phase shift amount of the interference fringes to a desired value φ is the wavelength selection direction (the first direction V1, the second direction V2, and the third direction V3). Any) and the angle θ formed by the x direction is expressed by the equation.

L = φ × P / (a × 4π × | cos θ |)
Incidentally, the shift amount L in the x direction of the diffraction grating 16 necessary for setting the phase shift amount φ of the interference fringes to 2π is P / (a × 2 × | cos θ |). This is an amount corresponding to a half period of the diffraction grating 16. In other words, the phase of the structured illumination light can be shifted by one period only by shifting the diffraction grating 16 by a half period (because the fringe period of the interference fringes composed of ± first-order diffracted light is 2 of the structural period of the diffraction grating 16. Equivalent to twice).

FIG. 13 is a diagram for explaining control by the control device 43. In this embodiment, in order to obtain one demodulated image, five modulated images using the diffracted light beam groups are used for each of the three diffracted light beam groups, that is, a total of 15 modulated images. The control device 43 includes a translation mechanism 15 of the light beam branching unit 14, a spatial light modulation unit 18A of the light amount adjustment unit 18, and a light beam selection unit so that each of the 15 modulated images is captured under a predetermined condition. The 19 rotation mechanisms 19C and the image sensor 42 are controlled.
Note that, for example, a method disclosed in US Pat. No. 8,115,806 is used as a demodulation operation to generate a demodulated image using the acquired 15 modulated images.

  As shown in FIG. 13, among the 15 modulated images, 5 modulated images using the first diffracted light beam group are taken in the time zone Td1. Five modulated images using the second diffracted light beam group are taken in a time zone Td2 following the time zone Td1. Five modulated images using the third diffracted light beam group are taken in a time zone Td3 following the time zone Td2. Since the time zones Td1 to Td3 are the same, the details of the time zone Td1 will be described as a representative.

  The time zone Td1 is composed of six time zones of time zones Tcd, Tp1, Tp2, Tp3, Tp4, and Tp5 in time order. In the first time zone Tcd, the control device 43 controls the rotation mechanism 19C so that the first diffracted light beam group is selected by the light beam selection unit 19. In the next time zone Tp1, the control device 43 performs control so that the phase of the interference fringes is captured in the first phase. Subsequently, in the time zone Tp2, the control device 43 performs control so as to capture a modulated image in which the phase of the interference fringes is the second phase. In each of the subsequent time zones Tp3, Tp4, and Tp5, the control device 43 performs control so that the phase of the interference fringes is captured in the third phase, the fourth phase, and the fifth phase, respectively.

  Since the time zones Tp1 to Tp5 are the same, the details of the time zone Tp2 will be described as a representative. In the first time zone Tcp of the time zone Tp2, the control device 43 controls the translation mechanism 15 so that the phase of the interference fringes becomes the second phase. In the time zone T101 following the time zone Tcp, the control device 43 is configured so that the region (region Ao in FIG. 4) through which the zero-order diffracted light beam of the spatial light modulator 18A passes is in the alignment state ST1. 18A is controlled. In a time zone T102 following the time zone T101, the control device 43 controls the spatial light modulation unit 18A so that the region through which the zero-order diffracted light beam of the spatial light modulation unit 18A passes is in the alignment state ST2.

  In a time zone T11, which is a time zone following the time zone Tcp and is a combination of the time zone T101 and the time zone T102, a region through which the ± 1st-order diffracted light beam of the spatial light modulator 18A passes (regions Ab to Ag in FIG. 4). However, the control device 43 controls the spatial light modulator 18A so as to be in the alignment state ST1. Then, during the time zone T111, the control device 43 performs control so that the image sensor 42 captures one modulated image.

The length T of the time zone Tl11 is set in advance. The control device 43 divides the time zone Tl11 into the time zone Tl01 and the time zone Tl02, that is, how many seconds the length of the time zone Tl02 is set as follows. decide. First, the control device 43 has a ratio C ₁ between the zero-order component and the first-order component of the optical transfer function (OTF) of the illumination optical system 10, a ratio C ₂ between the zero-order component and the second-order component, The wavelength λ of the laser emitted by the laser unit 100 is acquired. The control device 43 acquires these values set by the user, for example. The control device 43 reads out a value representing the length t ₁ of the time zone T101 stored in advance in association with the combination of the ratios C ₁ , C ₂ , and wavelength λ, and uses it.

The length t ₁ of the time zone T101 that is associated with the combination of the ratios C ₁ , C ₂ , and the wavelength λ is stored as calculated below. The ratios C ₁ and C ₂ and the length t ₁ of the time zone T101 are expressed by the following relational expression (1).
Even when varying the wavelength of the structured illumination light forming the sample 2, by changing the length t ₁ of the time zone TL01, the respective orders of the light components contained in the structured illumination regardless of the wavelength The ratio between these components can be made constant. If the intensity of the 0th-order diffracted light beam is smaller than the intensity of the 1st-order diffracted light beam, it is desirable to assign a part of the time zone T11 to the orientation state ST2.

Here, T is the total length of the time zone T101 and the time zone T102. I _{i, j} is the amount of light emitted from the light amount adjusting unit 18 of the i-th order diffracted light beam in the orientation state STj. I _i is the light amount when the i-th order diffracted light beam enters the light amount adjusting unit 18. Here, I _{i, j} and I _i are represented by the relationship shown in Expression (2). Further, the ratio between I ₀ and I ₁ is determined by the diffraction grating 16.

Α and β can be determined as follows according to the phase difference Δφ of the spatial light modulator 18A, the cone angle θ, and the axial direction θp of the polarizer 18B.
Assuming that the electric field amplitude of the incident light is E, the electric field amplitudes E ₁ and E _{s in} the major axis direction and minor axis direction of the molecules of the ferroelectric liquid crystal are expressed by equations (3) and (4).

Here, when Δφ = π at the design wavelength λ ₀ , the phase difference Δφ at a certain wavelength λ is expressed by Expression (5).

  The electric field amplitude Ep1 in the alignment state ST1 is expressed by the equation (6), and the electric field amplitude Ep2 in the alignment state ST2 is expressed by the equation (7).

  In general, since the relationship between the electric field and the light intensity is expressed by Expression (8), α and β are expressed by Expressions (9) and (10) below.

By calculating α and β using the equations (9) and (10), the correspondence between the length t ₁ of the time zone T101 and the ratios C ₁ and C ₂ can be calculated. When the wavelength of the laser is lambda _0, because it is [Delta] [phi = [pi, Equation (9), instead of (10), the following equation (11), can be used (12).

  Further, when θp = θ, the following equations (13) and (14) can be used instead of equations (9) and (10).

In the present embodiment, the orientation state ST2 is used only for the 0th-order diffracted light beam, but may be used for the ± 1st-order diffracted light beam.
In FIG. 12, the time order of the time zone T101 and the time zone T102 may be reversed. Alternatively, the time sequence of the time zone Tl01 and the time zone T102 in the time zone Tp1 and the time sequence of the time zone Tl01 and the time zone T102 in the time zone Tp2 may be reversed so as to be reversed. Good. By alternately changing, the number of changes in the orientation state can be halved.

[Modification of each embodiment]
In the above-described embodiment, in order to keep the ± first-order diffracted light beam incident on the specimen 2 as S-polarized light, the half-wave plate 19A that can rotate around the optical axis is used. A quarter-wave plate and a quarter-wave plate that can rotate around the optical axis may be used. However, in that case, the rotation position of the quarter-wave plate with respect to the first reference position is set to be the same as the rotation position of the light beam selection member 19B with reference to the second reference position.

  In the above-described embodiment, as means for branching the light beam emitted from the light source, the diffraction grating 16 having the number of directions M of the periodic structure of 2 or more, that is, diffraction that simultaneously generates a plurality of diffraction light beam groups having different branch directions. Although the grating 16 (see FIG. 2A) is used, a diffraction grating having a periodic structure direction number M of 1, that is, a diffraction grating that generates only one group of diffracted light beams having a common branching direction is used. Also good.

  However, in that case, the diffraction grating may be rotated around the optical axis in order to switch the direction of the interference fringes. In that case, instead of the rotatable light beam selection member 19B, a mask that cuts non-movable ± first-order diffracted light beams may be used.

  In other words, in the illumination optical system 10 described above, the diffractive means (16) may be a diffraction grating having a periodic structure over a single direction perpendicular to the optical axis, in which case the specimen (2) A rotating unit (not shown) that rotates the incident direction of the pair of light fluxes about the optical axis may be further provided.

  Further, in the above-described embodiment, since the interference fringes formed on the specimen 2 in the 3D-SIM mode are the three-beam interference fringes, the + 1st-order diffracted light flux and the −1st-order diffracted light are used as the diffracted light flux contributing to the structured illumination light. Although a combination of a light beam and a zero-order diffracted light beam is used, it goes without saying that other combinations may be used.

  Incidentally, in order to form a three-beam interference fringe, three-beam interference caused by three diffracted lights having the same diffraction order interval may be generated. For example, zero-order diffracted light, first-order diffracted light, and second-order diffracted light , A combination of ± 2nd order diffracted light and 0th order diffracted light, a combination of ± 3rd order diffracted light and 0th order diffracted light, and the like can be used.

  Note that the illumination optical system 10 of the present embodiment is configured by the epi-illumination optical system by the objective lens 31, but is not limited thereto, and is configured by a transmission / reflection illumination optical system by a condenser lens instead of the objective lens 31. Also good. In this case, the condensing point is formed on the pupil plane of the condenser lens.

  The light beam branching unit 14 of the present embodiment uses a diffraction grating as means for branching the light beam. For example, a spatial light modulator (SLM) that functions as a phase-type diffraction grating and a spatial light modulator are used. A combination with a control unit that supplies a drive signal to the device may be used. The controller can switch the structure period (structure period as a diffraction grating) of the spatial light modulator at high speed by switching the drive signal applied to the spatial light modulator.

  Further, a program for realizing the function of the control device 43 in FIG. 1 is recorded on a computer-readable recording medium, and the program recorded on the recording medium is read into the computer system and executed, thereby executing the control device 43. May be realized. Here, the “computer system” includes an OS and hardware such as peripheral devices.

  The “computer-readable recording medium” refers to a storage device such as a flexible medium, a magneto-optical disk, a portable medium such as a ROM and a CD-ROM, and a hard disk incorporated in a computer system. Furthermore, the “computer-readable recording medium” dynamically holds a program for a short time like a communication line when transmitting a program via a network such as the Internet or a communication line such as a telephone line. In this case, a volatile memory in a computer system serving as a server or a client in that case, and a program that holds a program for a certain period of time are also included. The program may be a program for realizing a part of the functions described above, and may be a program capable of realizing the functions described above in combination with a program already recorded in a computer system.

The embodiment of the present invention has been described in detail with reference to the drawings. However, the specific configuration is not limited to this embodiment, and includes design changes and the like within a scope not departing from the gist of the present invention.
Note that the requirements of the above-described embodiments can be combined as appropriate. Some components may not be used. In addition, as long as it is permitted by law, the disclosure of all publications and US patents relating to the devices cited in the above embodiments and modifications are incorporated herein by reference.

DESCRIPTION OF SYMBOLS 1 ... Structured illumination microscope apparatus 10 ... Illumination optical system 11 ... Optical fiber 12 ... Collector lens 13 ... Polarizing plate 14 ... Light beam splitting part 15 ... Translation mechanism 16 ... Diffractive optical element 17 ... Condensing lens 18 ... Light quantity adjustment part 18A ... Spatial light modulator 18B ... Polarizer 19 ... Flux selector 19A ... 1/2 wavelength plate 19B ... Flux selector 19C ... Rotating mechanism 21 ... Lens 22 ... Field stop 23 ... Field lens 24 ... Excitation filter 30 ... Imaging optics System 31 ... Objective lens 33 ... Dichroic mirror 42 ... Imaging element 43 ... Control device 44 ... Image storage / calculation device 45 ... Image display device 100 ... Laser unit

Claims (15)

A structured illumination device comprising an optical system for forming interference fringes on a specimen with a predetermined number of light beams,
The optical system is
A spatial light modulator that controls polarization of at least one of the predetermined number of light beams;
A structured illumination device comprising: a polarization member that extracts a component in a predetermined polarization direction from the light flux emitted from the spatial light modulation unit. A control unit for controlling the spatial light modulation unit;
The controller is
Controlling the spatial light modulation unit so that the polarization of the light beam emitted from the spatial light modulation unit is changed from the first polarization to the second polarization in a predetermined period;
The structured lighting device according to claim 1.   The control unit includes a first time zone in which the polarization of the light beam emitted from the spatial light modulation unit becomes the first polarization in a predetermined period, and the first polarization in the predetermined period. 2. The second time zone to be different second polarization is determined, and the spatial light modulation unit is controlled based on the determined first time zone and the second time zone. Structured lighting device.   The structured lighting device according to claim 3, wherein the control unit determines the first time zone and the second time zone according to a wavelength of the light beam.   The determination unit includes the first time zone, the second time zone, and the second time zone so that a ratio between coefficients of each component of the optical transfer function of the structured lighting device is constant regardless of the wavelength. The structured lighting device according to claim 4, wherein the structured lighting device is determined. A branching section for branching the emitted light beam from the light source into the predetermined number of light beams;
The structured illumination device according to any one of claims 1 to 5, further comprising: a rotation mechanism that rotates the predetermined number of light beams around an optical axis of the optical system. A branching section for branching the emitted light beam from the light source into a plurality of light beams;
A light beam selection member having a predetermined number of selection units for selecting only the predetermined number of light beams from the plurality of light beams;
A rotation mechanism that switches the combination of the predetermined number of light beams selected by the light beam selection member by rotating the light beam selection member around the optical axis of the optical system. The structured lighting device according to claim 1. The optical system is
An objective lens;
A first condensing lens that condenses each of the predetermined number of light fluxes on or near the pupil conjugate plane of the objective lens;
A second condenser lens that condenses each of the predetermined number of light beams that have passed through the pupil conjugate plane on or near the pupil plane of the objective lens,
The spatial light modulator and the polarizing member are provided between the first condenser lens and the second condenser lens,
The structured illumination device according to any one of claims 1 to 7. The optical system includes a collimating lens that converts each of the predetermined number of light beams that have passed through the pupil conjugate plane into substantially parallel light,
The second condenser lens condenses each of the predetermined number of light beams converted into substantially parallel light by the collimator lens on the pupil plane,
The spatial light modulator and the polarizing member are provided between the first condenser lens and the collimating lens.
The structured lighting device according to claim 8.   10. The structured illumination device according to claim 1, wherein the spatial light modulation unit includes a ferroelectric liquid crystal, and the polarization of the light beam is controlled by the ferroelectric liquid crystal. 11.   The structured illumination device according to claim 10, wherein the ferroelectric liquid crystal functions as a half-wave plate with respect to light having a predetermined wavelength by being provided with a phase difference π.   The structured illumination device according to claim 11, wherein the predetermined wavelength is on a shorter wavelength side than a center of a wavelength band of a plurality of lights respectively emitted from the light source.   The structured illumination device according to any one of claims 1 to 12, further comprising a phase shift unit that shifts a phase of the interference fringes. A structured lighting device according to any one of claims 1 to 13,
A structured illumination microscope apparatus comprising: an imaging unit that acquires a modulated image that is an image of the specimen spatially modulated by the interference fringes.   The structured illumination microscope apparatus according to claim 14, further comprising a calculation unit that generates a demodulated image of the specimen based on the modulated image.

Images