JP5861873B2 - Spectrometer and microspectroscopic system - Google Patents

Description

  The present invention relates to a spectroscope and a microspectroscopic system.

  In the conventional scanning fluorescence microscope, the illumination light (excitation light) emitted from the point light source is scanned on the specimen by the scanning unit, and the signal light (fluorescence) emitted from the specimen excited by the illumination light is scanned by the scanning unit. After descanning, the signal light separated by the spectroscopic element of the spectroscope is detected by the respective light receivers. As such a spectroscopic element for a spectroscope, a diffraction grating is widely used. This diffraction grating has polarization dependency, and the direction of the electric field oscillation of incident light is light of the S polarization component perpendicular to the marking line of the diffraction grating as compared to the light of the P polarization component parallel to the marking line of the diffraction grating. However, it is generally said that the diffraction efficiency is high. In particular, the above tendency appears remarkably in a long wavelength band where the wavelength of incident light is equal to or more than the period of the diffraction grating (the pitch of the engraving line). In other words, the shorter the diffraction grating period, the more the above-mentioned tendency appears and the higher the polarization dependency. This is a point to be considered when using a diffraction grating with high resolution. Therefore, in order to eliminate the difference in diffraction efficiency due to the difference in polarization direction, the signal light is separated into P-polarized component light and S-polarized component light, and P-polarized component light is converted into S-polarized component light. A configuration is disclosed in which these lights are separated by a diffraction grating (see, for example, Patent Document 1).

US Pat. No. 6,498,872

  However, if the separated P-polarized component light and S-polarized component light are incident on different positions of the diffraction grating, the diffraction grating becomes larger, and at least a part of the signal light is reflected on the diffraction grating. Increasing the size of the diffraction grating can be avoided if they are incident so as to overlap, but because the incident angles of these lights with respect to the diffraction grating are different, the condensing optics for condensing the spectral light split by the diffraction grating onto the receiver Two systems (for example, an imaging lens and a concave mirror) are required, resulting in a problem that the spectroscope becomes large.

  The present invention has been made in view of such problems, and a spectroscope capable of suppressing an increase in size and cost while taking into account the polarization dependence of the spectroscopic element, and a microspectroscopic system including this spectroscope. The purpose is to provide.

  In order to solve the above-described problems, a spectroscope according to the present invention includes a collimating lens that makes signal light a substantially parallel light beam, and a substantially parallel light beam emitted from the collimating lens in two optical paths, a first optical path and a second optical path. And a polarization separation element having the light in the first optical path as the P-polarized component and the light in the second optical path as the S-polarized component, and the P-polarized component in the first optical path disposed in the first optical path A polarization rotator that uses the S-polarized light as the light of S polarization, an optical path coupling member that reduces the beam diameter of the substantially parallel light flux in each of the first optical path and the second optical path while maintaining a parallel state, and an emission from the optical path coupling member A spectroscopic element that splits the substantially parallel light beam, a light receiver that detects the spectroscopic light dispersed by the spectroscopic element, and a condensing optical system that condenses the spectral light from the spectroscopic element on the light receiver. Features.

  Further, in such a spectroscope, the optical path coupling member includes a first reflecting surface and a second reflecting surface having an off-axis paraboloid shape that reflects a substantially parallel light beam emitted from the first optical path. And a second beam expander composed of a first reflecting surface and a second reflecting surface having an off-axis paraboloid shape that reflects a substantially parallel light beam emitted from the second optical path. Has been.

Further, in such a spectrometer, the optical path coupling member is preferably in a substantially parallel beam of each beam diameter is reduced to be adjacent to each other are configured so as to emit.

  Further, in such a spectroscope, the optical path coupling member has an overall diameter of each substantially parallel light beam emitted from the optical path coupling member, which is equal to or less than a diameter of the substantially parallel light beam emitted from the collimator lens and incident on the polarization separation element. It is preferable that

  In such a spectroscope, the polarization separation element is disposed at a position where the substantially parallel light beam emitted from the collimator lens is incident, and transmits the P-polarized light component of the substantially parallel light beam and guides it to the first optical path. The polarizing beam splitter preferably reflects the S-polarized component light, and the plane mirror reflects the S-polarized component light reflected by the polarizing beam splitter and guides it to the second optical path.

  In such a spectroscope, the polarization rotation element is preferably a Fresnel ROM half-wave plate.

  In addition, a microspectroscopic system according to the present invention includes a microscope that scans illumination light emitted from a light source, irradiates the specimen with an objective lens, and collects signal light emitted from the specimen with the objective lens; One of the above-described spectroscopes that spectrally detects signal light from the above.

  According to the present invention, it is possible to provide a spectroscope that suppresses the increase in size and cost while taking into account the polarization dependence of the spectroscopic element, and a microspectroscopic system including this spectroscope.

It is explanatory drawing which shows the structure of a microspectroscopic system. It is explanatory drawing which shows the structure of a spectrometer. It is explanatory drawing which shows the principal part of the said spectrometer.

  Hereinafter, preferred embodiments of the present invention will be described with reference to the drawings. First, the configuration of the microspectroscopic system will be described with reference to FIG. As shown in FIG. 1, the microspectroscopic system 1 includes a confocal microscope having a light source system 10, a confocal unit 20, and a microscope 30, a spectroscope 40, and a control unit 50. . In this microspectroscopy system 1, the confocal unit 20 and the spectroscope 40 are optically connected by an optical fiber 28 via fiber couplers 29a and 29b.

  The light source system 10 includes a laser device 11, an optical fiber 13, and fiber couplers 12 and 14. The laser device 11 includes, for example, a laser diode, and emits laser light (illumination light) having a target wavelength characteristic. This laser beam is guided to the confocal unit 20 through the optical fiber 13. In the example of FIG. 1, excitation light for exciting the specimen 33 to emit fluorescence is emitted as illumination light.

  The confocal unit 20 includes a collimating lens 21 that converts illumination light from the light source system 10 into a substantially parallel light beam, a dichroic mirror 22, a scanning unit 23, a scanner lens 24, a condensing lens 25, and a pinhole 26a. A pinhole plate 26 and a relay lens 27 are provided. The microscope 30 includes a second objective lens 31 and an objective lens 32, and a stage 34 on which the specimen 33 is placed. The confocal unit 20 and the microscope 30 are combined to form a scanning confocal microscope. The dichroic mirror 22 is configured to reflect the laser light emitted from the light source system 10 toward the microscope 30 and transmit the fluorescence (signal light) emitted from the specimen 33 excited by the laser light. . Further, the image side focal point of the condenser lens 25 is disposed so as to substantially coincide with the pinhole 26 a of the pinhole plate 26.

  Laser light (illumination light) emitted from the laser device 11 of the light source system 10 is introduced into the optical fiber 13 through the fiber coupler 12. Further, the laser light passing through the optical fiber 13 enters the collimating lens 21 of the confocal unit 20 from the fiber coupler 14. The laser light is converted into substantially parallel light by the collimator lens 21, then reflected by the dichroic mirror 22 to the optical path on the microscope 30 side, and applied to the scanning unit 23 and the scanner lens 24, which are composed of two orthogonally arranged galvanometer mirrors. Introduced and scanned in two dimensions. The scanned laser light is made into substantially parallel light by the second objective lens 31 and then condensed at one point on the specimen 33 by the objective lens 32. Note that the position on the specimen 33 scanned two-dimensionally by the scanning unit 23 is controlled by controlling the operation of the galvanometer mirror constituting the scanning unit 23 by the control unit 50. Then, the fluorescence (signal light) emitted from the specimen 33 excited by the laser light (illumination light) is converted into substantially parallel light by the objective lens 32, and follows a path opposite to the laser light (illumination light). After being descanned by the scanning unit 23, the light enters the dichroic mirror 22. Further, the fluorescence incident on the dichroic mirror 22 passes through the dichroic mirror 22 and is collected by the condenser lens 25 onto the pinhole 26 a of the pinhole plate 26.

  The fluorescence (signal light) that has passed through the pinhole 26a is guided to the optical fiber 28 from the fiber coupler 29a via the relay lens 27. Through the relay lens 27, as shown in FIG. 1, the light that has passed through the pinhole 26a becomes a divergent light beam as it is, and is condensed again, and apparently appears at the opening end of the optical fiber 28. Even with a small aperture diameter, it becomes possible to enter effectively (with little loss).

  Here, since the condensing point formed in the pinhole 26a is an image of a light spot on the specimen 33, even if there is light emitted from another point on the specimen 33, the pinhole 26a. Then, an image is not formed and is blocked by the pinhole plate 26 and hardly reaches the fiber coupler 29a. Therefore, only the light that has been able to pass through the pinhole 26a, that is, only the light emitted from the position of the light spot described above, can reach the fiber coupler 29a through the relay lens 27. As a result, the scanning confocal microscope is a microscope capable of observing a specimen not only with high lateral resolution but also with high vertical resolution.

  The fluorescence (signal light) incident on the fiber coupler 29a passes through the optical fiber 28 and is introduced into the spectroscope 40 via the fiber coupler 29b. Hereinafter, the configuration of the spectrometer 40 according to the present embodiment will be described.

  As shown in FIGS. 2 and 3, the spectroscope 40 includes a collimator lens 41 that makes signal light (fluorescence in the example of FIG. 1) incident from the optical fiber 28 through the fiber coupler 29b substantially parallel, and this collimator. The signal light converted into a substantially parallel light beam by the lens 41 is separated into P-polarized component light and S-polarized light component, the P-polarized light component is guided to the first optical path C1, and the S-polarized light component is input to the second optical path C2. The polarization separation element 42 for guiding, the polarization rotation element 43 provided in the first optical path C1, and converting the light of the P polarization component into the light of the S polarization component, and the light beam diameter of the light in the first optical path C1 and the second optical path C2. The optical path coupling element 44 that emits these lights in parallel and adjacent to each other, the reflective diffraction grating 45 that is a spectroscopic element that splits the substantially parallel light beam emitted from the optical path coupling element 44, and a plurality of light receiving elements Elements are arranged in an array Is a and the receiver 47 constituting the line detector, and is thereby condensing optical system 46 forms an image on the light receiving surface of the light receiver 47 is emitted from the diffraction grating 45 is diffracted light of (spectral light), the by. Here, the light receiver 47 uses 32 PMTs (Photomultiplier Tubes). The plurality of PMTs are arranged in the direction in which the plurality of score lines of the diffraction grating 45 are arranged, that is, in the light dispersion direction (spectral direction) of the diffraction grating 45. Thus, in this embodiment, since the photodetection array having a plurality of detection cells (PMT) is used as the light receiver 47, light in a plurality of wavelength regions can be detected simultaneously. Although FIG. 2 shows a case where a concave mirror (reflection optical system) is used as the condensing optical system 46, a lens having a positive refractive power (refractive optical system) may be used.

  The polarization separation element 42 includes a polarization beam splitter plate 42a disposed at a position where a substantially parallel light beam emitted from the collimating lens 41 is incident so that an incident angle of the substantially parallel light beam is approximately 45 °, and a polarization beam splitter plate. The plane mirror 42b is arranged so that the incident angle of the substantially parallel light beam reflected by 42a is about 45 ° and reflects the substantially parallel light beam. The incident angle is the angle of incident light with respect to the normal of the incident surface. Here, the polarization beam splitter plate 42a is obtained by applying a dielectric reflection coating to a parallel plate glass, and has a property of reflecting S-polarized light and transmitting P-polarized light among incident light. . Therefore, of the substantially parallel light beam (signal light) incident on the polarization beam splitter plate 42a, the P-polarized component light is transmitted through the polarization beam splitter plate 42a and guided to the first optical path C1, and the S-polarized component light. Is reflected by the polarization beam splitter plate 42a, further reflected by the plane mirror 42b, and guided to the second optical path C2. As described above, since the polarization beam splitter plate 42a and the plane mirror 42b are arranged so that the angle with respect to the incident light is about 45 °, the first optical path C1 and the second optical path C2 extend substantially in parallel. Are arranged. For convenience of the following explanation, the direction parallel to the optical axis of the collimating lens 41 is in the Z direction, and the light from the collimating lens 41 is reflected in the plane parallel to the direction in which the light is reflected by the polarization beam splitter plate 42a and the Z direction. A direction perpendicular to the Z direction is defined as a Y direction, and a direction perpendicular to the Y direction and the Z direction is defined as an X direction.

  The polarization rotation element 43 is configured by, for example, a Fresnel ROM half-wave plate. The Fresnel ROM 1/2 is a birefringent element using total reflection, and has a very small wavelength dependency, and can be used for light of various wavelengths. In the Fresnel ROM half-wave plate, the light incident from the incident surface 43a is reflected four times on one plane (reflected by the planes 43b to 43e) and emitted from the emission surface 43f. In the Fresnel ROM half-wave plate, the incident surface 43a and the output surface 43f are perpendicular to the Z direction, and the above-described one planes 43b to 43e are 45 ° to the ZX plane as shown in FIG. It is arranged to form. As described above, by providing the Fresnel ROM half-wave plate as the polarization rotation element 43, the signal light (substantially parallel light beam) that has passed through the polarizing beam splitter plate 42a and becomes the P-polarized component light is reflected in this Fresnel ROM 1 / For each total reflection in the two-wavelength plate, the polarization direction is changed by λ / 8, and the total is changed by λ / 2. As a result, the light is converted into light of the S polarization component. The S-polarized component light travels on the optical axis of the collimating lens 41, in other words, in a direction parallel to the Z direction.

  The optical path coupling element 44 includes a first beam expander 441 disposed in the first optical path C1 and a second beam expander 442 disposed in the second optical path C2. Each of the first and second beam expanders 441 and 442 is disposed at a position where light from the first and second optical paths C1 and C2 is incident, and has an off-axis paraboloid shape. First reflective surfaces 441a and 442a that reflect light, and an off-axis release that reflects light reflected by the first reflective surfaces 441a and 442a that are arranged to face the first reflective surfaces 441a and 442a. It is comprised by the two off-axis paraboloids which consist of the 2nd reflective surfaces 441b and 442b which have an object shape. In the present embodiment, the focal lengths of the first reflecting surfaces 441a and 442a are configured to be approximately twice the focal length of the second reflecting surfaces 441b and 442b. For this reason, the beam diameter of the signal light reflected by the second reflecting surfaces 441b and 442b is reduced to approximately half the beam diameter of the first and second optical paths C1 and C2. In the present embodiment, the second reflecting surface 441b used for the first beam expander 441 and the second reflecting surface 442b used for the second beam expander 442 are in contact with each other and Z Since the light is emitted so as to be substantially parallel to the axis, the maximum light beam diameter when entering the diffraction grating 45 is the same as the light beam diameter when entering the polarization separation element 42. Therefore, even if the signal light emitted from the collimating lens 41 is divided into two optical paths by the polarization separation element 42 and the polarization is aligned by the polarization rotation element 43, the beam diameter will not be doubled as a result. The diffraction grating 45 does not increase in size. Further, the signal light passing through each of the first and second optical paths C1 and C2 emitted from the optical path coupling element 44 is a substantially parallel light flux and has the same incident angle with respect to the diffraction grating 45. There is no need to prepare two, and an increase in the size of the spectrometer 40 can be prevented. In the present embodiment, the conversion magnifications of the light beam diameters of the first and second beam expanders 441 and 442 are uniform. However, the conversion magnifications that are different between the first beam expander 441 and the second beam expander 442. It may be. Also in this case, it is preferable that the maximum light beam diameter when entering the diffraction grating 45 is equal to or smaller than the light beam diameter when entering the polarization separation element 42.

  If the spectroscope 40 according to the present embodiment is configured as described above, the polarization direction of the signal light can be aligned with the S-polarized light, so that the electric field vibration direction of the signal light incident on the diffraction grating 45 is the time of the diffraction grating 45. Since it can be perpendicular to the line, the diffraction efficiency of the diffraction grating 45 can be increased, and bright spectral light can be obtained. Further, as described above, the light beam diameter of the signal light incident on the diffraction grating 45 by the optical path coupling element 44 (the total light beam diameter of the signal light that has passed through the first and second optical paths C1 and C2) is emitted from the collimator lens 41. Therefore, the diffraction grating 45 having the same size as a conventional spectroscope that does not align the polarization direction using the polarization separation element 42 or the like can be provided. Can be used. Further, since the signal light emitted from the optical path coupling element 44 is a substantially parallel light beam, only one condensing optical system 46 is required. Therefore, the enlargement of the spectroscope 40 can be prevented. Moreover, since the spectroscope 40 shown in this embodiment has the optical fiber 28 (fiber coupler 29b) as the incident end, it can be easily connected to a confocal microscope. That is, as described above, in a general confocal microscope, light detection is performed by a photodetector connected to the pinhole plate 26, but the spectroscope is obtained by making the light transmitted through the pinhole 26a enter the optical fiber 28. It is possible to easily introduce light into the optical fiber 28 using the optical fiber 28.

  In the above description, the case where one diffraction grating 45 is used is shown. However, a plurality of diffraction gratings having different wavelength resolutions are arranged on the rotating grating stage, and the rotating grating stage is rotated to couple the optical path. The diffraction grating on which the signal light emitted from the element 44 enters may be switchable. By rotating the diffraction grating stage on which the diffraction grating is mounted, the wavelength resolution of the diffraction grating can be switched, and the incident angle and the emission angle of light with respect to this diffraction grating can be changed. The wavelength region that can be detected by the device 47 can be changed.

  In the above description, the plane mirror 42b is provided in the second optical path C2, but it goes without saying that a plane mirror may be provided in the first optical path C1. In the above embodiment, the light dispersed by the diffraction grating 45 is collected by the condensing optical system 46 formed of a concave mirror and then received by the light receiver 47. However, instead of the light receiver 47, a specific light is received. You may provide the output slit which radiate | emits the light of this wavelength range.

  In the above description, the polarization separation element 42 is provided with the polarization beam splitter plate 42a which is a so-called planar polarization beam splitter. However, a prism type or wedge substrate type polarization beam splitter can also be used. It is.

DESCRIPTION OF SYMBOLS 1 Microscopic system 30 Microscope 32 Objective lens 40 Spectroscope 41 Collimating lens 42 Polarization separation element 42a Polarization beam splitter plate 42b Plane mirror 43 Polarization rotation element 44 Optical path coupling member 441 First beam expander 441a First reflection surface 441b Second Reflective surface 442 Second beam expander 442a Second reflective surface 442b Second reflective surface 45 Diffraction grating 46 Condensing optical system 47 Light receiver

Claims (6)

A collimating lens that converts the signal light into a substantially parallel light beam;
The substantially parallel light beam emitted from the collimating lens is divided into two optical paths, a first optical path and a second optical path, the light in the first optical path is a P-polarized component, and the light in the second optical path is S-polarized. A polarization separation element as a component;
A polarization rotation element that is disposed in the first optical path and uses the light of the P-polarized component in the first optical path as light of the S-polarized component;
An optical path coupling member that reduces the diameter of the substantially parallel light flux of each of the first optical path and the second optical path while maintaining a parallel state;
A spectroscopic element for dispersing the substantially parallel light beam emitted from the optical path coupling member;
A light receiver for detecting the spectroscopic light dispersed by the spectroscopic element;
Have a, a focusing optical system for focusing the partial beams from the spectral element to the light receiver,
The optical path coupling member is
A first beam expander composed of a first reflecting surface and a second reflecting surface having an off-axis paraboloid shape that reflects the substantially parallel light beam emitted from the first optical path;
And a second beam expander configured by an off-axis paraboloid-shaped first reflecting surface and a second reflecting surface that reflects the substantially parallel light beam emitted from the second optical path. Spectrometer characterized by.   The spectroscope according to claim 1, wherein the optical path coupling member is configured to emit the substantially parallel light beams with reduced light beam diameters adjacent to each other. The optical path coupling member is configured such that an overall diameter of each of the substantially parallel light beams emitted from the optical path coupling member is equal to or less than a diameter of the substantially parallel light beams emitted from the collimator lens and incident on the polarization separation element. The spectroscope according to claim 1 or 2 , characterized in that: The polarization separation element is disposed at a position where the substantially parallel light beam emitted from the collimating lens is incident, transmits light of the P-polarized component of the substantially parallel light beam, guides it to the first optical path, and A polarizing beam splitter that reflects light;
According to any one of claims 1 to 3, characterized in that it is composed of a plane mirror the reflected and reflecting light of S polarized component guided to the second optical path by the polarization beam splitter Spectroscope. The spectroscope according to claim 1, wherein the polarization rotation element is a Fresnel ROM half-wave plate. A microscope that scans illumination light emitted from a light source and irradiates the specimen with an objective lens, and collects signal light emitted from the specimen with the objective lens, and
A spectroscopic light system comprising: the spectroscope according to any one of claims 1 to 5 that spectrally detects the signal light from the microscope.

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